Publications by authors named "Vincent P Klink"

36 Publications

Xyloglucan endotransglycosylase/hydrolase increases tightly-bound xyloglucan and chain number but decreases chain length contributing to the defense response that Glycine max has to Heterodera glycines.

PLoS One 2021 14;16(1):e0244305. Epub 2021 Jan 14.

Department of Biological Sciences, Mississippi State University, Starkville, Mississippi State, United States of America.

The Glycine max xyloglucan endotransglycosylase/hydrolase (EC 2.4.1.207), GmXTH43, has been identified through RNA sequencing of RNA isolated through laser microdissection of Heterodera glycines-parasitized root cells (syncytia) undergoing the process of defense. Experiments reveal that genetically increasing XTH43 transcript abundance in the H. glycines-susceptible genotype G. max[Williams 82/PI 518671] decreases parasitism. Experiments presented here show decreasing XTH43 transcript abundance through RNA interference (RNAi) in the H. glycines-resistant G. max[Peking/PI 548402] increases susceptibility, but it is unclear what role XTH43 performs. The experiments presented here show XTH43 overexpression decreases the relative length of xyloglucan (XyG) chains, however, there is an increase in the amount of those shorter chains. In contrast, XTH43 RNAi increases XyG chain length. The experiments show that XTH43 has the capability to function, when increased in its expression, to limit XyG chain extension. This outcome would likely impair the ability of the cell wall to expand. Consequently, XTH43 could provide an enzymatically-driven capability to the cell that would allow it to limit the ability of parasitic nematodes like H. glycines to develop a feeding structure that, otherwise, would facilitate parasitism. The experiments presented here provide experimentally-based proof that XTHs can function in ways that could be viewed as being able to limit the expansion of the cell wall.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0244305PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7808671PMC
January 2021

The Conserved Oligomeric Golgi (COG) Complex Functions During a Defense Response to .

Front Plant Sci 2020 11;11:564495. Epub 2020 Nov 11.

Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Starkville, MS, United States.

The conserved oligomeric Golgi (COG) complex, functioning in retrograde trafficking, is a universal structure present among eukaryotes that maintains the correct Golgi structure and function. The COG complex is composed of eight subunits coalescing into two sub-complexes. COGs1-4 compose Sub-complex A. COGs5-8 compose Sub-complex B. The observation that COG interacts with the syntaxins, suppressors of the erd2-deletion 5 (Sed5p), is noteworthy because Sed5p also interacts with Sec17p [alpha soluble NSF attachment protein (α-SNAP)]. The α-SNAP gene is located within the major [soybean cyst nematode (SCN)] resistance locus () and functions in resistance. The study presented here provides a functional analysis of the COG complex. The analysis has identified two paralogs of each COG gene. Functional transgenic studies demonstrate at least one paralog of each COG gene family functions in during resistance. Furthermore, treatment of with the bacterial effector harpin, known to function in effector triggered immunity (ETI), leads to the induced transcription of at least one member of each COG gene family that has a role in resistance. In some instances, altered COG gene expression changes the relative transcript abundance of syntaxin 31. These results indicate that the COG complex functions through processes involving ETI leading to resistance.
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http://dx.doi.org/10.3389/fpls.2020.564495DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686354PMC
November 2020

Mitogen activated protein kinase (MAPK)-regulated genes with predicted signal peptides function in the Glycine max defense response to the root pathogenic nematode Heterodera glycines.

PLoS One 2020 4;15(11):e0241678. Epub 2020 Nov 4.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS, United States of America.

Glycine max has 32 mitogen activated protein kinases (MAPKs), nine of them exhibiting defense functions (defense MAPKs) to the plant parasitic nematode Heterodera glycines. RNA seq analyses of transgenic G. max lines overexpressing (OE) each defense MAPK has led to the identification of 309 genes that are increased in their relative transcript abundance by all 9 defense MAPKs. Here, 71 of those genes are shown to also have measurable amounts of transcript in H. glycines-induced nurse cells (syncytia) produced in the root that are undergoing a defense response. The 71 genes have been grouped into 7 types, based on their expression profile. Among the 71 genes are 8 putatively-secreted proteins that include a galactose mutarotase-like protein, pollen Ole e 1 allergen and extensin protein, endomembrane protein 70 protein, O-glycosyl hydrolase 17 protein, glycosyl hydrolase 32 protein, FASCICLIN-like arabinogalactan protein 17 precursor, secreted peroxidase and a pathogenesis-related thaumatin protein. Functional transgenic analyses of all 8 of these candidate defense genes that employ their overexpression and RNA interference (RNAi) demonstrate they have a role in defense. Overexpression experiments that increase the relative transcript abundance of the candidate defense gene reduces the ability that the plant parasitic nematode Heterodera glycines has in completing its life cycle while, in contrast, RNAi of these genes leads to an increase in parasitism. The results provide a genomic analysis of the importance of MAPK signaling in relation to the secretion apparatus during the defense process defense in the G. max-H. glycines pathosystem and identify additional targets for future studies.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0241678PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641413PMC
January 2021

Exocyst components promote an incompatible interaction between Glycine max (soybean) and Heterodera glycines (the soybean cyst nematode).

Sci Rep 2020 09 14;10(1):15003. Epub 2020 Sep 14.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS, 39762, USA.

Vesicle and target membrane fusion involves tethering, docking and fusion. The GTPase SECRETORY4 (SEC4) positions the exocyst complex during vesicle membrane tethering, facilitating docking and fusion. Glycine max (soybean) Sec4 functions in the root during its defense against the parasitic nematode Heterodera glycines as it attempts to develop a multinucleate nurse cell (syncytium) serving to nourish the nematode over its 30-day life cycle. Results indicate that other tethering proteins are also important for defense. The G. max exocyst is encoded by 61 genes: 5 EXOC1 (Sec3), 2 EXOC2 (Sec5), 5 EXOC3 (Sec6), 2 EXOC4 (Sec8), 2 EXOC5 (Sec10) 6 EXOC6 (Sec15), 31 EXOC7 (Exo70) and 8 EXOC8 (Exo84) genes. At least one member of each gene family is expressed within the syncytium during the defense response. Syncytium-expressed exocyst genes function in defense while some are under transcriptional regulation by mitogen-activated protein kinases (MAPKs). The exocyst component EXOC7-H4-1 is not expressed within the syncytium but functions in defense and is under MAPK regulation. The tethering stage of vesicle transport has been demonstrated to play an important role in defense in the G. max-H. glycines pathosystem, with some of the spatially and temporally regulated exocyst components under transcriptional control by MAPKs.
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http://dx.doi.org/10.1038/s41598-020-72126-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7490361PMC
September 2020

The heterologous expression of a soybean (Glycine max) xyloglucan endotransglycosylase/hydrolase (XTH) in cotton (Gossypium hirsutum) suppresses parasitism by the root knot nematode Meloidogyne incognita.

