Publications by authors named "Jane Y Wu"

96 Publications

Tongue as a first-line immune organ?

Authors:
Jane Y Wu

Protein Cell 2021 Mar 9;12(3):162-164. Epub 2021 Jan 9.

Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, USA.

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http://dx.doi.org/10.1007/s13238-020-00802-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7796681PMC
March 2021

SLIT2/ROBO1-signaling inhibits macropinocytosis by opposing cortical cytoskeletal remodeling.

Nat Commun 2020 08 17;11(1):4112. Epub 2020 Aug 17.

Program in Cell Biology, The Hospital for Sick Children, Peter Gilgan Centre for Research and Learning, 686 Bay Street, Toronto, ON, M5G 0A4, Canada.

Macropinocytosis is essential for myeloid cells to survey their environment and for growth of RAS-transformed cancer cells. Several growth factors and inflammatory stimuli are known to induce macropinocytosis, but its endogenous inhibitors have remained elusive. Stimulation of Roundabout receptors by Slit ligands inhibits directional migration of many cell types, including immune cells and cancer cells. We report that SLIT2 inhibits macropinocytosis in vitro and in vivo by inducing cytoskeletal changes in macrophages. In mice, SLIT2 attenuates the uptake of muramyl dipeptide, thereby preventing NOD2-dependent activation of NF-κB and consequent secretion of pro-inflammatory chemokine, CXCL1. Conversely, blocking the action of endogenous SLIT2 enhances CXCL1 secretion. SLIT2 also inhibits macropinocytosis in RAS-transformed cancer cells, thereby decreasing their survival in nutrient-deficient conditions which resemble tumor microenvironment. Our results identify SLIT2 as a physiological inhibitor of macropinocytosis and challenge the conventional notion that signals that enhance macropinocytosis negatively regulate cell migration, and vice versa.
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http://dx.doi.org/10.1038/s41467-020-17651-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7431850PMC
August 2020

Explant Culture of the Embryonic Mouse Spinal Cord and Gene Transfer by Electroporation.

Bio Protoc 2019 Sep 20;9(18):e3373. Epub 2019 Sep 20.

Department of Neurology, Lurie Comprehensive Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, IL, USA.

Developing axons change responsiveness to guidance cues during the journey to synapse with target cells. Axon crossing at the ventral midline serves as a model for studying how axons accomplish such a switch in their response. Although primary neuron culture has been a versatile technique for elucidating various developmental mechanisms, many characteristics of neurons, such as long axon-extending abilities and axonal compartments, are not thoroughly preserved. In explant cultures, such properties of differentiated neurons and tissue architecture are maintained. To examine how the midline repellent Slit regulated the distribution of the Robo receptor in spinal cord commissural axons upon midline crossing and whether Robo trafficking machinery was a determinant of midline crossing, novel explant culture systems were developed. We have combined an "open-book" spinal cord explant method with that devised for flat-mount retinae. Here we present our protocol for explant culture of embryonic mouse spinal cords, which allows flexible manipulation of experimental conditions, immunostaining of extending axons and quantitative analysis of individual axons. In addition, we present a modified method that combines electroporation and "closed-book" spinal cord explant culture. These culture systems provide new platforms for detailed analysis of axon guidance, by adapting gene knockdown, knockout and genome editing.
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http://dx.doi.org/10.21769/BioProtoc.3373DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7854209PMC
September 2019

TDP-43 induces mitochondrial damage and activates the mitochondrial unfolded protein response.

PLoS Genet 2019 05 17;15(5):e1007947. Epub 2019 May 17.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Mutations in or dys-regulation of the TDP-43 gene have been associated with TDP-43 proteinopathy, a spectrum of neurodegenerative diseases including Frontotemporal Lobar Degeneration (FTLD) and Amyotrophic Lateral Sclerosis (ALS). The underlying molecular and cellular defects, however, remain unclear. Here, we report a systematic study combining analyses of patient brain samples with cellular and animal models for TDP-43 proteinopathy. Electron microscopy (EM) analyses of patient samples revealed prominent mitochondrial impairment, including abnormal cristae and a loss of cristae; these ultrastructural changes were consistently observed in both cellular and animal models of TDP-43 proteinopathy. In these models, increased TDP-43 expression induced mitochondrial dysfunction, including decreased mitochondrial membrane potential and elevated production of reactive oxygen species (ROS). TDP-43 expression suppressed mitochondrial complex I activity and reduced mitochondrial ATP synthesis. Importantly, TDP-43 activated the mitochondrial unfolded protein response (UPRmt) in both cellular and animal models. Down-regulating mitochondrial protease LonP1 increased mitochondrial TDP-43 levels and exacerbated TDP-43-induced mitochondrial damage as well as neurodegeneration. Together, our results demonstrate that TDP-43 induced mitochondrial impairment is a critical aspect in TDP-43 proteinopathy. Our work has not only uncovered a previously unknown role of LonP1 in regulating mitochondrial TDP-43 levels, but also advanced our understanding of the pathogenic mechanisms for TDP-43 proteinopathy. Our study suggests that blocking or reversing mitochondrial damage may provide a potential therapeutic approach to these devastating diseases.
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http://dx.doi.org/10.1371/journal.pgen.1007947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6524796PMC
May 2019

Reduced USP33 expression in gastric cancer decreases inhibitory effects of Slit2-Robo1 signalling on cell migration and EMT.

