MAGI-1 Interacts with Nephrin to Maintain Slit Diaphragm Structure through Enhanced Rap1 Activation in Podocytes

TY - JOUR

T1 - MAGI-1 Interacts with Nephrin to Maintain Slit Diaphragm Structure through Enhanced Rap1 Activation in Podocytes

AU - Ni, Jie

AU - Bao, Sujin

AU - Johnson, Ruth I

AU - Zhu, Bingbing

AU - Li, Jianhua

AU - Vadaparampil, Justin

AU - Smith, Christopher M

AU - Campbell, Kirk N

AU - Grahammer, Florian

AU - Huber, Tobias B

AU - He, John C

AU - D'Agati, Vivette D

AU - Chan, Andrew

AU - Kaufman, Lewis

N1 - © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

PY - 2016/11/18

Y1 - 2016/11/18

N2 - MAGI-1 is a multidomain cytosolic scaffolding protein that in the kidney is specifically located at the podocyte slit diaphragm, a specialized junction that is universally injured in proteinuric diseases. There it interacts with several essential molecules, including nephrin and neph1, which are required for slit diaphragm formation and as an intracellular signaling hub. Here, we show that diminished MAGI-1 expression in cultured podocytes reduced nephrin and neph1 membrane localization and weakened tight junction integrity. Global magi1 knock-out mice, however, demonstrated normal glomerular histology and function into adulthood. We hypothesized that a second mild but complementary genetic insult might induce glomerular disease susceptibility in these mice. To identify such a gene, we utilized the developing fly eye to test for functional complementation between MAGI and its binding partners. In this way, we identified diminished expression of fly Hibris (nephrin) or Roughest (neph1) as dramatically exacerbating the effects of MAGI depletion. Indeed, when these combinations were studied in mice, the addition of nephrin, but not neph1, heterozygosity to homozygous deletion of MAGI-1 resulted in spontaneous glomerulosclerosis. In cultured podocytes, MAGI-1 depletion reduced intercellular contact-induced Rap1 activation, a pathway critical for proper podocyte function. Similarly, magi1 knock-out mice showed diminished glomerular Rap1 activation, an effect dramatically enhanced by concomitant nephrin haploinsufficiency. Finally, combined overexpression of MAGI-1 and nephrin increased Rap1 activation, but not when substituting a mutant MAGI-1 that cannot bind nephrin. We conclude that the interaction between nephrin and MAGI-1 regulates Rap1 activation in podocytes to maintain long term slit diaphragm structure.

AB - MAGI-1 is a multidomain cytosolic scaffolding protein that in the kidney is specifically located at the podocyte slit diaphragm, a specialized junction that is universally injured in proteinuric diseases. There it interacts with several essential molecules, including nephrin and neph1, which are required for slit diaphragm formation and as an intracellular signaling hub. Here, we show that diminished MAGI-1 expression in cultured podocytes reduced nephrin and neph1 membrane localization and weakened tight junction integrity. Global magi1 knock-out mice, however, demonstrated normal glomerular histology and function into adulthood. We hypothesized that a second mild but complementary genetic insult might induce glomerular disease susceptibility in these mice. To identify such a gene, we utilized the developing fly eye to test for functional complementation between MAGI and its binding partners. In this way, we identified diminished expression of fly Hibris (nephrin) or Roughest (neph1) as dramatically exacerbating the effects of MAGI depletion. Indeed, when these combinations were studied in mice, the addition of nephrin, but not neph1, heterozygosity to homozygous deletion of MAGI-1 resulted in spontaneous glomerulosclerosis. In cultured podocytes, MAGI-1 depletion reduced intercellular contact-induced Rap1 activation, a pathway critical for proper podocyte function. Similarly, magi1 knock-out mice showed diminished glomerular Rap1 activation, an effect dramatically enhanced by concomitant nephrin haploinsufficiency. Finally, combined overexpression of MAGI-1 and nephrin increased Rap1 activation, but not when substituting a mutant MAGI-1 that cannot bind nephrin. We conclude that the interaction between nephrin and MAGI-1 regulates Rap1 activation in podocytes to maintain long term slit diaphragm structure.

KW - Adaptor Proteins, Signal Transducing

KW - Animals

KW - Enzyme Activation

KW - Glomerulosclerosis, Focal Segmental

KW - Membrane Proteins

KW - Mice

KW - Mice, Knockout

KW - Podocytes

KW - rap1 GTP-Binding Proteins

KW - Journal Article

U2 - 10.1074/jbc.M116.745026

DO - 10.1074/jbc.M116.745026

M3 - SCORING: Journal article

C2 - 27707879

VL - 291

SP - 24406

EP - 24417

JO - J BIOL CHEM

JF - J BIOL CHEM

SN - 0021-9258

IS - 47

ER -