PLoS One 2020 6;15(7):e0235344. Epub 2020 Jul 6.

Department of Biological Sciences, Mississippi State University, Mississippi State, Mississippi, United States of America.

A Glycine max (soybean) hemicellulose modifying gene, xyloglucan endotransglycoslase/hydrolase (XTH43), has been identified as being expressed within a nurse cell known as a syncytium developing within the soybean root undergoing the process of defense to infection by the parasitic nematode, Heterodera glycines. The highly effective nature of XTH43 overexpression in suppressing H. glycines parasitism in soybean has led to experiments examining whether the heterologous expression of XTH43 in Gossypium hirsutum (upland cotton) could impair the parasitism of Meloidogyne incognita, that form a different type of nurse cell called a giant cell that is enclosed within a swollen root structure called a gall. The heterologous transgenic expression of XTH43 in cotton resulted in an 18% decrease in the number of galls, 70% decrease in egg masses, 64% decrease in egg production and a 97% decrease in second stage juvenile (J2) production as compared to transgenic controls. The heterologous XTH43 expression does not significantly affect root mass. The results demonstrate XTH43 expression functions effectively in impairing the development of M. incognita at numerous life cycle stages occurring within the cotton root. The experiments reveal that there are highly conserved aspects of the defense response of G. max that can function effectively in G. hirsutum to impair M. incognita having a different method of parasitism.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0235344PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7337317PMC
September 2020

MAPKDB: A MAP kinase database for signal transduction element identification.

Bioinformation 2019 15;15(5):338-341. Epub 2018 May 15.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, USA.

The mitogen activated protein kinase (MAPK) cascade is a central signal transduction platform, ubiquitous within the eukaryotes. MAPKs function prominently in different essential cellular processes such as proliferation, differentiation, survival and defense to pathogen attack. The 32 MAPKs of Glycine max (soybean) have been examined functionally to determine if they have any defense role, focusing in on infection by the plant-parasitic nematode Heterodera glycines. Of these 32 MAPKs, 9 have been shown to have a defense function. Hence, the Mitogen Activated Protein Kinase database (MAPKDB) has been developed to assist in such research. The MAPKDB allows users to search the annotations with sequence data for G. max transgenic lines undergoing overexpression (OE) or RNA interference (RNAi) of its defense map kinases. These defense MAPKs include map kinase 2 (MPK2), MPK3, MPK4, MPK5, MPK6, MPK13, MPK16, and MPK20. The database also contains data analysis information for each sample that helps to detect the differential expression of the genes identified within these samples. The database also contains data for each sample that helps to detect the differential expression of the genes identified within these samples. The database has been developed to manage G. max MAPK sequences with sequence alignment for 18 different samples along with two additional OE and RNAi control experiments for a total of 20.
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http://dx.doi.org/10.6026/97320630015338DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6589469PMC
May 2018

The mitogen activated protein kinase (MAPK) gene family functions as a cohort during the Glycine max defense response to Heterodera glycines.

Plant Physiol Biochem 2019 Apr 28;137:25-41. Epub 2019 Jan 28.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS, 39762, USA. Electronic address:

Mitogen activated protein kinases (MAPKs) play important signal transduction roles. However, little is known regarding how they influence the gene expression of other family members and the relationship to a biological process, including the Glycine max defense response to Heterodera glycines. Transcriptomics have identified MAPK gene expression occurring within root cells undergoing a defense response to a pathogenic event initiated by H. glycines in the allotetraploid Glycine max. Functional analyses are presented for its 32 MAPKs revealing 9 have a defense role, including homologs of Arabidopsis thaliana MAPK (MPK) MPK2, MPK3, MPK4, MPK5, MPK6, MPK13, MPK16 and MPK20. Defense signaling occurring through pathogen activated molecular pattern (PAMP) triggered immunity (PTI) and effector triggered immunity (ETI) have been determined in relation to these MAPKs. Five different types of gene expression relate to MAPK expression, influencing PTI and ETI gene expression and proven defense genes including an ABC-G transporter, 20S membrane fusion particle components, glycoside biosynthesis, carbon metabolism, hemicellulose modification, transcription and secretion. The experiments show MAPKs broadly influence defense MAPK gene expression, including the co-regulation of parologous MAPKs and reveal its relationship to proven defense genes. The experiments reveal each defense MAPK induces the expression of a G. max homolog of a PATHOGENESIS RELATED1 (PR1), itself shown to function in defense in the studied pathosystem.
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http://dx.doi.org/10.1016/j.plaphy.2019.01.018DOI Listing
April 2019

Testing the AC/DC hypothesis: Rock and roll is noise pollution and weakens a trophic cascade.

Ecol Evol 2018 Aug 10;8(15):7649-7656. Epub 2018 Jul 10.

Department of Biological Sciences Mississippi State University Mississippi State Mississippi.

Anthropogenic sound is increasingly considered a major environmental issue, but its effects are relatively unstudied. Organisms may be directly affected by anthropogenic sound in many ways, including interference with their ability to detect mates, predators, or food, and disturbances that directly affect one organism may in turn have indirect effects on others. Thus, to fully appreciate the net effect of anthropogenic sound, it may be important to consider both direct and indirect effects. We report here on a series of experiments to test the hypothesis that anthropogenic sound can generate cascading indirect effects within a community. We used a study system of lady beetles, soybean aphids, and soybean plants, which are a useful model for studying the direct and indirect effects of global change on food webs. For sound treatments, we used several types of music, as well as a mix of urban sounds (e.g., sirens, vehicles, and construction equipment), each at volumes comparable to a busy city street or farm tractor. In 18-hr feeding trials, rock music and urban sounds caused lady beetles to consume fewer aphids, but other types of music had no effect even at the same volume. We then tested the effect of rock music on the strength of trophic cascades in a 2-week experiment in plant growth chambers. When exposed to music by AC/DC, who articulated the null hypothesis that "rock and roll ain't noise pollution" in a song of the same name, lady beetles were less effective predators, resulting in higher aphid density and reduced final plant biomass relative to control (no music) treatments. While it is unclear what characteristics of sound generate these effects, our results reject the AC/DC hypothesis and demonstrate that altered interspecific interactions can transmit the indirect effects of anthropogenic noise through a community.
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http://dx.doi.org/10.1002/ece3.4273DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6106185PMC
August 2018

Harpin-inducible defense signaling components impair infection by the ascomycete Macrophomina phaseolina.