Cell Prolif 2019 May 21;52(3):e12606. Epub 2019 Mar 21.

Department of Gastric Surgery, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China.

Objectives: Gastric cancer (GC) is one of the most common cancers in the world, causing a large number of deaths every year. The Slit-Robo signalling pathway, initially discovered for its critical role in neuronal guidance, has recently been shown to modulate tumour invasion and metastasis in several human cancers. However, the role of Slit-Robo signalling and the molecular mechanisms underlying its role in the pathogenesis of gastric cancer remains to be elucidated.

Materials And Methods: Slit2, Robo1 and USP33 expressions were analysed in datasets obtained from the Oncomine database and measured in human gastric cancer specimens. The function of Slit2-Robo1-USP33 signalling on gastric cancer cells migration and epithelial-mesenchymal transition (EMT) was studied both in vitro and in vivo. The mechanism of the interaction between Robo1 and USP33 was explored by co-IP and ubiquitination protein analysis.

Results: The mRNA and protein levels of Slit2 and Robo1 are lower in GC tissues relative to those in adjacent healthy tissues. Importantly, Slit2 inhibits GC cell migration and suppresses EMT process in a Robo-dependent manner. The inhibitory function of Slit2-Robo1 is mediated by ubiquitin-specific protease 33 (USP33) via deubiquitinating and stabilizing Robo1. USP33 expression is decreased in GC tissues, and reduced USP33 level is correlated with poor patient survival.

Conclusions: Our study reveals the inhibitory function of Slit-Robo signalling in GC and uncovers a role of USP33 in suppressing cancer cell migration and EMT by enhancing Slit2-Robo1 signalling. USP33 represents a feasible choice as a prognostic biomarker for GC.
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http://dx.doi.org/10.1111/cpr.12606DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6536419PMC
May 2019

A crucial role for Arf6 in the response of commissural axons to Slit.

Development 2019 02 4;146(3). Epub 2019 Feb 4.

Department of Neurology, Lurie Comprehensive Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA

A switch in the response of commissural axons to the repellent Slit is crucial for ensuring that they cross the ventral midline only once. However, the underlying mechanisms remain to be elucidated. We have found that both endocytosis and recycling of Robo1 receptor are crucial for modulating Slit sensitivity in vertebrate commissural axons. Robo1 endocytosis and its recycling back to the cell surface maintained the stability of axonal Robo1 during Slit stimulation. We identified Arf6 guanosine triphosphatase and its activators, cytohesins, as previously unknown components in Slit-Robo1 signalling in vertebrate commissural neurons. Slit-Robo1 signalling activated Arf6. The -deficient mice exhibited marked defects in commissural axon midline crossing. Our data showed that a Robo1 endocytosis-triggered and Arf6-mediated positive-feedback strengthens the Slit response in commissural axons upon their midline crossing. Furthermore, the cytohesin-Arf6 pathways modulated this self-enhancement of the Slit response before and after midline crossing, resulting in a switch that reinforced robust regulation of axon midline crossing. Our study provides insights into endocytic trafficking-mediated mechanisms for spatiotemporally controlled axonal responses and uncovers new players in the midline switch in Slit responsiveness of commissural axons.
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http://dx.doi.org/10.1242/dev.172106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6382006PMC
February 2019

FUS interacts with ATP synthase beta subunit and induces mitochondrial unfolded protein response in cellular and animal models.

Proc Natl Acad Sci U S A 2018 10 24;115(41):E9678-E9686. Epub 2018 Sep 24.

State Key Laboratory of Brain and Cognitive Sciences, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China;

FUS (fused in sarcoma) proteinopathy is a group of neurodegenerative diseases characterized by the formation of inclusion bodies containing the FUS protein, including frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Previous studies show that mitochondrial damage is an important aspect of FUS proteinopathy. However, the molecular mechanisms by which FUS induces mitochondrial damage remain to be elucidated. Our biochemical and genetic experiments demonstrate that FUS interacts with the catalytic subunit of mitochondrial ATP synthase (ATP5B), disrupts the formation of ATP synthase complexes, and inhibits mitochondrial ATP synthesis. FUS expression activates the mitochondrial unfolded protein response (UPR). Importantly, down-regulating expression of ATP5B or UPR genes in FUS transgenic flies ameliorates neurodegenerative phenotypes. Our data show that mitochondrial impairment is a critical early event in FUS proteinopathy, and provide insights into the pathogenic mechanism of FUS-induced neurodegeneration.
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http://dx.doi.org/10.1073/pnas.1806655115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6187197PMC
October 2018

Measure and model a 3-D space-variant PSF for fluorescence microscopy image deblurring.

Opt Express 2018 May;26(11):14375-14391

Conventional deconvolution methods assume that the microscopy system is spatially invariant, introducing considerable errors. We developed a method to more precisely estimate space-variant point-spread functions from sparse measurements. To this end, a space-variant version of deblurring algorithm was developed and combined with a total-variation regularization. Validation with both simulation and real data showed that our PSF model is more accurate than the piecewise-invariant model and the blending model. Comparing with the orthogonal basis decomposition based PSF model, our proposed model also performed with a considerable improvement. We also evaluated the proposed deblurring algorithm. Our new deblurring algorithm showed a significantly better signal-to-noise ratio and higher image quality than those of the conventional space-invariant algorithm.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6005672PMC
http://dx.doi.org/10.1364/OE.26.014375DOI Listing
May 2018

Traumatic injury induces stress granule formation and enhances motor dysfunctions in ALS/FTD models.

