Publications by authors named "Kevin M Wright"

44 Publications

Characterization of PARP6 Function in Knockout Mice and Patients with Developmental Delay.

Cells 2021 May 22;10(6). Epub 2021 May 22.

Department of Physiology and Pharmacology, Oregon Health & Science University, Portland, OR 97210, USA.

PARP6, a member of a family of enzymes (17 in humans) known as poly-ADP-ribose polymerases (PARPs), is a neuronally enriched PARP. While previous studies from our group show that Parp6 is a regulator of dendrite morphogenesis in rat hippocampal neurons, its function in the nervous system in vivo is poorly understood. Here, we describe the generation of a loss-of-function mouse model for examining the function of Parp6 during neurodevelopment in vivo. Using CRISPR-Cas9 mutagenesis, we generated a mouse line that expressed a Parp6 truncated variant (Parp6) in place of Parp6. Unlike Parp6, Parp6 is devoid of catalytic activity. Homozygous Parp6 do not exhibit obvious neuromorphological defects during development, but nevertheless die perinatally. This suggests that Parp6 catalytic activity is important for postnatal survival. We also report PARP6 mutations in six patients with several neurodevelopmental disorders, including microencephaly, intellectual disabilities, and epilepsy. The most severe mutation in (C563R) results in the loss of catalytic activity. Expression of Parp6 in hippocampal neurons decreases dendrite morphogenesis. To gain further insight into function in neurons we also performed a BioID proximity labeling experiment in hippocampal neurons and identified several microtubule-binding proteins (e.g., MAP-2) using proteomics. Taken together, our results suggest that PARP6 is an essential microtubule-regulatory gene in mice, and that the loss of PARP6 catalytic activity has detrimental effects on neuronal function in humans.
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http://dx.doi.org/10.3390/cells10061289DOI Listing
May 2021

Immunohistochemical and Genetic Labeling of Hairy and Glabrous Skin Innervation.

Curr Protoc 2021 May;1(5):e121

Vollum Institute, Oregon Health & Science University, Portland, Oregon.

Cutaneous innervation is an essential component of the mammalian sensory nervous system. During development, genetically and morphologically diverse subtypes of sensory neurons use distinct molecular pathways to innervate end organs or form free nerve endings in glabrous and hairy skin. Peripheral neurons can be damaged by acute injury or degenerate due to chronic conditions including diabetes and chemotherapy, leading to peripheral neuropathy. The analysis of skin and cutaneous innervation can be applied to many research endeavors, from developmental neuroscience to pharmaceutical testing. Due to the natural hydrophobicity and heterogenous makeup of the skin (dense, keratinized cells as well as sparse, extracellular-matrix-bound cells), its histological analysis presents unique challenges compared to that of many other tissues. This series of protocols describes histological methods for generalized immunohistochemistry and subtype-specific genetic labeling of sensory neurons in mouse skin in both whole-mount and section formats. We provide detailed methodology of tissue preparation for hairy and glabrous skin, several types of labeling, and counting of hair follicles in flat-mounted mouse skin. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Cryosectioning and immunostaining of mouse hairy skin Alternate Protocol 1: Alternate method for preparation and fixation of mouse hairy skin Basic Protocol 2: Sectioning of mouse paw glabrous skin Basic Protocol 3: Whole-mount immunolabeling of mouse skin Basic Protocol 4: Sparse labeling of skin-innervating neurons with a Cre-dependent membrane-bound alkaline phosphatase reporter Alternate Protocol 2: Sparse labeling of skin-innervating neurons with a Cre-dependent fluorescent reporter Basic Protocol 5: Oil Red O staining of skin.
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http://dx.doi.org/10.1002/cpz1.121DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8162737PMC
May 2021

Spatially mapped single-cell chromatin accessibility.

Nat Commun 2021 02 24;12(1):1274. Epub 2021 Feb 24.

Molecular and Medical Genetics, Oregon Health & Science University, Portland, OR, USA.