Plant Physiol Biochem 2018 Aug 18;129:331-348. Epub 2018 Jun 18.

Department of Biological Sciences, College of Arts and Sciences, Mississippi State University, Mississippi State, MS, 39762, USA. Electronic address:

Soybean (Glycine max) infection by the charcoal rot (CR) ascomycete Macrophomina phaseolina is enhanced by the soybean cyst nematode (SCN) Heterodera glycines. We hypothesized that G. max genetic lines impairing infection by M. phaseolina would also limit H. glycines parasitism, leading to resistance. As a part of this M. phaseolina resistance process, the genetic line would express defense genes already proven to impair nematode parasitism. Using G. max, exhibiting partial resistance to M. phaseolina, experiments show the genetic line also impairs H. glycines parasitism. Furthermore, comparative studies show G. max exhibits induced expression of the effector triggered immunity (ETI) gene NON-RACE SPECIFIC DISEASE RESISTANCE 1/HARPIN INDUCED1 (NDR1/HIN1) that functions in defense to H. glycines as compared to the H. glycines and M. phaseolina susceptible line G. max. Other defense genes that are induced in G. max include the pathogen associated molecular pattern (PAMP) triggered immunity (PTI) genes ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1), NONEXPRESSOR OF PR1 (NPR1) and TGA2. These observations link G. max defense processes that impede H. glycines parasitism to also potentially function toward impairing M. phaseolina pathogenicity. Testing this hypothesis, G. max genetically engineered to experimentally induce GmNDR1-1, EDS1-2, NPR1-2 and TGA2-1 expression leads to impaired M. phaseolina pathogenicity. In contrast, G. max engineered to experimentally suppress the expression of GmNDR1-1, EDS1-2, NPR1-2 and TGA2-1 by RNA interference (RNAi) enhances M. phaseolina pathogenicity. The results show components of PTI and ETI impair both nematode and M. phaseolina pathogenicity.
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http://dx.doi.org/10.1016/j.plaphy.2018.06.020DOI Listing
August 2018

A harpin elicitor induces the expression of a coiled-coil nucleotide binding leucine rich repeat (CC-NB-LRR) defense signaling gene and others functioning during defense to parasitic nematodes.

Plant Physiol Biochem 2017 Dec 13;121:161-175. Epub 2017 Oct 13.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, United States. Electronic address:

The bacterial effector harpin induces the transcription of the Arabidopsis thaliana NON-RACE SPECIFIC DISEASE RESISTANCE 1/HARPIN INDUCED1 (NDR1/HIN1) coiled-coil nucleotide binding leucine rich repeat (CC-NB-LRR) defense signaling gene. In Glycine max, Gm-NDR1-1 transcripts have been detected within root cells undergoing a natural resistant reaction to parasitism by the syncytium-forming nematode Heterodera glycines, functioning in the defense response. Expressing Gm-NDR1-1 in Gossypium hirsutum leads to resistance to Meloidogyne incognita parasitism. In experiments presented here, the heterologous expression of Gm-NDR1-1 in G. hirsutum impairs Rotylenchulus reniformis parasitism. These results are consistent with the hypothesis that Gm-NDR1-1 expression functions broadly in generating a defense response. To examine a possible relationship with harpin, G. max plants topically treated with harpin result in induction of the transcription of Gm-NDR1-1. The result indicates the topical treatment of plants with harpin, itself, may lead to impaired nematode parasitism. Topical harpin treatments are shown to impair G. max parasitism by H. glycines, M. incognita and R. reniformis and G. hirsutum parasitism by M. incognita and R. reniformis. How harpin could function in defense has been examined in experiments showing it also induces transcription of G. max homologs of the proven defense genes ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1), TGA2, galactinol synthase, reticuline oxidase, xyloglucan endotransglycosylase/hydrolase, alpha soluble N-ethylmaleimide-sensitive fusion protein (α-SNAP) and serine hydroxymethyltransferase (SHMT). In contrast, other defense genes are not directly transcriptionally activated by harpin. The results indicate harpin induces pathogen associated molecular pattern (PAMP) triggered immunity (PTI) and effector-triggered immunity (ETI) defense processes in the root, activating defense to parasitic nematodes.
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http://dx.doi.org/10.1016/j.plaphy.2017.10.004DOI Listing
December 2017

A Glycine max homolog of NON-RACE SPECIFIC DISEASE RESISTANCE 1 (NDR1) alters defense gene expression while functioning during a resistance response to different root pathogens in different genetic backgrounds.

Plant Physiol Biochem 2017 May 1;114:60-71. Epub 2017 Mar 1.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, United States. Electronic address:

A Glycine max homolog of the Arabidopsis thaliana NON-RACE SPECIFIC DISEASE RESISTANCE 1 (NDR1) coiled-coil nucleotide binding leucine rich repeat (CC-NB-LRR) defense signaling gene (Gm-NDR1-1) is expressed in root cells undergoing a defense response to the root pathogenic nematode, Heterodera glycines. Gm-NDR1-1 overexpression in the H. glycines-susceptible genotype G. max impairs parasitism. In contrast, Gm-NDR1-1 RNA interference (RNAi) in the H. glycines-resistant genotype G. max facilitates parasitism. The broad effectiveness of Gm-NDR1-1 in impairing parasitism has then been examined by engineering its heterologous expression in Gossypium hirsutum which is susceptible to the root pathogenic nematode Meloidogyne incognita. The heterologous expression of Gm-NDR1-1 in G. hirsutum effectively impairs M. incognita parasitism, reducing gall, egg mass, egg and juvenile numbers. In contrast to our prior experiments examining the effectiveness of the heterologous expression of a G. max homolog of the A. thaliana salicyclic acid signaling (SA) gene NONEXPRESSOR OF PR1 (Gm-NPR1-2), no cumulative negative effect on M. incognita parasitism has been observed in G. hirsutum expressing Gm-NDR1-1. The results indicate a common genetic basis exists for plant resistance to parasitic nematodes that involves Gm-NDR1. However, the Gm-NDR1-1 functions in ways that are measurably dissimilar to Gm-NPR1-2. Notably, Gm-NDR1-1 overexpression leads to increased relative transcript levels of its homologs of A. thaliana genes functioning in SA signaling, including NPR1-2, TGA2-1 and LESION SIMULATING DISEASE1 (LSD1-2) that is lost in Gm-NDR1-1 RNAi lines. Similar observations have been made regarding the expression of other defense genes.
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http://dx.doi.org/10.1016/j.plaphy.2017.02.022DOI Listing
May 2017

Components of the SNARE-containing regulon are co-regulated in root cells undergoing defense.