Hum Mol Genet 2018 04;27(8):1366-1381

Division of Child Neurology, Department of Pediatrics, Children's Hospital of Pittsburgh, University of Pittsburgh Medical Center, Pittsburgh, PA 15224, USA.

Traumatic brain injury (TBI) has been predicted to be a predisposing factor for amyotrophic lateral sclerosis (ALS) and other neurological disorders. Despite the importance of TBI in ALS progression, the underlying cellular and molecular mechanisms are still an enigma. Here, we examined the contribution of TBI as an extrinsic factor and investigated whether TBI influences the susceptibility of developing neurodegenerative symptoms. To evaluate the effects of TBI in vivo, we applied mild to severe trauma to Drosophila and found that TBI leads to the induction of stress granules (SGs) in the brain. The degree of SGs induction directly correlates with the level of trauma. Furthermore, we observed that the level of mortality is directly proportional to the number of traumatic hits. Interestingly, trauma-induced SGs are ubiquitin, p62 and TDP-43 positive, and persistently remain over time suggesting that SGs might be aggregates and exert toxicity in our fly models. Intriguingly, TBI on animals expressing ALS-linked genes increased mortality and locomotion dysfunction suggesting that mild trauma might aggravate neurodegenerative symptoms associated with ALS. Furthermore, we found elevated levels of high molecular weight ubiquitinated proteins and p62 in animals expressing ALS-causing genes with TBI, suggesting that TBI may lead to the defects in protein degradation pathways. Finally, we observed that genetic and pharmacological induction of autophagy enhanced the clearance of SGs and promoted survival of flies in vivo. Together, our study demonstrates that trauma can induce SG formation in vivo and might enhance neurodegenerative phenotypes in the fly models of ALS.
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http://dx.doi.org/10.1093/hmg/ddy047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6455923PMC
April 2018

TDP-43 regulates cancer-associated microRNAs.

Protein Cell 2018 10 26;9(10):848-866. Epub 2017 Sep 26.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.

Aberrant regulation of miRNA genes contributes to pathogenesis of a wide range of human diseases, including cancer. The TAR DNA binding protein 43 (TDP-43), a RNA/DNA binding protein associated with neurodegeneration, is involved in miRNA biogenesis. Here, we systematically examined miRNAs regulated by TDP-43 using RNA-Seq coupled with an siRNA-mediated knockdown approach. TDP-43 knockdown affected the expression of a number of miRNAs. In addition, TDP-43 down-regulation led to alterations in the patterns of different isoforms of miRNAs (isomiRs) and miRNA arm selection, suggesting a previously unknown role of TDP-43 in miRNA processing. A number of TDP-43 associated miRNAs, and their candidate target genes, are associated with human cancers. Our data reveal highly complex roles of TDP-43 in regulating different miRNAs and their target genes. Our results suggest that TDP-43 may promote migration of lung cancer cells by regulating miR-423-3p. In contrast, TDP-43 increases miR-500a-3p expression and binds to the mature miR-500a-3p sequence. Reduced expression of miR-500a-3p is associated with poor survival of lung cancer patients, suggesting that TDP-43 may have a suppressive role in cancer by regulating miR-500a-3p. Cancer-associated genes LIF and PAPPA are possible targets of miR-500a-3p. Our work suggests that TDP-43-regulated miRNAs may play multifaceted roles in the pathogenesis of cancer.
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http://dx.doi.org/10.1007/s13238-017-0480-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6160384PMC
October 2018

Preparation of intact mitochondria using free-flow isoelectric focusing with post-pH gradient sample injection for morphological, functional and proteomics studies.

Anal Chim Acta 2017 Aug 17;982:200-208. Epub 2017 Jun 17.

State Key Laboratory of Microbial Metabolism, Laboratory of Analytical Biochemistry and Bioseparation, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. Electronic address:

Mitochondria play essential roles in both energy metabolism and cell signaling, which are critical for cell survival. Although significant efforts have been invested in understanding mitochondrial biology, methods for intact mitochondria preparation are technically challenging and remain to be improved. New methods for heterogeneous mitochondria purification will therefore boost our understanding on their physiological and biophysical properties. Herein, we developed a novel recycling free-flow isoelectric focusing (RFFIEF) with post-pH gradient sample injection (post-PGSI) for preparative separation of mitochondria. Crude mitochondria of rabbit liver obtained from differential centrifugation were purified by the developed method according to their pI values as six fractions. Transmission electron microscope images revealed that intact mitochondria existed in two fractions of pH 6.24 and 6.61, degenerative mitochondria were in two fractions of pH 5.46 and 5.72, and inner membrane vesicles (IMVs) appeared in the fractions of pH 4.70 and 5.04. Membrane potential measurement proved a dramatic difference between intact mitochondria and IMVs, which reflected the bioactivity of obtained populations. Particularly, proteomics analyses revealed that more number of proteins were identified in the intact fractions than that of IMVs or crude mitochondria, which demonstrated that RFFIEF could be powerful tool for the preparation of intact organelle as well as their proteomic and in-depth biological analysis.
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http://dx.doi.org/10.1016/j.aca.2017.06.007DOI Listing
August 2017

PINK1 and Parkin are genetic modifiers for FUS-induced neurodegeneration.

