Robo2 is the cell surface receptor for the repulsive guidance cue Slit and is involved in axon guidance and neuronal migration. Nephrin is a podocyte slit-diaphragm protein that functions in the ...kidney glomerular filtration barrier. Here, we report that Robo2 is expressed at the basal surface of mouse podocytes and colocalizes with nephrin. Biochemical studies indicate that Robo2 forms a complex with nephrin in the kidney through adaptor protein Nck. In contrast to the role of nephrin that promotes actin polymerization, Slit2-Robo2 signaling inhibits nephrin-induced actin polymerization. In addition, the amount of F-actin associated with nephrin is increased in Robo2 knockout mice that develop an altered podocyte foot process structure. Genetic interaction study further reveals that loss of Robo2 alleviates the abnormal podocyte structural phenotype in nephrin null mice. These results suggest that Robo2 signaling acts as a negative regulator on nephrin to influence podocyte foot process architecture.
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► Robo2 colocalizes with nephrin in kidney podocytes ► Robo2 forms a complex with nephrin through the Nck protein ► Slit2-Robo2 signaling inhibits nephrin-induced actin polymerization ► Robo2 loss alleviates podocyte structural phenotype in nephrin mutant mice
The unique interdigitating foot processes of kidney podocytes, together with intervening slit diaphragms, confer selective permeability to the glomerular filtration barrier. Nephrin, a major slit-diaphragm component, regulates permeability and promotes actin polymerization. Lu and colleagues now identify the neuronal guidance cue receptor Robo2 as a podocyte protein that forms a complex with nephrin through the adaptor protein Nck. Their study suggests that Robo2 signaling acts as a negative regulator of nephrin-induced actin polymerization, thereby modulating podocyte foot process structure.
This Review summarizes recent research on the podocyte slit diaphragm. A growing number of molecules that function at the slit diaphragm have been identified in patients with inherited and sporadic ...nephrotic syndromes. Genetic deletion of nearly all of these molecules results in proteinuria and effacement of foot processes. Nephrin, Neph1 and podocin seem to form a multifunctional receptor complex at the slit diaphragm. Most of the other components of the slit diaphragm interact directly with this complex, in many cases coupling slit diaphragm components to the podocyte's actin cytoskeleton. These molecular findings are being applied to patients with glomerular disease. Over the next decade, these data might help to improve disease classification and prediction of which patients will respond to immunosuppressive treatment.
Despite growing interest in species' range shifts, little is known about the ecological and evolutionary factors that control geographic range boundaries. We investigated the processes that maintain ...the northern range limit of the mud fiddler crab (Uca pugnax) at North Scituate, Massachusetts, USA (42°14′ N), located ∼60 km north of Cape Cod. Larvae from five populations in Massachusetts were reared under controlled temperatures to test whether cooler water near the edge of this species' range inhibits planktonic development. Few larvae completed development at temperatures <18°C, a threshold that larvae would regularly encounter north of Cape Cod. Extensive salt marshes are present north of the current range boundary, and a transplant experiment using field enclosures confirmed that benthic fiddler crabs can survive severe winter conditions in this northern habitat. Taken with oceanographic data, these results suggest that the range boundary of fiddler crabs is likely maintained by the influence of cooler water temperatures on the larval phase. Analyses of mitochondrial DNA sequences from a neutral marker (COI) indicate high gene flow among U. pugnax populations in Massachusetts with little differentiation across Cape Cod. Consistent with predictions regarding the homogenizing influence of gene flow, larvae from source populations north and south of Cape Cod shared a common lower threshold for development. However, larvae produced near the range edge had faster growth rates than those from the south side of Cape Cod (typically reaching the final megalopal stage 1.0-5.5 d faster at 18°C). Additional studies are needed to determine the mechanism underlying this counter-gradient variation in development time. We hypothesize that dispersal into cooler water on the north side of Cape Cod may act as a selection filter that sieves out slow developers from the larval pool by increasing planktonic duration and exposure to associated sources of mortality. Thus while high gene flow may prevent the evolution of greater cold tolerance in northern populations, recurrent selection on existing variation may lead to an unexpected concentration of favorable adaptations at the edge of the range. Such a pattern could permit edge populations to play a dominant and unrecognized role in future range extensions.