Plant Signal Behav 2017 02;12(2):e1274481

b Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology , Mississippi State University , Mississippi State , MS , USA.

The term regulon has been coined in the genetic model plant Arabidopsis thaliana, denoting a structural and physiological defense apparatus defined genetically through the identification of the penetration (pen) mutants. The regulon is composed partially by the soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor (SNARE) syntaxin PEN1. PEN1 has homology to a Saccharomyces cerevisae gene that regulates a Secretion (Sec) protein, Suppressor of Sec 1 (Sso1p). The regulon is also composed of the β-glucosidase (PEN2) and an ATP binding cassette (ABC) transporter (PEN3). While important in inhibiting pathogen infection, limited observations have been made regarding the transcriptional regulation of regulon genes until now. Experiments made using the model agricultural Glycine max (soybean) have identified co-regulated gene expression of regulon components. The results explain the observation of hundreds of genes expressed specifically in the root cells undergoing the natural process of defense. Data regarding additional G. max genes functioning within the context of the regulon are presented here, including Sec 14, Sec 4 and Sec 23. Other examined G. max homologs of membrane fusion genes include an endosomal bromo domain-containing protein1 (Bro1), syntaxin6 (SYP6), SYP131, SYP71, SYP8, Bet1, coatomer epsilon (ϵ-COP), a coatomer zeta (ζ-COP) paralog and an ER to Golgi component (ERGIC) protein. Furthermore, the effectiveness of biochemical pathways that would function within the context of the regulon ave been examined, including xyloglucan xylosyltransferase (XXT), reticuline oxidase (RO) and galactinol synthase (GS). The experiments have unveiled the importance of the regulon during defense in the root and show how the deposition of callose relates to the process.
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http://dx.doi.org/10.1080/15592324.2016.1274481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5351740PMC
February 2017

The syntaxin 31-induced gene, LESION SIMULATING DISEASE1 (LSD1), functions in Glycine max defense to the root parasite Heterodera glycines.

Plant Signal Behav 2015 ;10(1):e977737

a Department of Biological Sciences ; Mississippi State University ; Starkville , MS USA.

Experiments show the membrane fusion genes α soluble NSF attachment protein (α-SNAP) and syntaxin 31 (Gm-SYP38) contribute to the ability of Glycine max to defend itself from infection by the plant parasitic nematode Heterodera glycines. Accompanying their expression is the transcriptional activation of the defense genes ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1) and NONEXPRESSOR OF PR1 (NPR1) that function in salicylic acid (SA) signaling. These results implicate the added involvement of the antiapoptotic, environmental response gene LESION SIMULATING DISEASE1 (LSD1) in defense. Roots engineered to overexpress the G. max defense genes Gm-α-SNAP, SYP38, EDS1, NPR1, BOTRYTIS INDUCED KINASE1 (BIK1) and xyloglucan endotransglycosylase/hydrolase (XTH) in the susceptible genotype G. max[Williams 82/PI 518671] have induced Gm-LSD1 (Gm-LSD1-2) transcriptional activity. In reciprocal experiments, roots engineered to overexpress Gm-LSD1-2 in the susceptible genotype G. max[Williams 82/PI 518671] have induced levels of SYP38, EDS1, NPR1, BIK1 and XTH, but not α-SNAP prior to infection. In tests examining the role of Gm-LSD1-2 in defense, its overexpression results in ∼52 to 68% reduction in nematode parasitism. In contrast, RNA interference (RNAi) of Gm-LSD1-2 in the resistant genotype G. max[Peking/PI 548402] results in an 3.24-10.42 fold increased ability of H. glycines to parasitize. The results identify that Gm-LSD1-2 functions in the defense response of G. max to H. glycines parasitism. It is proposed that LSD1, as an antiapoptotic protein, may establish an environment whereby the protected, living plant cell could secrete materials in the vicinity of the parasitizing nematode to disarm it. After the targeted incapacitation of the nematode the parasitized cell succumbs to its targeted demise as the infected root region is becoming fortified.
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http://dx.doi.org/10.4161/15592324.2014.977737DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4622666PMC
October 2015

Quantitative field testing Heterodera glycines from metagenomic DNA samples isolated directly from soil under agronomic production.

PLoS One 2014 24;9(2):e89887. Epub 2014 Feb 24.

Department of Biological Sciences, Mississippi State University, Mississippi State, Mississippi, United States of America.

A quantitative PCR procedure targeting the Heterodera glycines ortholog of the Caenorhabditis elegans uncoordinated-78 gene was developed. The procedure estimated the quantity of H. glycines from metagenomic DNA samples isolated directly from field soil under agronomic production. The estimation of H. glycines quantity was determined in soil samples having other soil dwelling plant parasitic nematodes including Hoplolaimus, predatory nematodes including Mononchus, free-living nematodes and biomass. The methodology provides a framework for molecular diagnostics of nematodes from metagenomic DNA isolated directly from field soil.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0089887PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3933691PMC
January 2015

Syntaxin 31 functions in Glycine max resistance to the plant parasitic nematode Heterodera glycines.

Plant Mol Biol 2014 May 23;85(1-2):107-21. Epub 2014 Jan 23.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS, 39762, USA,

A Glycine max syntaxin 31 homolog (Gm-SYP38) was identified as being expressed in nematode-induced feeding structures known as syncytia undergoing an incompatible interaction with the plant parasitic nematode Heterodera glycines. The observed Gm-SYP38 expression was consistent with prior gene expression analyses that identified the alpha soluble NSF attachment protein (Gm-α-SNAP) resistance gene because homologs of these genes physically interact and function together in other genetic systems. Syntaxin 31 is a protein that resides on the cis face of the Golgi apparatus and binds α-SNAP-like proteins, but has no known role in resistance. Experiments presented here show Gm-α-SNAP overexpression induces Gm-SYP38 transcription. Overexpression of Gm-SYP38 rescues G. max [Williams 82/PI 518671], genetically rhg1 (-/-), by suppressing H. glycines parasitism. In contrast, Gm-SYP38 RNAi in the rhg1 (+/+) genotype G. max [Peking/PI 548402] increases susceptibility. Gm-α-SNAP and Gm-SYP38 overexpression induce the transcriptional activity of the cytoplasmic receptor-like kinase BOTRYTIS INDUCED KINASE 1 (Gm-BIK1-6) which is a family of defense proteins known to anchor to membranes through a 5' MGXXXS/T(R) N-myristoylation sequence. Gm-BIK1-6 had been identified previously by RNA-seq experiments as expressed in syncytia undergoing an incompatible reaction. Gm-BIK1-6 overexpression rescues the resistant phenotype. In contrast, Gm-BIK1-6 RNAi increases parasitism. The analysis demonstrates a role for syntaxin 31-like genes in resistance that until now was not known.
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http://dx.doi.org/10.1007/s11103-014-0172-2DOI Listing
May 2014

A novel sample preparation method that enables nucleic acid analysis from ultrathin sections.