High-throughput single-cell epigenomic assays can resolve cell type heterogeneity in complex tissues, however, spatial orientation is lost. Here, we present single-cell combinatorial indexing on Microbiopsies Assigned to Positions for the Assay for Transposase Accessible Chromatin, or sciMAP-ATAC, as a method for highly scalable, spatially resolved, single-cell profiling of chromatin states. sciMAP-ATAC produces data of equivalent quality to non-spatial sci-ATAC and retains the positional information of each cell within a 214 micron cubic region, with up to hundreds of tracked positions in a single experiment. We apply sciMAP-ATAC to assess cortical lamination in the adult mouse primary somatosensory cortex and in the human primary visual cortex, where we produce spatial trajectories and integrate our data with non-spatial single-nucleus RNA and other chromatin accessibility single-cell datasets. Finally, we characterize the spatially progressive nature of cerebral ischemic infarction in the mouse brain using a model of transient middle cerebral artery occlusion.
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http://dx.doi.org/10.1038/s41467-021-21515-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7904839PMC
February 2021

GABAergic neurons are susceptible to BAX-dependent apoptosis following isoflurane exposure in the neonatal period.

PLoS One 2021 12;16(1):e0238799. Epub 2021 Jan 12.

Vollum Institute, Oregon Health & Science University, Portland, Oregon, United States of America.

Exposure to volatile anesthetics during the neonatal period results in acute neuron death. Prior work suggests that apoptosis is the dominant mechanism mediating neuron death. We show that Bax deficiency blocks neuronal death following exposure to isoflurane during the neonatal period. Blocking Bax-mediated neuron death attenuated the neuroinflammatory response of microglia following isoflurane exposure. We find that GABAergic interneurons are disproportionately overrepresented among dying neurons. Despite the increase in neuronal apoptosis induced by isoflurane exposure during the neonatal period, seizure susceptibility, spatial memory retention, and contextual fear memory were unaffected later in life. However, Bax deficiency alone led to mild deficiencies in spatial memory and contextual fear memory, suggesting that normal developmental apoptotic death is important for cognitive function. Collectively, these findings show that while GABAergic neurons in the neonatal brain undergo elevated Bax-dependent apoptotic cell death following exposure to isoflurane, this does not appear to have long-lasting consequences on overall neurological function later in life.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0238799PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7802958PMC
April 2021

Gbx2 Identifies Two Amacrine Cell Subtypes with Distinct Molecular, Morphological, and Physiological Properties.

Cell Rep 2020 11;33(7):108382

Vollum Institute, Oregon Health and Science University, Portland, OR 97239, USA; Department of Ophthalmology, Casey Eye Institute, Oregon Health and Science University, Portland, OR 97239, USA. Electronic address:

Our understanding of nervous system function is limited by our ability to identify and manipulate neuronal subtypes within intact circuits. We show that the Gbx2 mouse line labels two amacrine cell (AC) subtypes in the mouse retina that have distinct morphological, physiological, and molecular properties. Using a combination of RNA-seq, genetic labeling, and patch clamp recordings, we show that one subtype is GABAergic that receives excitatory input from On bipolar cells. The other population is a non-GABAergic, non-glycinergic (nGnG) AC subtype that lacks the expression of standard neurotransmitter markers. Gbx2 nGnG ACs have smaller, asymmetric dendritic arbors that receive excitatory input from both On and Off bipolar cells. Gbx2 nGnG ACs also exhibit spatially restricted tracer coupling to bipolar cells (BCs) through gap junctions. This study identifies a genetic tool for investigating the two distinct AC subtypes, and it provides a model for studying synaptic communication and visual circuit function.
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http://dx.doi.org/10.1016/j.celrep.2020.108382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7713908PMC
November 2020

FDA Approves Pralsetinib for Treatment of Adults With Metastatic RET Fusion-Positive NSCLC.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 Oct;34(10):406-406;431

The US FDA granted accelerated approval to pralsetinib for the treatment of adult patients with metastatic RET fusion-positive non-small cell lung cancer.
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http://dx.doi.org/10.46883/ONC.2020.3410.0406DOI Listing
October 2020

Two Retrospective Analyses Show No Associated Adverse Outcomes With Delayed RP.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 09;34(9):344-345

The results of 2 studies showed no association between delayed radical prostatectomy(RP) and adverse oncological outcomes, supporting current recommendations of urologic societies for surgical treatment of patients with intermediate- and high-risk prostate cancer during the coronavirus disease 2019 (COVID-19)pandemic.
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http://dx.doi.org/10.46883/ONC.2020.3409.0344DOI Listing
September 2020

Final Data Analysis Supports Tivozanib as Superior Treatment for Patients With RCC.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 Jul;34(7):257

Tivozanib (Fotivda) significantly improved progression-free survival (PFS), compared with sorafenib (Nexavar), in patients with highly relapsed or refractory metastatic renal cell carcinoma (RCC), according to results from the phase 3 TIVO-3 trial presented at the 2020 American Society of Clinical Oncology Virtual Scientific Program.
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July 2020

