SLIT2/ROBO2 signaling pathway inhibits nonmuscle myosin IIA activity and destabilizes kidney podocyte adhesion

Abstract

The repulsive guidance cue SLIT2 and its receptor ROBO2 are required for kidney development and podocyte foot process structure, but the SLIT2/ROBO2 signaling mechanism regulating podocyte function is not known. Here we report that a potentially novel signaling pathway consisting of SLIT/ROBO Rho GTPase activating protein 1 (SRGAP1) and nonmuscle myosin IIA (NMIIA) regulates podocyte adhesion downstream of ROBO2. We found that the myosin II regulatory light chain (MRLC), a subunit of NMIIA, interacts directly with SRGAP1 and forms a complex with ROBO2/SRGAP1/NMIIA in the presence of SLIT2. Immunostaining demonstrated that SRGAP1 is a podocyte protein and is colocalized with ROBO2 on the basal surface of podocytes. In addition, SLIT2 stimulation inhibits NMIIA activity, decreases focal adhesion formation, and reduces podocyte attachment to collagen. In vivo studies further showed that podocyte-specific knockout of protects mice from hypertension-induced podocyte detachment and albuminuria and also partially rescues the podocyte-loss phenotype in knockout mice. Thus, we have identified SLIT2/ROBO2/SRGAP1/NMIIA as a potentially novel signaling pathway in kidney podocytes, which may play a role in regulating podocyte adhesion and attachment. Our findings also suggest that SLIT2/ROBO2 signaling might be a therapeutic target for kidney diseases associated with podocyte detachment and loss.

Dissection of Glomerular Transcriptional Profile in Patients With Diabetic Nephropathy: SRGAP2a Protects Podocyte Structure and Function

“…Given that the alteration of cellular F-actin by SRGAP2 is through its interaction with the Rho family of GTPases, these results suggest that the interactions of SRGAP2a with Cdc42, RhoA, or Rac1 and their functions may vary in different cell types and under different pathological conditions. Studies by Fan and colleagues (52,53) showed that SRGAP1, another SRGAP family member, is expressed on the basal surface of podocytes and promotes podocyte detachment through interacting with myosin II regulatory light chain. Different from the function of SRGAP2a that we observed to stabilize podocyte F-actin fibers, they showed that SRGAP1 actually decreased polymerization of F-actin.…”

Disruption of Robo2-Baiap2 integrated signaling drives cystic disease

“…To assess the effects of Slit2 and of silencing of Robo2 on gene expression, we starved MDCK cells overnight in DMEM/F-12 (Invitrogen) supplemented with 0.1% FBS, and cells were treated with ROBO2 siR-NAs and then with 50 nM recombinant human SLIT2 N-terminus (MilliporeSigma, SRP3155) or buffer for 24 hours as previously described (59,60). After the media were removed, cells were lysed for Western blotting.…”

Proteinuric Kidney Diseases: A Podocyte's Slit Diaphragm and Cytoskeleton Approach

“…However, there is much more to discover. Novel therapeutic target genes or proteins in podocytes are constantly being discovered as we write this review ( 269 – 271 ). Longstanding concepts, such as the function of the SD and the lack of mitotic ability of the podocyte, are already being challenged ( 272 – 274 ).…”