Microsc Microanal 2013 Jun 21;19(3):635-41. Epub 2013 Mar 21.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, USA.

The ability to isolate and perform nucleic acid analyses of individual cells is critical to studying the development of various cell types and structures. We present a novel biological sample preparation method developed for laser capture microdissection-assisted nucleic acid analysis of ultrathin cell/tissue sections. We used cells of the mitotic bed of the tadpole teeth of Lithobates sphenocephalus (Southern Leopard Frog). Cells from the mitotic beds at the base of the developing teeth series were isolated and embedded in the methacrylate resin, Technovit® 9100®. Intact cells of the mitotic beds were thin sectioned and examined by bright-field and transmission electron microscopy. The cytological and ultrastructural anatomy of the immature and progressively more mature tooth primordia appeared well preserved and intact. A developmental series of tooth primordia were isolated by laser capture microdissection (LCM). Processing of these cells for RNA showed that intact RNA could be isolated. The study demonstrates that Technovit® 9100® can be used as an embedding medium for extremely small tissues and from individual cells, a prerequisite step to LCM and nucleic acid analyses. A relatively small amount of sample material was needed for the analysis, which makes this technique ideal for cell-specific analyses when the desired cells are limited in quantity.
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http://dx.doi.org/10.1017/S1431927613000044DOI Listing
June 2013

The expression of a naturally occurring, truncated allele of an α-SNAP gene suppresses plant parasitic nematode infection.

Plant Mol Biol 2012 Sep 12;80(2):131-55. Epub 2012 Jun 12.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, USA.

Transcriptional mapping experiments of the major soybean cyst nematode resistance locus, rhg1, identified expression of the vesicular transport machinery component, α soluble NSF attachment protein (α-SNAP), occurring during defense. Sequencing the α-SNAP coding regions from the resistant genotypes G. max ([Peking/PI 548402]) and G. max ([PI 437654]) revealed they are identical, but differ from the susceptible G. max ([Williams 82/PI 518671]) by the presence of several single nucleotide polymorphisms. Using G. max ([Williams 82/PI 518671]) as a reference, a G → T(2,822) transversion in the genomic DNA sequence at a functional splice site of the α-SNAP([Peking/PI 548402]) allele produced an additional 17 nucleotides of mRNA sequence that contains an in-frame stop codon caused by a downstream G → A(2,832) transition. The G. max ([Peking/PI 548402]) genotype has cell wall appositions (CWAs), structures identified as forming as part of a defense response by the activity of the vesicular transport machinery. In contrast, the 17 nt α-SNAP([Peking/PI 548402]) mRNA motif is not found in G. max ([PI 88788]) that exhibits defense to H. glycines, but lack CWAs. The α-SNAP([PI 88788]) promoter contains sequence elements that are nearly identical to the α-SNAP([Peking/PI 548402]) allele, but differs from the G. max ([Williams 82/PI 518671]) ortholog. Overexpressing the α-SNAP([Peking/PI 548402]) allele in the susceptible G. max ([Williams 82/PI 518671]) genotype suppressed H. glycines infection. The experiments indicate a role for the vesicular transport machinery during infection of soybean by the soybean cyst nematode. However, increased GmEREBP1, PR1, PR2, PR5 gene activity but suppressed PR3 expression accompanied the overexpression of the α-SNAP([Peking/PI 548402]) allele prior to infection.
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http://dx.doi.org/10.1007/s11103-012-9932-zDOI Listing
September 2012

Quantitative field testing Rotylenchulus reniformis DNA from metagenomic samples isolated directly from soil.

PLoS One 2011 16;6(12):e28954. Epub 2011 Dec 16.

Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, Mississippi, United States of America.

A quantitative PCR procedure targeting the β-tubulin gene determined the number of Rotylenchulus reniformis Linford & Oliveira 1940 in metagenomic DNA samples isolated from soil. Of note, this outcome was in the presence of other soil-dwelling plant parasitic nematodes including its sister genus Helicotylenchus Steiner, 1945. The methodology provides a framework for molecular diagnostics of nematodes from metagenomic DNA isolated directly from soil.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0028954PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3241691PMC
August 2012

Mapping cell fate decisions that occur during soybean defense responses.

Plant Mol Biol 2011 Nov 11;77(4-5):513-28. Epub 2011 Oct 11.

Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, USA.

The soybean defense response to the soybean cyst nematode was used as a model to map at cellular resolution its genotype-defined cell fate decisions occurring during its resistant reactions. The defense responses occur at the site of infection, a nurse cell known as the syncytium. Two major genotype-defined defense responses exist, the G. max ([Peking])- and G. max ([PI 88788])-types. Resistance in G. max ([Peking]) is potent and rapid, accompanied by the formation of cell wall appositions (CWAs), structures known to perform important defense roles. In contrast, defense occurs by a potent but more prolonged reaction in G. max ([PI 88788]), lacking CWAs. Comparative transcriptomic analyses with confirmation by Illumina® deep sequencing were organized through a custom-developed application, Pathway Analysis and Integrated Coloring of Experiments (PAICE) that presents gene expression of these cytologically and developmentally distinct defense responses using the Kyoto Encyclopedia of Genes and Genomes (KEGG) framework. The analyses resulted in the generation of 1,643 PAICE pathways, allowing better understanding of gene activity across all chromosomes. Analyses of the rhg1 resistance locus, defined within a 67 kb region of DNA demonstrate expression of an amino acid transporter and an α soluble NSF attachment protein gene specifically in syncytia undergoing their defense responses.
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http://dx.doi.org/10.1007/s11103-011-9828-3DOI Listing
November 2011

Extremes in rapid cellular morphogenesis: post-transcriptional regulation of spermatogenesis in Marsilea vestita.

Protoplasma 2011 Jul 13;248(3):457-73. Epub 2011 Apr 13.

Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742, USA.