Patient Reported Outcomes Show Newer Drug Combinations Maintain Quality of Life Longer Than Current Standard of Care Treatments.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 Feb;34(2):61-62

Multiple studies showed improved patient outcomes and increased quality of life among various treatments that previously demonstrated clinical benefits. these patient-centric findings, along with a promising new combination therapy for patients with advanced hepatocellular carcinoma highlight some of the latest research to come out of the 2020 Gastrointestinal Cancers Symposium.
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February 2020

FDA Approves Pembrolizumab for BCG-Unresponsive NMIBC.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 Feb;34(2):44

The FDA approved pembrolizumab (Keytruda) for the treatment of patients with Bacillus Calmette-Guerin (BCG)-unresponsive, high-risk, non-muscle invasive bladder cancer (NMIBC) with carcinoma in situ (CIS) with or without papillary tumors who are ineligible for or chose to not undergo cystectomy.
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February 2020

Further Analyses Highlight Benefits of PARP Inhibitors As Frontline Maintenance in Ovarian Cancer.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 May;34(5):175

Evidence for maintenance treatment with PARP inhibitors as the standard of care in the frontline setting for patients with ovarian cancer continues to mount, according to additional analyses from the phase 3 PRIMA trial and the phase 3 PAOLA-1 trial released as part of the virtual platform for the Society of Gynecologic Oncology (SGO) 2020 Annual Meeting on Women's Cancer.
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May 2020

FDA Approves 2 Therapies for Treatment of Metastatic Non-Small Cell Lung Cancer.

Authors:
Kevin M Wright

Oncology (Williston Park) 2020 Jun;34(6):202

The FDA approved 2 new therapies, each with its own companion diagnostic, for the treatment of patients with metastatic non-small cell lung cancer.
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June 2020

Single-cell transcriptomics of the naked mole-rat reveals unexpected features of mammalian immunity.

PLoS Biol 2019 11 21;17(11):e3000528. Epub 2019 Nov 21.

Calico Life Sciences LLC, South San Francisco, California, United States of America.

The immune system comprises a complex network of specialized cells that protects against infection, eliminates cancerous cells, and regulates tissue repair, thus serving a critical role in homeostasis, health span, and life span. The subterranean-dwelling naked mole-rat (NM-R; Heterocephalus glaber) exhibits prolonged life span relative to its body size, is unusually cancer resistant, and manifests few physiological or molecular changes with advancing age. We therefore hypothesized that the immune system of NM-Rs evolved unique features that confer enhanced cancer immunosurveillance and prevent the age-associated decline in homeostasis. Using single-cell RNA-sequencing (scRNA-seq) we mapped the immune system of the NM-R and compared it to that of the short-lived, cancer-prone mouse. In contrast to the mouse, we find that the NM-R immune system is characterized by a high myeloid-to-lymphoid cell ratio that includes a novel, lipopolysaccharide (LPS)-responsive, granulocyte cell subset. Surprisingly, we also find that NM-Rs lack canonical natural killer (NK) cells. Our comparative genomics analyses support this finding, showing that the NM-R genome lacks an expanded gene family that controls NK cell function in several other species. Furthermore, we reconstructed the evolutionary history that likely led to this genomic state. The NM-R thus challenges our current understanding of mammalian immunity, favoring an atypical, myeloid-biased mode of innate immunosurveillance, which may contribute to its remarkable health span.
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http://dx.doi.org/10.1371/journal.pbio.3000528DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6894886PMC
November 2019

A Prospective Analysis of Genetic Variants Associated with Human Lifespan.

G3 (Bethesda) 2019 09 4;9(9):2863-2878. Epub 2019 Sep 4.

Calico Life Sciences LLC. 1170 Veterans Blvd, South San Francisco 94080

We present a massive investigation into the genetic basis of human lifespan. Beginning with a genome-wide association (GWA) study using a de-identified snapshot of the unique database - more than 300,000 genotyped individuals linked to pedigrees of over 400,000,000 people - we mapped six genome-wide significant loci associated with parental lifespan. We compared these results to a GWA analysis of the traditional lifespan proxy trait, age, and found only one locus, , to be associated with both age and lifespan. By combining the results with those of an independent UK Biobank dataset, we conducted a meta-analysis of more than 650,000 individuals and identified fifteen parental lifespan-associated loci. Beyond just those significant loci, our genome-wide set of polymorphisms accounts for up to 8% of the variance in human lifespan; this value represents a large fraction of the heritability estimated from phenotypic correlations between relatives.
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http://dx.doi.org/10.1534/g3.119.400448DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6723124PMC
September 2019

The expanding functional roles and signaling mechanisms of adhesion G protein-coupled receptors.