Hum Mol Genet 2016 12;25(23):5059-5068

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Science, Beijing, China.

Dysregulation of Fused in Sarcoma (FUS) gene expression is associated with fronto-temporal lobar degeneration (FTLD), and missense mutations in the FUS gene have been identified in patients affected by amyotrophic lateral sclerosis (ALS). However, molecular and cellular defects underlying FUS proteinopathy remain to be elucidated. Here, we examined whether genes important for mitochondrial quality control play a role in FUS proteinopathy. In our genetic screening, Pink1 and Park genes were identified as modifiers of neurodegeneration phenotypes induced by wild type (Wt) or ALS-associated P525L-mutant human FUS. Down-regulating expression of either Pink1 or Parkin genes ameliorated FUS-induced neurodegeneration phenotypes. The protein levels of PINK1 and Parkin were elevated in cells overexpressing FUS. Remarkably, ubiquitinylation of Miro1 protein, a downstream target of the E3 ligase activity of Parkin, was also increased in cells overexpressing FUS protein. In fly motor neurons expressing FUS, both motility and processivity of mitochondrial axonal transport were reduced by expression of either Wt- or P525L-mutant FUS. Finally, down-regulating PINK1 or Parkin partially rescued the locomotive defects and enhanced the survival rate in transgenic flies expressing FUS. Our data indicate that PINK1 and Parkin play an important role in FUS-induced neurodegeneration. This study has uncovered a previously unknown link between FUS proteinopathy and PINK1/Parkin genes, providing new insights into the pathogenesis of FUS proteinopathy.
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http://dx.doi.org/10.1093/hmg/ddw310DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6078632PMC
December 2016

A new method for quantifying mitochondrial axonal transport.

Protein Cell 2016 Nov 25;7(11):804-819. Epub 2016 May 25.

State Key Laboratory for Brain & Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.

Axonal transport of mitochondria is critical for neuronal survival and function. Automatically quantifying and analyzing mitochondrial movement in a large quantity remain challenging. Here, we report an efficient method for imaging and quantifying axonal mitochondrial transport using microfluidic-chamber-cultured neurons together with a newly developed analysis package named "MitoQuant". This tool-kit consists of an automated program for tracking mitochondrial movement inside live neuronal axons and a transient-velocity analysis program for analyzing dynamic movement patterns of mitochondria. Using this method, we examined axonal mitochondrial movement both in cultured mammalian neurons and in motor neuron axons of Drosophila in vivo. In 3 different paradigms (temperature changes, drug treatment and genetic manipulation) that affect mitochondria, we have shown that this new method is highly efficient and sensitive for detecting changes in mitochondrial movement. The method significantly enhanced our ability to quantitatively analyze axonal mitochondrial movement and allowed us to detect dynamic changes in axonal mitochondrial transport that were not detected by traditional kymographic analyses.
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http://dx.doi.org/10.1007/s13238-016-0268-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5084152PMC
November 2016

Noncanonical Myo9b-RhoGAP Accelerates RhoA GTP Hydrolysis by a Dual-Arginine-Finger Mechanism.

J Mol Biol 2016 07 27;428(15):3043-57. Epub 2016 Jun 27.

National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China. Electronic address:

The GTP hydrolysis activities of Rho GTPases are stimulated by GTPase-activating proteins (GAPs), which contain a RhoGAP domain equipped with a characteristic arginine finger and an auxiliary asparagine for catalysis. However, the auxiliary asparagine is missing in the RhoGAP domain of Myo9b (Myo9b-RhoGAP), a unique motorized RhoGAP that specifically targets RhoA for controlling cell motility. Here, we determined the structure of Myo9b-RhoGAP in complex with GDP-bound RhoA and magnesium fluoride. Unexpectedly, Myo9b-RhoGAP contains two arginine fingers at its catalytic site. The first arginine finger resembles the one within the canonical RhoGAP domains and inserts into the nucleotide-binding pocket of RhoA, whereas the second arginine finger anchors the Switch I loop of RhoA and interacts with the nucleotide, stabilizing the transition state of GTP hydrolysis and compensating for the lack of the asparagine. Mutating either of the two arginine fingers impaired the catalytic activity of Myo9b-RhoGAP and affected the Myo9b-mediated cell migration. Our data indicate that Myo9b-RhoGAP accelerates RhoA GTP hydrolysis by a previously unknown dual-arginine-finger mechanism, which may be shared by other noncanonical RhoGAP domains lacking the auxiliary asparagine.
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http://dx.doi.org/10.1016/j.jmb.2016.06.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5714546PMC
July 2016

MiR-130a regulates neurite outgrowth and dendritic spine density by targeting MeCP2.

Protein Cell 2016 07 1;7(7):489-500. Epub 2016 Jun 1.

State Key Laboratory for Brain & Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.