The endosporic male gametophyte of the water fern, Marsilea vestita, provides a unique opportunity to study the mechanisms that control cell fate determination during a burst of rapid development. In this review, we show how the spatial and temporal control of development in this simple gametophyte involves several distinct modes of RNA processing that allow the translation of specific mRNAs at distinct stages during gametogenesis. During the early part of development, nine successive cell division cycles occur in precise planes within a closed volume to produce seven sterile cells and 32 spermatids. There is no cell movement in the gametophyte; so, cell position and size within the spore wall define cell fate. After the division cycles have been completed, the spermatids become sites for the de novo formation of basal bodies, for the assembly of a complex cytoskeleton, for nuclear and cell elongation, and for ciliogenesis. In contrast, the adjacent sterile cells exhibit none of these changes. The spermatids differentiate into multiciliated, corkscrew-shaped gametes that resemble no other cells in the entire plant. Development is controlled post-transcriptionally. The transcripts stored in the microspore are released (unmasked) in the gametophyte at different times during development. At the start of these studies, we identified several key mRNAs that undergo translation at specific stages of gametophyte development. We developed RNA silencing protocols that enabled us to block the translation of these proteins and thereby establish their necessity and sufficiency for the completion of specific stages of gametogenesis. In addition, RNAi enabled us to identify additional proteins that are essential for other phases of development. Since the distributions of mRNAs and the proteins they encode are not identical in the gametophyte, transcript processing is apparently important in allowing translation to occur under strict temporal and spatial control. Transcript polyadenylation occurs in the spermatogenous cells in ways that match the translation of specific mRNAs. We have found that the exon junction complex plays key roles in transcript regulation and modifications that underlie cell specification in the gametophyte. We have recently become interested in the mechanisms that control the unmasking of the stored transcripts and have linked the synthesis and redistribution of spermidine in the gametophyte to the control of mRNA release from storage during early development and later to basal body formation, cytoskeletal assembly, and nuclear and cell elongation in the differentiating spermatids.
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http://dx.doi.org/10.1007/s00709-011-0276-3DOI Listing
July 2011

Differences in gene expression amplitude overlie a conserved transcriptomic program occurring between the rapid and potent localized resistant reaction at the syncytium of the Glycine max genotype Peking (PI 548402) as compared to the prolonged and potent resistant reaction of PI 88788.

Plant Mol Biol 2011 Jan 14;75(1-2):141-65. Epub 2010 Dec 14.

Department of Biological Sciences, Harned Hall, Mississippi State University, Mississippi State, MS 39762, USA.

Glycine max L. Merr. (soybean) resistance to Heterodera glycines Ichinohe occurs at the site of infection, a nurse cell known as the syncytium. Resistance is classified into two cytologically-defined responses, the G. max ([Peking])- and G. max ([PI 88788])-types. Each type represents a cohort of G. max genotypes. Resistance in G. max ([Peking]) occurs by a potent and rapid localized response, affecting parasitic second stage juveniles (p-J2). In contrast, resistance occurs by a potent but more prolonged reaction in the genotype G. max ([PI 88788]) that affects nematode development at the J3 and J4 stages. Microarray analyses comparing these cytologically and developmentally distinct resistant reactions reveal differences in gene expression in pericycle and surrounding cells even before infection. The differences include higher relative levels of the differentially expressed in response to arachidonic acid 1 gene (DEA1 [Gm-DEA1]) (+224.19-fold) and a protease inhibitor (+68.28-fold) in G. max ([Peking/PI 548402]) as compared to G. max ([PI 88788]). Gene pathway analyses compare the two genotypes (1) before, (2) at various times during, (3) constitutively throughout the resistant reaction and (4) at all time points prior to and during the resistant reaction. The amplified levels of transcriptional activity of defense genes may explain the rapid and potent reaction in G. max ([Peking/PI 548402]) as compared to G. max ([PI 88788]). In contrast, the shared differential expression levels of genes in G. max ([Peking/PI 548402]) and G. max ([PI 88788]) may indicate a conserved genomic program underlying the G. max resistance on which the genotype-specific gene expression programs are built off.
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http://dx.doi.org/10.1007/s11103-010-9715-3DOI Listing
January 2011

Microarray Detection Call Methodology as a Means to Identify and Compare Transcripts Expressed within Syncytial Cells from Soybean (Glycine max) Roots Undergoing Resistant and Susceptible Reactions to the Soybean Cyst Nematode (Heterodera glycines).

J Biomed Biotechnol 2010 19;2010:491217. Epub 2010 May 19.

Department of Biological Sciences, Harned Hall, Mississippi State University, Mississippi State, MS 39762, USA.

Background. A comparative microarray investigation was done using detection call methodology (DCM) and differential expression analyses. The goal was to identify genes found in specific cell populations that were eliminated by differential expression analysis due to the nature of differential expression methods. Laser capture microdissection (LCM) was used to isolate nearly homogeneous populations of plant root cells. Results. The analyses identified the presence of 13,291 transcripts between the 4 different sample types. The transcripts filtered down into a total of 6,267 that were detected as being present in one or more sample types. A comparative analysis of DCM and differential expression methods showed a group of genes that were not differentially expressed, but were expressed at detectable amounts within specific cell types. Conclusion. The DCM has identified patterns of gene expression not shown by differential expression analyses. DCM has identified genes that are possibly cell-type specific and/or involved in important aspects of plant nematode interactions during the resistance response, revealing the uniqueness of a particular cell population at a particular point during its differentiation process.
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http://dx.doi.org/10.1155/2010/491217DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2875038PMC
September 2012

Syncytium gene expression in Glycine max([PI 88788]) roots undergoing a resistant reaction to the parasitic nematode Heterodera glycines.

Plant Physiol Biochem 2010 Feb-Mar;48(2-3):176-93. Epub 2010 Jan 4.

Department of Biological Sciences, Harned Hall, Mississippi State University, Mississippi State, MS, 39762, USA.

The plant parasitic nematode, Heterodera glycines is the major pathogen of Glycine max (soybean). H. glycines accomplish parasitism by creating a nurse cell known as the syncytium from which it feeds. The syncytium undergoes two developmental phases. The first is a parasitism phase where feeding sites are selected, initiating the development of the syncytium. During this earlier phase (1-4 days post infection), syncytia undergoing resistant and susceptible reactions appear the same. The second phase is when the resistance response becomes evident (between 4 and 6dpi) and is completed by 9dpi. Analysis of the resistant reaction of G. max genotype PI 88788 (G. max([PI 88788])) to H. glycines population NL1-RHg/HG-type 7 (H. glycines([NL1-RHg/HG-type 7])) is accomplished by laser microdissection of syncytia at 3, 6 and 9dpi. Comparative analyses are made to pericycle and their neighboring cells isolated from mock-inoculated roots. These analyses reveal induced levels of the jasmonic acid biosynthesis and 13-lipoxygenase pathways. Direct comparative analyses were also made of syncytia at 6 days post infection to those at 3dpi (base line). The comparative analyses were done to identify localized gene expression that characterizes the resistance phase of the resistant reaction. The most highly induced pathways include components of jasmonic acid biosynthesis, 13-lipoxygenase pathway, S-adenosyl methionine pathway, phenylpropanoid biosynthesis, suberin biosynthesis, adenosylmethionine biosynthesis, ethylene biosynthesis from methionine, flavonoid biosynthesis and the methionine salvage pathway. In comparative analyses of 9dpi to 6dpi (base line), these pathways, along with coumarin biosynthesis, cellulose biosynthesis and homogalacturonan degradation are induced. The experiments presented here strongly implicate the jasmonic acid defense pathway as a factor involved in the localized resistant reaction of G. max([PI 88788]) to H. glycines([NL1-RHg/HG-type 7]).
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http://dx.doi.org/10.1016/j.plaphy.2009.12.003DOI Listing
September 2010

A gene expression analysis of syncytia laser microdissected from the roots of the Glycine max (soybean) genotype PI 548402 (Peking) undergoing a resistant reaction after infection by Heterodera glycines (soybean cyst nematode).