Ann N Y Acad Sci 2019 11 6;1456(1):5-25. Epub 2019 Jun 6.

Vollum Institute, Oregon Health & Science University, Portland, Oregon.

The adhesion class of G protein-coupled receptors (GPCRs) is the second largest family of GPCRs (33 members in humans). Adhesion GPCRs (aGPCRs) are defined by a large extracellular N-terminal region that is linked to a C-terminal seven transmembrane (7TM) domain via a GPCR-autoproteolysis inducing (GAIN) domain containing a GPCR proteolytic site (GPS). Most aGPCRs undergo autoproteolysis at the GPS motif, but the cleaved fragments stay closely associated, with the N-terminal fragment (NTF) bound to the 7TM of the C-terminal fragment (CTF). The NTFs of most aGPCRs contain domains known to be involved in cell-cell adhesion, while the CTFs are involved in classical G protein signaling, as well as other intracellular signaling. In this workshop report, we review the most recent findings on the biology, signaling mechanisms, and physiological functions of aGPCRs.
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http://dx.doi.org/10.1111/nyas.14094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7891679PMC
November 2019

Dystroglycan is a scaffold for extracellular axon guidance decisions.

Elife 2019 02 13;8. Epub 2019 Feb 13.

Vollum Institute, Oregon Health & Science University, Portland, United States.

Axon guidance requires interactions between extracellular signaling molecules and transmembrane receptors, but how appropriate context-dependent decisions are coordinated outside the cell remains unclear. Here we show that the transmembrane glycoprotein Dystroglycan interacts with a changing set of environmental cues that regulate the trajectories of extending axons throughout the mammalian brain and spinal cord. Dystroglycan operates primarily as an extracellular scaffold during axon guidance, as it functions non-cell autonomously and does not require signaling through its intracellular domain. We identify the transmembrane receptor Celsr3/Adgrc3 as a binding partner for Dystroglycan, and show that this interaction is critical for specific axon guidance events in vivo. These findings establish Dystroglycan as a multifunctional scaffold that coordinates extracellular matrix proteins, secreted cues, and transmembrane receptors to regulate axon guidance.
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http://dx.doi.org/10.7554/eLife.42143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6395066PMC
February 2019

Estimates of the Heritability of Human Longevity Are Substantially Inflated due to Assortative Mating.

Genetics 2018 11;210(3):1109-1124

Ancestry, San Francisco, California 94107

Human life span is a phenotype that integrates many aspects of health and environment into a single ultimate quantity: the elapsed time between birth and death. Though it is widely believed that long life runs in families for genetic reasons, estimates of life span "heritability" are consistently low (∼15-30%). Here, we used pedigree data from public trees, including hundreds of millions of historical persons, to estimate the heritability of human longevity. Although "nominal heritability" estimates based on correlations among genetic relatives agreed with prior literature, the majority of that correlation was also captured by correlations among nongenetic (in-law) relatives, suggestive of highly assortative mating around life span-influencing factors (genetic and/or environmental). We used structural equation modeling to account for assortative mating, and concluded that the true heritability of human longevity for birth cohorts across the 1800s and early 1900s was well below 10%, and that it has been generally overestimated due to the effect of assortative mating.
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http://dx.doi.org/10.1534/genetics.118.301613DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6218226PMC
November 2018

Retinal ganglion cell axon sorting at the optic chiasm requires dystroglycan.

Dev Biol 2018 10 24;442(2):210-219. Epub 2018 Aug 24.

Neuroscience Graduate Program, Oregon Health&Science University, Portland, OR 97239, USA; Vollum Institute, Oregon Health&Science University, Portland, OR 97239, USA. Electronic address:

In the developing visual system, retinal ganglion cell (RGC) axons project from the retina to several distal retinorecipient regions in the brain. Several molecules have been implicated in guiding RGC axons in vivo, but the role of extracellular matrix molecules in this process remains poorly understood. Dystroglycan is a laminin-binding transmembrane protein important for formation and maintenance of the extracellular matrix and basement membranes and has previously been implicated in axon guidance in the developing spinal cord. Using two genetic models of functional dystroglycan loss, we show that dystroglycan is necessary for correct sorting of contralateral and ipsilateral RGC axons at the optic chiasm. Mis-sorted axons still target retinorecipient brain regions and persist in adult mice, even after axon pruning is complete. Our results highlight the importance of the extracellular matrix for axon sorting at an intermediate choice point in the developing visual circuit.
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http://dx.doi.org/10.1016/j.ydbio.2018.08.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6190595PMC
October 2018

Cas Adaptor Proteins Coordinate Sensory Axon Fasciculation.