Male germ cells are connected by intercellular bridges (ICBs) in a syncytium due to incomplete cytokinesis. Syncytium is thought to be important for synchronized germ cell development by interchange ...of cytoplasmic factors via ICBs. Mammalian ADP-ribosylation factor 6 (ARF6) is a small GTPase that is involved in many cellular mechanisms including but not limited to regulating cellular structure, motility, vesicle trafficking and cytokinesis. ARF6 localizes to ICBs in spermatogonia and spermatocytes in mice. Here we report that mice with global depletion of ARF6 in adulthood using Ubc-CreERT2 display no observable phenotypes but are male sterile. ARF6-deficient males display a progressive loss of germ cells, including LIN28A-expressing spermatogonia, and ultimately develop Sertoli-cell-only syndrome. Specifically, intercellular bridges are lost in ARF6-deficient testis. Furthermore, germ cell-specific inactivation using the Ddx4-CreERT2 results in the same testicular morphological phenotype, showing the germ cell-intrinsic requirement of ARF6. Therefore, ARF6 is essential for spermatogenesis in mice and this function is conserved from Drosophila to mammals.
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•Arf6 null adult male mice were infertile, exhibiting a Sertoli-cell-only phenotype.•Germline-specific Arf6 deletion results in similar progressive loss of germ cells.•ARF6 is a component of intercellular bridges in male germ cells.•Arf6 deletion resulted in disruption of germ cell intercellular bridges.•ARF6 plays an essential germ-cell-autonomous role in spermatogenesis.
This study describes a quality improvement (QI) process to reduce bias and increase inclusion and equity in the recruitment of health service psychology interns in an American Psychological ...Association-accredited psychology internship program at a national children's hospital.
This QI project utilized two Plan-Do-Study-Act (PDSA) cycles targeting the application review and the interview processes primarily using supervisor engagement and feedback to inform these processes. The goal of the PDSA cycles was to increase diversity in psychology doctoral interns offered interviews and ultimately recruited to the internship program.
The application rating form was revised to place a greater emphasis on factors related to diversity, such as increasing the number of points applicants could earn for being bilingual. Regarding the interview process, structured interview questions were created, and a new, unified rubric was used to score interviewees. The changes in demographics of applicants selected for interviews and feedback from applicants who interviewed are reported.
The QI process resulted in tangible changes to improve equitable and inclusive internship recruitment. Lessons learned throughout this process included the need for continual auditing of practices through an equity lens, engaging supervisors at all stages of the process, and implementing incremental actions.
Glomerular injury is often characterized by the effacement of podocytes, loss of slit diaphragms, and proteinuria. Renal ischemia or the loss of blood flow to the kidneys has been widely associated ...with tubular and endothelial injury but rarely has been shown to induce podocyte damage and disruption of the slit diaphragm. In this study, we have used an in vivo rat ischemic model to demonstrate that renal ischemia induces podocyte effacement with loss of slit diaphragm and proteinuria. Biochemical analysis of the ischemic glomerulus shows that ischemia induces rapid loss of interaction between slit diaphragm junctional proteins Neph1 and ZO-1. To further understand the effect of ischemia on molecular interactions between slit diaphragm proteins, a cell culture model was employed to study the binding between Neph1 and ZO-1. Under physiologic conditions, Neph1 co-localized with ZO-1 at cell-cell contacts in cultured human podocytes. Induction of injury by ATP depletion resulted in rapid loss of Neph1 and ZO-1 binding and redistribution of Neph1 and ZO-1 proteins from cell membrane to the cytoplasm. Recovery resulted in increased Neph1 tyrosine phosphorylation, restoring Neph1 and ZO-1 binding and their localization at the cell membrane. We further demonstrate that tyrosine phosphorylation of Neph1 mediated by Fyn results in significantly increased Neph1 and ZO-1 binding, suggesting a critical role for Neph1 tyrosine phosphorylation in reorganizing the Neph1-ZO-1 complex. This study documents that renal ischemia induces dynamic changes in the molecular interactions between slit diaphragm proteins, leading to podocyte damage and proteinuria.
