Identification of direct negative cross-talk between the SLIT2 and bone morphogenetic protein-Gremlin signaling pathways

Slit guidance ligand 2 (SLIT2) is a large, secreted protein that binds roundabout (ROBO) receptors on multiple cell types, including neurons and kidney podocytes. S LIT2-ROBO mediated signaling regulates neuronal migration and ureteric bud (UB) outgrowth during kidney development as well as glomerular filtration in adult kidneys. Additionally, SLIT2 binds Gremlin, an antagonist of bone morphogenetic proteins (BMPs), and BMP Gremlin signaling also regulates UB formation. However, direct cross-talk between the ROBO2-SLIT2 and BMP Gremlin signaling pathways has not been established. Here, we report the discovery of negative feedback between the SLIT2 and BMP Gremlin signaling pathways. We found that the SLIT2 Gremlin interaction inhibited both SLIT2-ROBO2 signaling in neurons and Gremlin antagonism of BMP activity in myoblasts and fibroblasts. Furthermore, BMP2 down-regulated SLIT2 expression and promoter activity through canonical BMP signaling. Gremlin treatment, BMP receptor inhibition, and SMAD family member 4 (SMAD4) knockdown rescued BMPmediated repression of SLIT2. BMP2 treatment of nephron progenitor cells derived from human embryonic stem cells decreased SLIT2 expression, further suggesting an interaction between the BMP2 Gremlin and SLIT2 pathways in human kidney cells. In conclusion, our study has revealed direct negative cross-talk between two pathways, previously thought to be unassociated, that may regulate both kidney development and adult tissue maintenance.