Sci Rep 2018 04 16;8(1):5996. Epub 2018 Apr 16.

Neuroscience Program, Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA, 92521, USA.

Development of complex neural circuits like the peripheral somatosensory system requires intricate mechanisms to ensure axons make proper connections. While much is known about ligand-receptor pairs required for dorsal root ganglion (DRG) axon guidance, very little is known about the cytoplasmic effectors that mediate cellular responses triggered by these guidance cues. Here we show that members of the Cas family of cytoplasmic signaling adaptors are highly phosphorylated in central projections of the DRG as they enter the spinal cord. Furthermore, we provide genetic evidence that Cas proteins regulate fasciculation of DRG sensory projections. These data establish an evolutionarily conserved requirement for Cas adaptor proteins during peripheral nervous system axon pathfinding. They also provide insight into the interplay between axonal fasciculation and adhesion to the substrate.
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http://dx.doi.org/10.1038/s41598-018-24261-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5902548PMC
April 2018

Dystroglycan Maintains Inner Limiting Membrane Integrity to Coordinate Retinal Development.

J Neurosci 2017 08 31;37(35):8559-8574. Epub 2017 Jul 31.

Neuroscience Graduate Program and

Proper neural circuit formation requires the precise regulation of neuronal migration, axon guidance, and dendritic arborization. Mutations affecting the function of the transmembrane glycoprotein dystroglycan cause a form of congenital muscular dystrophy that is frequently associated with neurodevelopmental abnormalities. Despite its importance in brain development, the role of dystroglycan in regulating retinal development remains poorly understood. Using a mouse model of dystroglycanopathy ( ) and conditional mutants of both sexes, we show that dystroglycan is critical for the proper migration, axon guidance, and dendritic stratification of neurons in the inner retina. Using genetic approaches, we show that dystroglycan functions in neuroepithelial cells as an extracellular scaffold to maintain the integrity of the retinal inner limiting membrane. Surprisingly, despite the profound disruptions in inner retinal circuit formation, spontaneous retinal activity is preserved. These results highlight the importance of dystroglycan in coordinating multiple aspects of retinal development. The extracellular environment plays a critical role in coordinating neuronal migration and neurite outgrowth during neural circuit development. The transmembrane glycoprotein dystroglycan functions as a receptor for multiple extracellular matrix proteins and its dysfunction leads to a form of muscular dystrophy frequently associated with neurodevelopmental defects. Our results demonstrate that dystroglycan is required for maintaining the structural integrity of the inner limiting membrane (ILM) in the developing retina. In the absence of functional dystroglycan, ILM degeneration leads to defective migration, axon guidance, and mosaic spacing of neurons and a loss of multiple neuron types during retinal development. These results demonstrate that disorganization of retinal circuit development is a likely contributor to visual dysfunction in patients with dystroglycanopathy.
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http://dx.doi.org/10.1523/JNEUROSCI.0946-17.2017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5577860PMC
August 2017

Astrocytes follow ganglion cell axons to establish an angiogenic template during retinal development.

Glia 2017 10 19;65(10):1697-1716. Epub 2017 Jul 19.

Department of Neurobiology, Duke University School of Medicine, Durham, North Carolina, 27710.