Plant Mol Biol 2009 Dec 29;71(6):525-67. Epub 2009 Sep 29.

Department of Biological Sciences, Mississippi State University, Harned Hall, Mississippi State, MS 39762, USA.

The syncytium is a nurse cell formed within the roots of Glycine max by the plant parasitic nematode Heterodera glycines. Its development and maintenance are essential for nematode survival. The syncytium appears to undergo two developmental phases during its maturation into a functional nurse cell. The first phase is a parasitism phase where the nematode establishes the molecular circuitry that during the second phase ensures a compatible interaction with the plant cell. The cytological features of syncytia undergoing susceptible or resistant reactions appear the same during the parasitism phase. Depending on the outcome of any defense response, the second phase is a period of syncytium maintenance (susceptible reaction) or failure (resistant reaction). In the analyses presented here, the localized gene expression occurring at the syncytium during the resistant reaction was studied. This was accomplished by isolating syncytial cells from Glycine max genotype Peking (PI 548402) by laser capture microdissection. Microarray analyses using the Affymetrix soybean GeneChip directly compared Peking syncytia undergoing a resistant reaction to those undergoing a susceptible reaction during the parasitism phase of the resistant reaction. Those analyses revealed lipoxygenase-9 and lipoxygenase-4 as the most highly induced genes in the resistant reaction. The analysis also identified induced levels of components of the phenylpropanoid pathway. These genes included phenylalanine ammonia lyase, chalcone isomerase, isoflavone reductase, cinnamoyl-CoA reductase and caffeic acid O-methyltransferase. The presence of induced levels of these genes implies the importance of jasmonic acid and phenylpropanoid signaling pathways locally at the site of the syncytium during the resistance phase of the resistant reaction. The analysis also identified highly induced levels of four S-adenosylmethionine synthetase genes, the EARLY-RESPONSIVE TO DEHYDRATION 2 gene and the 14-3-3 gene known as GENERAL REGULATORY FACTOR 2. Subsequent analyses studied microdissected syncytial cells at 3, 6 and 9 days post infection (dpi) during the course of the resistant reaction, resulting in the identification of signature gene expression profiles at each time point in a single G. max genotype, Peking.
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http://dx.doi.org/10.1007/s11103-009-9539-1DOI Listing
December 2009

Emerging approaches to broaden resistance of soybean to soybean cyst nematode as supported by gene expression studies.

Plant Physiol 2009 Nov 12;151(3):1017-22. Epub 2009 Aug 12.

Department of Biological Sciences, Mississippi State University, Mississippi State, Mississippi 39762, USA.

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http://dx.doi.org/10.1104/pp.109.144006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2773110PMC
November 2009

A correlation between host-mediated expression of parasite genes as tandem inverted repeats and abrogation of development of female Heterodera glycines cyst formation during infection of Glycine max.

Planta 2009 Jun 4;230(1):53-71. Epub 2009 Apr 4.

Department of Biological Sciences, Mississippi State University, Harned Hall, Rm 310, Mississippi State, MS 39762, USA.

Host-mediated (hm) expression of parasite genes as tandem inverted repeats was investigated as a means to abrogate the formation of mature Heterodera glycines (soybean cyst nematode) female cysts during its infection of Glycine max (soybean). A Gateway-compatible hm plant transformation system was developed specifically for these experiments in G. max. Three steps then were taken to identify H. glycines candidate genes. First, a pool of 150 highly conserved H. glycines homologs of genes having lethal mutant phenotypes or phenocopies from the free living nematode Caenorhabditis elegans were identified. Second, annotation of those 150 genes on the Affymetrix soybean GeneChip allowed for the identification of a subset of 131 genes whose expression could be monitored during the parasitic phase of the H. glycines life cycle. Third, a microarray analyses identified a core set of 32 genes with induced expression (>2.0-fold, log base 2) during the parasitic stages of infection. H. glycines homologs of small ribosomal protein 3a and 4 (Hg-rps-3a [accession number CB379877] and Hg-rps-4 [accession number CB278739]), synaptobrevin (Hg-snb-1 [accession number BF014436]) and a spliceosomal SR protein (Hg-spk-1 [accession number BI451523.1]) were tested for functionality in hm expression studies. Effects on H. glycines development were observed 8 days after infection. Experiments demonstrated that 81-93% fewer females developed on transgenic roots containing the genes engineered as tandem inverted repeats. The effect resembles RNA interference. The methodology has been used here as an alternative approach to engineer resistance to H. glycines.
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http://dx.doi.org/10.1007/s00425-009-0926-2DOI Listing
June 2009

Population-specific gene expression in the plant pathogenic nematode Heterodera glycines exists prior to infection and during the onset of a resistant or susceptible reaction in the roots of the Glycine max genotype Peking.

BMC Genomics 2009 Mar 16;10:111. Epub 2009 Mar 16.

Department of Biological Sciences, Harned Hall, Mississippi State University, Mississippi State, MS 39762, USA.

Background: A single Glycine max (soybean) genotype (Peking) reacts differently to two different populations of Heterodera glycines (soybean cyst nematode) within the first twelve hours of infection during resistant (R) and susceptible (S) reactions. This suggested that H. glycines has population-specific gene expression signatures. A microarray analysis of 7539 probe sets representing 7431 transcripts on the Affymetrix soybean GeneChip were used to identify population-specific gene expression signatures in pre-infective second stage larva (pi-L2) prior to their infection of Peking. Other analyses focused on the infective L2 at 12 hours post infection (i-L2(12h)), and the infective sedentary stages at 3 days post infection (i-L2(3d)) and 8 days post infection (i-L2/L3(8d)).