MicroRNAs (miRNAs) are critical for both development and function of the central nervous system. Significant evidence suggests that abnormal expression of miRNAs is associated with neurodevelopmental disorders. MeCP2 protein is an epigenetic regulator repressing or activating gene transcription by binding to methylated DNA. Both loss-of-function and gain-of-function mutations in the MECP2 gene lead to neurodevelopmental disorders such as Rett syndrome, autism and MECP2 duplication syndrome. In this study, we demonstrate that miR-130a inhibits neurite outgrowth and reduces dendritic spine density as well as dendritic complexity. Bioinformatics analyses, cell cultures and biochemical experiments indicate that miR-130a targets MECP2 and down-regulates MeCP2 protein expression. Furthermore, expression of the wild-type MeCP2, but not a loss-of-function mutant, rescues the miR-130a-induced phenotype. Our study uncovers the MECP2 gene as a previous unknown target for miR-130a, supporting that miR-130a may play a role in neurodevelopment by regulating MeCP2. Together with data from other groups, our work suggests that a feedback regulatory mechanism involving both miR-130a and MeCP2 may serve to ensure their appropriate expression and function in neural development.
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http://dx.doi.org/10.1007/s13238-016-0272-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4930766PMC
July 2016

Myo9b is a key player in SLIT/ROBO-mediated lung tumor suppression.

J Clin Invest 2015 Nov 3;125(12):4407-20. Epub 2015 Nov 3.

Emerging evidence indicates that the neuronal guidance molecule SLIT plays a role in tumor suppression, as SLIT-encoding genes are inactivated in several types of cancer, including lung cancer; however, it is not clear how SLIT functions in lung cancer. Here, our data show that SLIT inhibits cancer cell migration by activating RhoA and that myosin 9b (Myo9b) is a ROBO-interacting protein that suppresses RhoA activity in lung cancer cells. Structural analyses revealed that the RhoGAP domain of Myo9b contains a unique patch that specifically recognizes RhoA. We also determined that the ROBO intracellular domain interacts with the Myo9b RhoGAP domain and inhibits its activity; therefore, SLIT-dependent activation of RhoA is mediated by ROBO inhibition of Myo9b. In a murine model, compared with control lung cancer cells, SLIT-expressing cells had a decreased capacity for tumor formation and lung metastasis. Evaluation of human lung cancer and adjacent nontumor tissues revealed that Myo9b is upregulated in the cancer tissue. Moreover, elevated Myo9b expression was associated with lung cancer progression and poor prognosis. Together, our data identify Myo9b as a key player in lung cancer and as a ROBO-interacting protein in what is, to the best of our knowledge, a newly defined SLIT/ROBO/Myo9b/RhoA signaling pathway that restricts lung cancer progression and metastasis. Additionally, our work suggests that targeting the SLIT/ROBO/Myo9b/RhoA pathway has potential as a diagnostic and therapeutic strategy for lung cancer.
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http://dx.doi.org/10.1172/JCI81673DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4665778PMC
November 2015

Autism, an area that needs public attention and investment.

Authors:
Jane Y Wu

Sci China Life Sci 2015 Oct;58(10):931-2

Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611, USA.

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http://dx.doi.org/10.1007/s11427-015-4940-2DOI Listing
October 2015

FUS Interacts with HSP60 to Promote Mitochondrial Damage.

PLoS Genet 2015 Sep 3;11(9):e1005357. Epub 2015 Sep 3.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China; Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States of America.

FUS-proteinopathies, a group of heterogeneous disorders including ALS-FUS and FTLD-FUS, are characterized by the formation of inclusion bodies containing the nuclear protein FUS in the affected patients. However, the underlying molecular and cellular defects remain unclear. Here we provide evidence for mitochondrial localization of FUS and its induction of mitochondrial damage. Remarkably, FTLD-FUS brain samples show increased FUS expression and mitochondrial defects. Biochemical and genetic data demonstrate that FUS interacts with a mitochondrial chaperonin, HSP60, and that FUS translocation to mitochondria is, at least in part, mediated by HSP60. Down-regulating HSP60 reduces mitochondrially localized FUS and partially rescues mitochondrial defects and neurodegenerative phenotypes caused by FUS expression in transgenic flies. This is the first report of direct mitochondrial targeting by a nuclear protein associated with neurodegeneration, suggesting that mitochondrial impairment may represent a critical event in different forms of FUS-proteinopathies and a common pathological feature for both ALS-FUS and FTLD-FUS. Our study offers a potential explanation for the highly heterogeneous nature and complex genetic presentation of different forms of FUS-proteinopathies. Our data also suggest that mitochondrial damage may be a target in future development of diagnostic and therapeutic tools for FUS-proteinopathies, a group of devastating neurodegenerative diseases.
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http://dx.doi.org/10.1371/journal.pgen.1005357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4559378PMC
September 2015

CD146 acts as a novel receptor for netrin-1 in promoting angiogenesis and vascular development.

Cell Res 2015 Mar 6;25(3):275-87. Epub 2015 Feb 6.

Key Laboratory of Protein and Peptide Pharmaceuticals, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.