Immature astrocytes and blood vessels enter the developing mammalian retina at the optic nerve head and migrate peripherally to colonize the entire retinal nerve fiber layer (RNFL). Retinal vascularization is arrested in retinopathy of prematurity (ROP), a major cause of bilateral blindness in children. Despite their importance in normal development and ROP, the factors that control vascularization of the retina remain poorly understood. Because astrocytes form a reticular network that appears to provide a substrate for migrating endothelial cells, they have long been proposed to guide angiogenesis. However, whether astrocytes do in fact impose a spatial pattern on developing vessels remains unclear, and how astrocytes themselves are guided is unknown. Here we explore the cellular mechanisms that ensure complete retinal coverage by astrocytes and blood vessels in mouse. We find that migrating astrocytes associate closely with the axons of retinal ganglion cells (RGCs), their neighbors in the RNFL. Analysis of Robo1; Robo2 mutants, in which RGC axon guidance is disrupted, and Math5 (Atoh7) mutants, which lack RGCs, reveals that RGCs provide directional information to migrating astrocytes that sets them on a centrifugal trajectory. Without this guidance, astrocytes exhibit polarization defects, fail to colonize the peripheral retina, and display abnormal fine-scale spatial patterning. Furthermore, using cell type-specific chemical-genetic tools to selectively ablate astrocytes, we show that the astrocyte template is required for angiogenesis and vessel patterning. Our results are consistent with a model whereby RGC axons guide formation of an astrocytic network that subsequently directs vessel development.
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http://dx.doi.org/10.1002/glia.23189DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5561467PMC
October 2017

Selection on meiosis genes in diploid and tetraploid Arabidopsis arenosa.

Mol Biol Evol 2015 Apr 26;32(4):944-55. Epub 2014 Dec 26.

Department of Evolutionary and Organismic Biology, Harvard University

Meiotic chromosome segregation is critical for fertility across eukaryotes, and core meiotic processes are well conserved even between kingdoms. Nevertheless, recent work in animals has shown that at least some meiosis genes are highly diverse or strongly differentiated among populations. What drives this remains largely unknown. We previously showed that autotetraploid Arabidopsis arenosa evolved stable meiosis, likely through reduced crossover rates, and that associated with this there is strong evidence for selection in a subset of meiosis genes known to affect axis formation, synapsis, and crossover frequency. Here, we use genome-wide data to study the molecular evolution of 70 meiosis genes in a much wider sample of A. arenosa. We sample the polyploid lineage, a diploid lineage from the Carpathian Mountains, and a more distantly related diploid lineage from the adjacent, but biogeographically distinct Pannonian Basin. We find that not only did selection act on meiosis genes in the polyploid lineage but also independently on a smaller subset of meiosis genes in Pannonian diploids. Functionally related genes are targeted by selection in these distinct contexts, and in two cases, independent sweeps occurred in the same loci. The tetraploid lineage has sustained selection on more genes, has more amino acid changes in each, and these more often affect conserved or potentially functional sites. We hypothesize that Pannonian diploid and tetraploid A. arenosa experienced selection on structural proteins that mediate sister chromatid cohesion, the formation of meiotic chromosome axes, and synapsis, likely for different underlying reasons.
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http://dx.doi.org/10.1093/molbev/msu398DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4379401PMC
April 2015

The glucuronyltransferase B4GAT1 is required for initiation of LARGE-mediated α-dystroglycan functional glycosylation.

Elife 2014 Oct 3;3. Epub 2014 Oct 3.

Department of Molecular Physiology and Biophysics, University of Iowa, Carver College of Medicine, Iowa City, United States.

Dystroglycan is a cell membrane receptor that organizes the basement membrane by binding ligands in the extracellular matrix. Proper glycosylation of the α-dystroglycan (α-DG) subunit is essential for these activities, and lack thereof results in neuromuscular disease. Currently, neither the glycan synthesis pathway nor the roles of many known or putative glycosyltransferases that are essential for this process are well understood. Here we show that FKRP, FKTN, TMEM5 and B4GAT1 (formerly known as B3GNT1) localize to the Golgi and contribute to the O-mannosyl post-phosphorylation modification of α-DG. Moreover, we assigned B4GAT1 a function as a xylose β1,4-glucuronyltransferase. Nuclear magnetic resonance studies confirmed that a glucuronic acid β1,4-xylose disaccharide synthesized by B4GAT1 acts as an acceptor primer that can be elongated by LARGE with the ligand-binding heteropolysaccharide. Our findings greatly broaden the understanding of α-DG glycosylation and provide mechanistic insight into why mutations in B4GAT1 disrupt dystroglycan function and cause disease.
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http://dx.doi.org/10.7554/eLife.03941DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4227050PMC
October 2014

Divergent population structure and climate associations of a chromosomal inversion polymorphism across the Mimulus guttatus species complex.

Mol Ecol 2014 Jun;23(11):2844-60

Department of Biology, Duke University, PO Box 90338, Durham, NC 27708, USA.