Alterations in glomerular podocyte cell-cell and cell-matrix contacts are key events in progressive glomerular failure. Integrin-linked kinase (ILK) has been implicated in podocyte cell-matrix ...interaction and is induced in proteinuria. For evaluation of ILK function in vivo, mice with a Cre-mediated podocyte-specific ILK inactivation were generated. These mice seemed normal at birth but developed progressive focal segmental glomerulosclerosis and died in terminal renal failure. The first ultrastructural lesions that are seen at onset of albuminuria are glomerular basement membrane (GBM) alterations with a significant increase in true harmonic mean GBM thickness. Podocyte foot process effacement and loss of slit diaphragm followed with progression to unselective proteinuria. No significant reduction of slit membrane molecules (podocin and nephrin), key GBM components (fibronectin, laminins, and collagen IV isoforms), or podocyte integrins could be observed at onset of proteinuria. However, alpha3-integrins were relocalized into a granular pattern along the GBM, consistent with altered integrin-mediated matrix assembly in ILK-deficient podocytes. As the increased GBM thickness precedes structural podocyte lesions and key components of the GBM were expressed at comparable levels to controls, these data suggest an essential role of ILK for the close interconnection of GBM structure and podocyte function.
The 14th International Podocyte Conference took place in Philadelphia, Pennsylvania, USA from May 23 to 26, 2023. It commenced with an early-career researchers' meeting on May 23, providing young ...scientists with a platform to present and discuss their research findings. Throughout the main conference, 29 speakers across 9 sessions shared their insights on podocyte biology, glomerular medicine, novel technologic advancements, and translational approaches. Additionally, the event featured 3 keynote lectures addressing engineered chimeric antigen receptor T cell– and mRNA-based therapies and the use of biobanks for enhanced disease comprehension. Furthermore, 4 brief oral abstract sessions allowed scientists to present their findings to a broad audience. The program also included a panel discussion addressing the challenges of conducting human research within the American Black community. Remarkably, after a 5-year hiatus from in-person conferences, the 14th International Podocyte Conference successfully convened scientists from around the globe, fostering the presentation and discussion of crucial research findings, as summarized in this review. Furthermore, to ensure continuous and sustainable education, research, translation, and trial medicine related to podocyte and glomerular diseases for the benefit of patients, the International Society of Glomerular Disease was officially launched during the conference.
NPHS2 was recently identified as a gene whose mutations cause autosomal recessive steroid-resistant nephrotic syndrome. Its product, podocin, is a new member of the stomatin family, which consists of ...hairpin-like integral membrane proteins with intracellular NH(2)- and COOH-termini. Podocin is expressed in glomerular podocytes, but its subcellular distribution and interaction with other proteins are unknown. Here we show, by immunoelectron microscopy, that podocin localizes to the podocyte foot process membrane, at the insertion site of the slit diaphragm. Podocin accumulates in an oligomeric form in lipid rafts of the slit diaphragm. Moreover, GST pull-down experiments reveal that podocin associates via its COOH-terminal domain with CD2AP, a cytoplasmic binding partner of nephrin, and with nephrin itself. That podocin interacts with CD2AP and nephrin in vivo is shown by coimmunoprecipitation of these proteins from glomerular extracts. Furthermore, in vitro studies reveal direct interaction of podocin and CD2AP. Hence, as with the erythrocyte lipid raft protein stomatin, podocin is present in high-order oligomers and may serve a scaffolding function. We postulate that podocin serves in the structural organization of the slit diaphragm and the regulation of its filtration function.
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