Angiogenesis, a process that newly-formed blood vessels sprout from pre-existing ones, is vital for vertebrate development and adult homeostasis. Previous studies have demonstrated that the neuronal guidance molecule netrin-1 participates in angiogenesis and morphogenesis of the vascular system. Netrin-1 exhibits dual activities in angiogenesis: either promoting or inhibiting angiogenesis. The anti-angiogenic activity of netrin-1 is mediated by UNC5B receptor. However, how netrin-1 promotes angiogenesis remained unclear. Here we report that CD146, an endothelial transmembrane protein of the immunoglobulin superfamily, is a receptor for netrin-1. Netrin-1 binds to CD146 with high affinity, inducing endothelial cell activation and downstream signaling in a CD146-dependent manner. Conditional knockout of the cd146 gene in the murine endothelium or disruption of netrin-CD146 interaction by a specific anti-CD146 antibody blocks or reduces netrin-1-induced angiogenesis. In zebrafish embryos, downregulating either netrin-1a or CD146 results in vascular defects with striking similarity. Moreover, knocking down CD146 blocks ectopic vascular sprouting induced by netrin-1 overexpression. Together, our data uncover CD146 as a previously unknown receptor for netrin-1 and also reveal a functional ligand for CD146 in angiogenesis, demonstrating the involvement of netrin-CD146 signaling in angiogenesis during vertebrate development.
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http://dx.doi.org/10.1038/cr.2015.15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4349246PMC
March 2015

Increased dysbindin-1B isoform expression in schizophrenia and its propensity in aggresome formation.

Cell Discov 2015 10;1:15032. Epub 2015 Nov 10.

National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Tsinghua University , Beijing, China.

Genetic variations in the human dysbindin-1 gene (DTNBP1) have been associated with schizophrenia. As a result of alternative splicing, the human DTNBP1 gene generates at least three distinct protein isoforms, dysbindin-1A, -1B and -1C. Significant effort has focused on dysbindin-1A, an important player in multiple steps of neurodevelopment. However, the other isoforms, dysbindin-1B and dysbindin-1C have not been well characterized. Nor have been associated with human diseases. Here we report an increase in expression of DTNBP1b mRNA in patients with paranoid schizophrenia as compared with healthy controls. A single-nucleotide polymorphism located in intron 9, rs117610176, has been identified and associated with paranoid schizophrenia, and its C allele leads to an increase of DTNBP1b mRNA splicing. Our data show that different dysbindin splicing isoforms exhibit distinct subcellular distribution, suggesting their distinct functional activities. Dysbindin-1B forms aggresomes at the perinuclear region, whereas dysbindin-1A and -1C proteins exhibit diffused patterns in the cytoplasm. Dysbindin-1A interacts with dysbindin-1B, getting recruited to the aggresome structure when co-expressed with dysbindin-1B. Moreover, cortical neurons over-expressing dysbindin-1B show reduction in neurite outgrowth, suggesting that dysbindin-1B may interfere with dysbindin-1A function in a dominant-negative manner. Taken together, our study uncovers a previously unknown association of DTNBP1b expression with schizophrenia in addition to its distinct biochemical and functional properties.
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http://dx.doi.org/10.1038/celldisc.2015.32DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4860834PMC
July 2016

USP33 mediates Slit-Robo signaling in inhibiting colorectal cancer cell migration.

Int J Cancer 2015 Apr 29;136(8):1792-802. Epub 2014 Sep 29.

Wuxi Oncology Institute, the Affiliated Hospital of Jiangnan University, Wuxi, Jiangsu, China; Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, 303 E. Chicago Ave., Chicago, IL.

Originally discovered in neuronal guidance, the Slit-Robo pathway is emerging as an important player in human cancers. However, its involvement and mechanism in colorectal cancer (CRC) remains to be elucidated. Here, we report that Slit2 expression is reduced in CRC tissues compared with adjacent noncancerous tissues. Extensive promoter hypermethylation of the Slit2 gene has been observed in CRC cells, which provides a mechanistic explanation for the Slit2 downregulation in CRC. Functional studies showed that Slit2 inhibits CRC cell migration in a Robo-dependent manner. Robo-interacting ubiquitin-specific protease 33 (USP33) is required for the inhibitory function of Slit2 on CRC cell migration by deubiquitinating and stabilizing Robo1. USP33 expression is downregulated in CRC samples, and reduced USP33 mRNA levels are correlated with increased tumor grade, lymph node metastasis and poor patient survival. Taken together, our data reveal USP33 as a previously unknown tumor-suppressing gene for CRC by mediating the inhibitory function of Slit-Robo signaling on CRC cell migration. Our work suggests the potential value of USP33 as an independent prognostic marker of CRC.
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http://dx.doi.org/10.1002/ijc.29226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4323690PMC
April 2015

An ALS-mutant TDP-43 neurotoxic peptide adopts an anti-parallel β-structure and induces TDP-43 redistribution.

Hum Mol Genet 2014 Dec 11;23(25):6863-77. Epub 2014 Aug 11.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, USA

TDP-43 proteinopathies are clinically and genetically heterogeneous diseases that had been considered distinct from classical amyloid diseases. Here, we provide evidence for the structural similarity between TDP-43 peptides and other amyloid proteins. Atomic force microscopy and electron microscopy examination of peptides spanning a previously defined amyloidogenic fragment revealed a minimal core region that forms amyloid fibrils similar to the TDP-43 fibrils detected in FTLD-TDP brain tissues. An ALS-mutant A315E amyloidogenic TDP-43 peptide is capable of cross-seeding other TDP-43 peptides and an amyloid-β peptide. Sequential Nuclear Overhauser Effects and double-quantum-filtered correlation spectroscopy in nuclear magnetic resonance (NMR) analyses of the A315E-mutant TDP-43 peptide indicate that it adopts an anti-parallel β conformation. When added to cell cultures, the amyloidogenic TDP-43 peptides induce TDP-43 redistribution from the nucleus to the cytoplasm. Neuronal cultures in compartmentalized microfluidic-chambers demonstrate that the TDP-43 peptides can be taken up by axons and induce axonotoxicity and neuronal death, thus recapitulating key neuropathological features of TDP-43 proteinopathies. Importantly, a single amino acid change in the amyloidogenic TDP-43 peptide that disrupts fibril formation also eliminates neurotoxicity, supporting that amyloidogenesis is critical for TDP-43 neurotoxicity.
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http://dx.doi.org/10.1093/hmg/ddu409DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4245047PMC
December 2014

USP33, a new player in lung cancer, mediates Slit-Robo signaling.