Chromosomal rearrangement polymorphisms are common and increasingly found to be associated with adaptive ecological divergence and speciation. Rearrangements, such as inversions, reduce recombination in heterozygous individuals and thus can protect favourable allelic combinations at linked loci, facilitating their spread in the presence of gene flow. Recently, we identified a chromosomal inversion polymorphism that contributes to ecological adaptation and reproductive isolation between annual and perennial ecotypes of the yellow monkeyflower, Mimulus guttatus. Here we evaluate the population genetic structure of this inverted region in comparison with the collinear regions of the genome across the M. guttatus species complex. We tested whether annual and perennial M. guttatus exhibit different patterns of divergence for loci in the inverted and noninverted regions of the genome. We then evaluated whether there are contrasting climate associations with these genomic regions through redundancy analysis. We found that the inversion exhibits broadly different patterns of divergence among annual and perennial M. guttatus and is associated with environmental variation across population accessions. This study is the first widespread population genetic survey of the diversity of the M. guttatus species complex. Our findings contribute to a greater understanding of morphological, ecological, and genetic evolutionary divergence across this highly diverse group of closely related ecotypes and species. Finally, understanding species relationships among M. guttatus sp. has hitherto been stymied by accumulated evidence of substantial gene flow among populations as well as designated species. Nevertheless, our results shed light on these relationships and provide insight into adaptation in life history traits within the complex.
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http://dx.doi.org/10.1111/mec.12778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4111140PMC
June 2014

Cytological techniques to analyze meiosis in Arabidopsis arenosa for investigating adaptation to polyploidy.

Front Plant Sci 2014 3;4:546. Epub 2014 Jan 3.

School of Biosciences, The University of Birmingham Birmingham, UK.

Arabidopsis arenosa is a close relative of the model plant A. thaliana, and exists in nature as stable diploid and autotetraploid populations. Natural tetraploids have adapted to whole genome duplication and do not commonly show meiotic errors such as multivalent and univalent formation, which can lead to chromosome non-disjunction and reduced fertility. A genome scan for genes strongly differentiated between diploid and autotetraploid A. arenosa identified a subset of meiotic genes that may be responsible for adaptation to polyploid meiosis. To investigate the mechanisms by which A. arenosa adapted to its polyploid state, and the functionality of the identified potentially adaptive polymorphisms, a thorough cytological analysis is required. Therefore, in this chapter we describe methods and techniques to analyze male meiosis in A. arenosa, including optimum plant growth conditions, and immunocytological and cytological approaches developed with the specific purpose of understanding meiotic adaptation in an autotetraploid. In addition we present a meiotic cytological atlas to be used as a reference for particular stages and discuss observations arising from a comparison of meiosis between diploid and autotetraploid A. arenosa.
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http://dx.doi.org/10.3389/fpls.2013.00546DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3879461PMC
January 2014

Fine-scale variation in meiotic recombination in Mimulus inferred from population shotgun sequencing.

Proc Natl Acad Sci U S A 2013 Nov 13;110(48):19478-82. Epub 2013 Nov 13.

Department of Energy Joint Genome Institute, Walnut Creek, CA 94598.

Meiotic recombination rates can vary widely across genomes, with hotspots of intense activity interspersed among cold regions. In yeast, hotspots tend to occur in promoter regions of genes, whereas in humans and mice, hotspots are largely defined by binding sites of the positive-regulatory domain zinc finger protein 9. To investigate the detailed recombination pattern in a flowering plant, we use shotgun resequencing of a wild population of the monkeyflower Mimulus guttatus to precisely locate over 400,000 boundaries of historic crossovers or gene conversion tracts. Their distribution defines some 13,000 hotspots of varying strengths, interspersed with cold regions of undetectably low recombination. Average recombination rates peak near starts of genes and fall off sharply, exhibiting polarity. Within genes, recombination tracts are more likely to terminate in exons than in introns. The general pattern is similar to that observed in yeast, as well as in positive-regulatory domain zinc finger protein 9-knockout mice, suggesting that recombination initiation described here in Mimulus may reflect ancient and conserved eukaryotic mechanisms.
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http://dx.doi.org/10.1073/pnas.1319032110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3845195PMC
November 2013

Meiotic adaptation to genome duplication in Arabidopsis arenosa.

Curr Biol 2013 Nov 17;23(21):2151-6. Epub 2013 Oct 17.

Department of Organismic and Evolutionary Biology, Harvard University, 16 Divinity Ave., Cambridge, MA, 02138, USA.

