Gq signaling causes glomerular injury by activating TRPC6
Liming Wang, … , Michelle P. Winn, Robert F. Spurney
Liming Wang, … , Michelle P. Winn, Robert F. Spurney
Published April 6, 2015
Citation Information: J Clin Invest. 2015;125(5):1913-1926. https://doi.org/10.1172/JCI76767.
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Research Article Nephrology

Gq signaling causes glomerular injury by activating TRPC6

Abstract

Familial forms of focal segmental glomerulosclerosis (FSGS) have been linked to gain-of-function mutations in the gene encoding the transient receptor potential channel C6 (TRPC6). GPCRs coupled to Gq signaling activate TRPC6, suggesting that Gq-dependent TRPC6 activation underlies glomerular diseases. Here, we developed a murine model in which a constitutively active Gq α subunit (GqQ209L, referred to herein as GqQ>L) is specifically expressed in podocytes and examined the effects of this mutation in response to puromycin aminonucleoside (PAN) nephrosis. We found that compared with control animals, animals expressing GqQ>L exhibited robust albuminuria, structural features of FSGS, and reduced numbers of glomerular podocytes. Gq activation stimulated calcineurin (CN) activity, resulting in CN-dependent upregulation of TRPC6 in murine kidneys. Deletion of TRPC6 in GqQ>L-expressing mice prevented FSGS development and inhibited both tubular damage and podocyte loss induced by PAN nephrosis. Similarly, administration of the CN inhibitor FK506 reduced proteinuria and tubular injury but had more modest effects on glomerular pathology and podocyte numbers in animals with constitutive Gq activation. Moreover, these Gq-dependent effects on podocyte injury were generalizable to diabetic kidney disease, as expression of GqQ>L promoted albuminuria, mesangial expansion, and increased glomerular basement membrane width in diabetic mice. Together, these results suggest that targeting Gq/TRPC6 signaling may have therapeutic benefits for the treatment of glomerular diseases.

Authors

Liming Wang, Grant Jirka, Paul B. Rosenberg, Anne F. Buckley, Jose A. Gomez, Timothy A. Fields, Michelle P. Winn, Robert F. Spurney

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Figure 3

Induction of GqQ>L in podocytes enhances TRPC6 expression in vivo.

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Induction of GqQ>L in podocytes enhances TRPC6 expression in vivo. (A...
(A) Treatment with DOX enhanced Trpc6 mRNA levels in glomerular preparations from GqQ>L mice compared with levels detected in controls. (B) FK506 inhibited expression of Trpc6 mRNA in glomerular preparations from GqQ>L mice treated with DOX. (C and D) TRPC6 expression was similar in controls treated with DOX and in GqQ>L mice treated with sucrose. Treatment with DOX enhanced TRPC6 expression in GqQ>L mice compared with expression in either controls treated with DOX or in GqQ>L mice treated with sucrose. Induction of TRPC6 protein in GqQ>L mice treated with DOX was blocked by FK506. Actin was used as a loading control, and HEK293 cells transfected with a TRPC6 construct were used as a positive control for the immunoblotting studies. Because of the high levels of TRPC6 expression in the HEK293 cells, small amounts of the HEK293 cell lysate were used, and the actin loading control was not visible at this length of exposure. For the qRT-PCR analysis, 11 controls treated with DOX, 4 GqQ>L mice treated with DOX, 8 GqQ>L mice treated with DOX and vehicle, and 4 GqQ>L mice treated with DOX and FK506 were used. For the immunoblotting studies, 4 controls treated with DOX, 4 GqQ>L mice treated with sucrose, 4 GqQ>L mice treated with DOX, and 4 GqQ>L mice treated with DOX and FK506 were used. *P < 0.025 or †P < 0.01 versus the indicated groups using a 2-tailed t test for qRT-PCR data and ANOVA, followed by a Bonferroni’s post-hoc test, for densitometric data.