Protein Cell 2014 Sep 2;5(9):704-13. Epub 2014 Jul 2.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.

Ubiquitin specific protease 33 (USP33) is a multifunctional protein regulating diverse cellular processes. The expression and role of USP33 in lung cancer remain unexplored. In this study, we show that USP33 is down-regulated in multiple cohorts of lung cancer patients and that low expression of USP33 is associated with poor prognosis. USP33 mediates Slit-Robo signaling in lung cancer cell migration. Downregulation of USP33 reduces the protein stability of Robo1 in lung cancer cells, providing a previously unknown mechanism for USP33 function in mediating Slit activity in lung cancer cells. Taken together, USP33 is a new player in lung cancer that regulates Slit-Robo signaling. Our data suggest that USP33 may be a candidate tumor suppressor for lung cancer with potential as a prognostic marker.
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http://dx.doi.org/10.1007/s13238-014-0070-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4145083PMC
September 2014

RNA-binding proteins in neurological diseases.

Sci China Life Sci 2014 Apr;57(4):432-44

Emerging studies support that RNA-binding proteins (RBPs) play critical roles in human biology and pathogenesis. RBPs are essential players in RNA processing and metabolism, including pre-mRNA splicing, polyadenylation, transport, surveillance, mRNA localization, mRNA stability control, translational control and editing of various types of RNAs. Aberrant expression of and mutations in RBP genes affect various steps of RNA processing, altering target gene function. RBPs have been associated with various diseases, including neurological diseases. Here, we mainly focus on selected RNA-binding proteins including Nova-1/Nova-2, HuR/HuB/HuC/HuD, TDP-43, Fus, Rbfox1/Rbfox2, QKI and FMRP, discussing their function and roles in human diseases.
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http://dx.doi.org/10.1007/s11427-014-4647-9DOI Listing
April 2014

AxonQuant: A Microfluidic Chamber Culture-Coupled Algorithm That Allows High-Throughput Quantification of Axonal Damage.

Neurosignals 2014 28;22(1):14-29. Epub 2014 Feb 28.

School of Electronic Science and Engineering, Nanjing University, Nanjing, China.

Published methods for imaging and quantitatively analyzing morphological changes in neuronal axons have serious limitations because of their small sample sizes, and their time-consuming and nonobjective nature. Here we present an improved microfluidic chamber design suitable for fast and high-throughput imaging of neuronal axons. We developed the AxonQuant algorithm, which is suitable for automatic processing of axonal imaging data. This microfluidic chamber-coupled algorithm allows calculation of an 'axonal continuity index' that quantitatively measures axonal health status in a manner independent of neuronal or axonal density. This method allows quantitative analysis of axonal morphology in an automatic and nonbiased manner. Our method will facilitate large-scale high-throughput screening for genes or therapeutic compounds for neurodegenerative diseases involving axonal damage. When combined with imaging technologies utilizing different gene markers, this method will provide new insights into the mechanistic basis for axon degeneration. Our microfluidic chamber culture-coupled AxonQuant algorithm will be widely useful for studying axonal biology and neurodegenerative disorders.
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http://dx.doi.org/10.1159/000358092DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4148468PMC
October 2015

Axon guidance factor SLIT2 inhibits neural invasion and metastasis in pancreatic cancer.

Cancer Res 2014 Mar 21;74(5):1529-40. Epub 2014 Jan 21.

Authors' Affiliations: Experimental and Clinical Research Center, a joint cooperation between the Charité Medical Faculty and the Max-Delbrück Center for Molecular Medicine, Berlin, Germany; Medizinische Klinik mit Schwerpunkt Hepatologie und Gastroenterologie; Klinik für Viszeral-, Allgemein-, und Transplantationschirurgie; Institut für Pathologie, Charité-Universitätsmedizin Berlin, Germany; and Department of Neurology, Lurie Cancer Center, Center for Genetic Medicine, Northwestern University Feinberg School of Medicine.