Whole genome duplication (WGD) is a major factor in the evolution of multicellular eukaryotes, yet by doubling the number of homologs, WGD severely challenges reliable chromosome segregation, a process conserved across kingdoms. Despite this, numerous genome-duplicated (polyploid) species persist in nature, indicating early problems can be overcome. Little is known about which genes are involved--only one has been molecularly characterized. To gain new insights into the molecular basis of adaptation to polyploidy, we investigated genome-wide patterns of differentiation between natural diploids and tetraploids of Arabidopsis arenosa, an outcrossing relative of A. thaliana. We first show that diploids are not preadapted to polyploid meiosis. We then use a genome scanning approach to show that although polymorphism is extensively shared across ploidy levels, there is strong ploidy-specific differentiation in 39 regions spanning 44 genes. These are discrete, mostly single-gene peaks of sharply elevated differentiation. Among these peaks are eight meiosis genes whose encoded proteins coordinate a specific subset of early meiotic functions, suggesting these genes comprise a polygenic solution to WGD-associated chromosome segregation challenges. Our findings indicate that even conserved meiotic processes can be capable of nimble evolutionary shifts when required.
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http://dx.doi.org/10.1016/j.cub.2013.08.059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3859316PMC
November 2013

Evolutionary genetics: inheritance of a complex pollination syndrome.

Curr Biol 2013 Jun;23(12):R525-7

Department of Organismic and Evolutionary Biology, Harvard University, 16 Divinity Avenue, Cambridge, MA, USA.

How adaptive traits that are controlled by multiple genes evolve is an intriguing question in evolutionary genetics. A recent study shows that tight linkage allows genes that contribute to a multitrait pollination syndrome to be inherited together as a unit.
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http://dx.doi.org/10.1016/j.cub.2013.04.069DOI Listing
June 2013

Indirect evolution of hybrid lethality due to linkage with selected locus in Mimulus guttatus.

PLoS Biol 2013 26;11(2):e1001497. Epub 2013 Feb 26.

Department of Biology, Duke University, Durham, North Carolina, USA.

Most species are superbly and intricately adapted to the environments in which they live. Adaptive evolution by natural selection is the primary force shaping biological diversity. Differences between closely related species in ecologically selected characters such as habitat preference, reproductive timing, courtship behavior, or pollinator attraction may prevent interbreeding in nature, causing reproductive isolation. But does ecological adaptation cause reproductive incompatibilities such as hybrid sterility or lethality? Although several genes causing hybrid incompatibilities have been identified, there is intense debate over whether the genes that contribute to ecological adaptations also cause hybrid incompatibilities. Thirty years ago, a genetic study of local adaptation to copper mine soils in the wildflower Mimulus guttatus identified a locus that appeared to cause copper tolerance and hybrid lethality in crosses to other populations. But do copper tolerance and hybrid lethality have the same molecular genetic basis? Here we show, using high-resolution genome mapping, that copper tolerance and hybrid lethality are not caused by the same gene but are in fact separately controlled by two tightly linked loci. We further show that selection on the copper tolerance locus indirectly caused the hybrid incompatibility allele to go to high frequency in the copper mine population because of hitchhiking. Our results provide a new twist on Darwin's original supposition that hybrid incompatibilities evolve as an incidental by-product of ordinary adaptation to the environment.
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http://dx.doi.org/10.1371/journal.pbio.1001497DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3582499PMC
August 2013

Dystroglycan organizes axon guidance cue localization and axonal pathfinding.

Neuron 2012 Dec;76(5):931-44

The Solomon H. Snyder Department of Neuroscience and Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

Precise patterning of axon guidance cue distribution is critical for nervous system development. Using a murine forward genetic screen for novel determinants of axon guidance, we identified B3gnt1 and ISPD as required for the glycosylation of dystroglycan in vivo. Analysis of B3gnt1, ISPD, and dystroglycan mutant mice revealed a critical role for glycosylated dystroglycan in the development of several longitudinal axon tracts. Remarkably, the axonal guidance defects observed in B3gnt1, ISPD, and dystroglycan mutants resemble several of the axon guidance defects found in mice lacking the axon guidance cue Slit and its receptor Robo. This similarity is explained by our observations that dystroglycan binds directly to Slit and is required for proper Slit localization within the basement membrane and floor plate in vivo. These findings establish a novel role for glycosylated dystroglycan as a key determinant of axon guidance cue distribution and function in the mammalian nervous system.
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http://dx.doi.org/10.1016/j.neuron.2012.10.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3526105PMC
December 2012