Results: Differential expression and false discovery rate (FDR) analyses comparing populations of pi-L2 (i.e., incompatible population, NL1-RHg to compatible population, TN8) identified 71 genes that were induced in NL1-RHg as compared to TN8. These genes included putative gland protein G23G12, putative esophageal gland protein Hgg-20 and arginine kinase. The comparative analysis of pi-L2 identified 44 genes that were suppressed in NL1-RHg as compared to TN8. These genes included a different Hgg-20 gene, an EXPB1 protein and a cuticular collagen. By 12 h, there were 7 induced genes and 0 suppressed genes in NL1-RHg. By 3d, there were 9 induced and 10 suppressed genes in NL1-RHg. Substantial changes in gene expression became evident subsequently. At 8d there were 13 induced genes in NL1-RHg. This included putative gland protein G20E03, ubiquitin extension protein, putative gland protein G30C02 and beta-1,4 endoglucanase. However, 1668 genes were found to be suppressed in NL1-RHg. These genes included steroid alpha reductase, serine proteinase and a collagen protein.

Conclusion: These analyses identify a genetic expression signature for these two populations both prior to and subsequently as they undergo an R or S reaction. The identification of genes like steroid alpha reductase and serine proteinase that are involved in feeding and nutritional uptake as being highly suppressed during the R response at 8d may indicate genes that the plant is targeting. The analyses also identified numerous putative parasitism genes that are differentially expressed. The 1668 genes that are suppressed in NL1-RHg, and hence induced in TN8 may represent genes that are important during the parasitic stages of H. glycines development. The potential for different arrays of putative parasitism genes to be expressed in different nematode populations may indicate how H. glycines evolve mechanisms to overcome resistance.
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http://dx.doi.org/10.1186/1471-2164-10-111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2662880PMC
March 2009

The use of laser capture microdissection to study the infection of Glycine max (soybean) by Heterodera glycines (soybean cyst nematode).

Plant Signal Behav 2008 Feb;3(2):105-7

United States Department of Agriculture; Soybean Genomics and Improvement Laboratory; Beltsville, Maryland USA.

The soybean cyst nematode (Heterodera glycines) is an obligate parasite of soybean (Glycine max). It is the most destructive pathogen of G. max, accounting for approximately 0.46-0.82 billion dollars in crop losses, annually, in the U.S. Part of the infection process involves H. glycines establishing feeding sites (syncytia) that it derives its nourishment from throughout its lifecycle. Microscopic methods (i.e., laser capture microdissection [LCM]) that faithfully dissect out those feeding sites are important improvements to the study of this significant plant pathogen. Our isolation of developing feeding sites during an incompatible or a compatible reaction is providing new ways by which this important plant-pathogen interaction can be studied. We have used these methods to create cDNA libraries, clone genes and perform microarray analyses. Importantly, it is providing insight not only into how the root is responding at the organ level to H. glycines, but also how the syncytium is responding during its maturation into a functional feeding site.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2633993PMC
http://dx.doi.org/10.4161/psb.3.2.4962DOI Listing
February 2008

Laser capture microdissection (LCM) and comparative microarray expression analysis of syncytial cells isolated from incompatible and compatible soybean (Glycine max) roots infected by the soybean cyst nematode (Heterodera glycines).

Planta 2007 Nov 1;226(6):1389-409. Epub 2007 Aug 1.

United States Department of Agriculture, Soybean Genomics and Improvement Laboratory, Bldg. 006, Rm. 118, 10300 Baltimore Ave., Beltsville, MD 20705-2350, USA.

Syncytial cells in soybean (Glycine max cultivar [cv.] Peking) roots infected by incompatible and compatible populations of soybean cyst nematode (SCN [Heterodera glycines]) were collected using laser capture microdissection (LCM). Gene transcript abundance was assayed using Affymetrix soybean GeneChips, each containing 37,744 probe sets. Our analyses identified differentially expressed genes in syncytial cells that are not differentially expressed in the whole root analyses. Therefore, our results show that the mass of transcriptional activity occurring in the whole root is obscuring identification of transcriptional events occurring within syncytial cells. In syncytial cells from incompatible roots at three dpi, genes encoding lipoxygenase (LOX), heat shock protein (HSP) 70, superoxidase dismutase (SOD) were elevated almost tenfold or more, while genes encoding several transcription factors and DNA binding proteins were also elevated, albeit at lower levels. In syncytial cells formed during the compatible interaction at three dpi, genes encoding prohibitin, the epsilon chain of ATP synthase, allene oxide cyclase and annexin were more abundant. By 8 days, several genes of unknown function and genes encoding a germin-like protein, peroxidase, LOX, GAPDH, 3-deoxy-D-arabino-heptolosonate 7-phosphate synthase, ATP synthase and a thioesterase were abundantly expressed. These observations suggest that gene expression is different in syncytial cells as compared to whole roots infected with nematodes. Our observations also show that gene expression is different between syncytial cells that were isolated from incompatible and compatible roots and that gene expression is changing over the course of syncytial cell development as it matures into a functional feeding site.
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http://dx.doi.org/10.1007/s00425-007-0578-zDOI Listing
November 2007

A time-course comparative microarray analysis of an incompatible and compatible response by Glycine max (soybean) to Heterodera glycines (soybean cyst nematode) infection.

Planta 2007 Nov 25;226(6):1423-47. Epub 2007 Jul 25.

United States Department of Agriculture, Soybean Genomics and Improvement Laboratory, 10300 Baltimore Ave. Bldg 006, Beltsville, MD 20705, USA.

The development of an infection in soybean [Glycine max L. cultivar (cv.) Peking] roots by incompatible (I) and compatible (C) populations of soybean cyst nematode (SCN) (Heterodera glycines) was assayed using an AffymetriX soybean GeneChip. This time-course microarray analysis, using 37,744 probe sets, measured transcript abundance during I and C. These analyses reveal that infection by individual I and C H. glycines populations influence the transcription of G. max genes differently. A substantial difference in gene expression is present between I and C at 12 h post infection. Thus, G. max can differentiate between I and C nematode populations even before they have begun to select their feeding sites. The microarray analysis identified genes induced earlier in infection during I than C. MA also identified amplitude differences in transcript abundance between I and C reactions. Some of the probe sets measuring increased transcript levels during I represented no apical meristem (NAM) and WRKY transcription factors as well as NBS-LRR kinases. Later during I, heat shock protein (HSPs) probe sets (i.e. HSP90, HSP70, ClpB/HSP101) measured increased transcript abundance. These results demonstrate that G. max roots respond very differently to the different H. glycines races even before their feeding site selection has occurred. The ability of G. max to engage an I reaction, thus, appears to be dependent on the ability of root cells to recognize the different races of H. glycines because these experiments were conducted in the identical G. max genetic background.
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http://dx.doi.org/10.1007/s00425-007-0581-4DOI Listing
November 2007