Pancreatic ductal adenocarcinoma (PDAC) metastasizes by neural, vascular, and local invasion routes, which limit patient survival. In nerves and vessels, SLIT2 and its ROBO receptors constitute repellent guidance cues that also direct epithelial branching. Thus, the SLIT2-ROBO system may represent a key pinch point to regulate PDAC spread. In this study, we examined the hypothesis that escaping from repellent SLIT2-ROBO signaling is essential to enable PDAC cells to appropriate their local stromal infrastructure for dissemination. Through immunohistochemical analysis, we detected SLIT2 receptors ROBO1 and ROBO4 on epithelia, nerves, and vessels in healthy pancreas and PDAC specimens, respectively. SLIT2 mRNA expression was reduced in PDAC compared with nontransformed pancreatic tissues and cell lines, suggesting a reduction in SLIT2-ROBO pathway activity in PDAC. In support of this interpretation, restoring the SLIT2 expression in SLIT2-deficient PDAC cells inhibited their bidirectional chemoattraction with neural cells, and more specifically, impaired unidirectional PDAC cell navigation along outgrowing neurites in models of neural invasion. Restoring autocrine/paracrine SLIT2 signaling was also sufficient to inhibit the directed motility of PDAC cells, but not their random movement. Conversely, RNA interference-mediated silencing of ROBO1 stimulated the motility of SLIT2-competent PDAC cells. Furthermore, culture supernatants from SLIT2-competent PDAC cells impaired migration of endothelial cells (human umbilical vein endothelial cells), whereas an N-terminal SLIT2 cleavage fragment stimulated such migration. In vivo investigations of pancreatic tumors with restored SLIT2 expression demonstrated reduced invasion, metastasis, and vascularization, with opposing effects produced by ROBO1 silencing in tumor cells or sequestration of endogenous SLIT2. Analysis of clinical specimens of PDAC showed that those with low SLIT2 mRNA expression exhibited a higher incidence and a higher fraction of tumor-infiltrated lymph nodes. Taken together, our findings argue that disrupting SLIT2-ROBO signaling in PDAC may enhance metastasis and predispose PDAC cells to neural invasion.
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http://dx.doi.org/10.1158/0008-5472.CAN-13-1012DOI Listing
March 2014

Alternative Pre-mRNA Splicing, Cell Death, and Cancer.

Cancer Treat Res 2013 ;158:181-212

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Science, Beijing, China.

Alternative splicing is one of the most powerful mechanisms for generating functionally distinct products from a single genetic loci and for fine-tuning gene activities at the post-transcriptional level. Alternative splicing plays important roles in regulating genes critical for cell death. These cell death genes encode death ligands, cell surface death receptors, intracellular death regulators, signal transduction molecules, and death executor enzymes such as caspases and nucleases. Alternative splicing of these genes often leads to the formation of functionally different products, some of which have antagonistic effects that are either cell death-promoting or cell death-preventing. Differential alternative splicing can affect expression, subcellular distribution, and functional activities of the gene products. Molecular defects in splicing regulation of cell death genes have been associated with cancer development and resistance to treatment. Studies using molecular, biochemical, and systems-based approaches have begun to reveal mechanisms underlying the regulation of alternative splicing of cell death genes. Systematic studies have begun to uncover the multi-level interconnected networks that regulate alternative splicing. A global picture of the complex mechanisms that regulate cell death genes at the pre-mRNA splicing level has thus begun to emerge.
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http://dx.doi.org/10.1007/978-3-642-31659-3_8DOI Listing
April 2016

AMPK interacts with DSCAM and plays an important role in netrin-1 induced neurite outgrowth.

Protein Cell 2013 Feb 12;4(2):155-61. Epub 2013 Mar 12.

State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.

Down syndrome cell adhesion molecule (DSCAM) acts as a netrin-1 receptor and mediates attractive response of axons to netrin-1 in neural development. However, the signaling mechanisms of netrin-DSCAM remain unclear. Here we report that AMP-activated protein kinase (AMPK) interacts with DSCAM through its γ subunit, but does not interact with DCC (deleted in colorectal cancer), another major receptor for netrin-1. Netrin-treatment of cultured cortical neurons leads to increased phosphorylation of AMPK. Both AMPK mutant with dominant-negative effect and AMPK inhibitor can significantly suppress netrin-1 induced neurite outgrowth. Together, these findings demonstrate that AMPK interacts with DSCAM and plays an important role in netrin-1 induced neurite outgrowth. Our study uncovers a previously unknown component, AMPK, in netrin-DSCAM signaling pathway.
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http://dx.doi.org/10.1007/s13238-012-2126-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3893083PMC
February 2013

Characterization of β-domains in C-terminal fragments of TDP-43 by scanning tunneling microscopy.

J Struct Biol 2013 Jan 5;181(1):11-6. Epub 2012 Nov 5.

Key Laboratory for Biological Effects of Nanomaterials and Nanosafety (Chinese Academy of Sciences), National Center for Nanoscience and Technology, Beijing 100190, China.

The TAR DNA-binding protein 43 (TDP-43) has been identified as a critical player in a range of neurodegenerative diseases, including frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Recent discoveries demonstrate the important role of carboxyl-terminal fragments of TDP-43 in its proteinopathy. Herein, we report the characterization of β-domains in the C-terminal fragments of TDP-43 using scanning tunneling microscopy (STM). Careful comparison of the wild-type TDP-43 (Wt) and the three mutant TDP-43 peptides: an ALS-related mutant peptide: phosphorylated A315T mutant TDP-43 (A315T(p)) and two model peptides: A315T mutant TDP-43 (A315T), A315E mutant TDP-43 (A315E) reveals that A315T(p) has a longer core region of the β-domain than Wt. A315E possesses the longest core region of the β-domain and A315T(p) mutant TDP-43 has the second longest core region of the β-domain. The core regions of the β-domains for A315T and Wt TDP-43 have the same length. This observation provides a supportive evidence of a higher tendency in beta-sheet formation of A315T(p) containing TDP-43 fragment, and structural mechanism for the higher cytotoxicity and accelerated fibril formation of the A315T(p) mutation-containing TDP-43 peptide as compared with Wt TDP-43.
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http://dx.doi.org/10.1016/j.jsb.2012.10.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3787119PMC
January 2013