Gα12 Activation in Podocytes Leads to Cumulative Changes in Glomerular Collagen Expression, Proteinuria and Glomerulosclerosis

Ilene Boucher; Wanfeng Yu; Sarah Beaudry; Hideyuki Negoro; Mei Tran; Martin R Pollak; Joel M Henderson; Bradley M Denker


Lab Invest. 2012;92(5):662-675. 

In This Article

Abstract and Introduction


Glomerulosclerosis is a common pathological finding that often progresses to renal failure. The mechanisms of chronic kidney disease progression are not well defined, but may include activation of numerous vasoactive and inflammatory pathways. We hypothesized that podocytes are susceptible to filtered plasma components, including hormones and growth factors that stimulate signaling pathways leading to glomerulosclerosis. Gα12 couples to numerous G-protein-coupled receptors (GPCRs) and regulates multiple epithelial responses, including proliferation, apoptosis, permeability and the actin cytoskeleton. Herein, we report that genetic activation of Gα12 in podocytes leads to time-dependent increases in proteinuria and glomerulosclerosis. To mimic activation of Gα12 pathways, constitutively active Gα12 (QL) was conditionally expressed in podocytes using Nphs2-Cre and LacZ/floxed QLα12 transgenic mice. Some QLα12LacZ+/Cre+ mice developed proteinuria at 4–6 months, and most were proteinuric by 12 months. Proteinuria increased with age, and by 12–14 months, many demonstrated glomerulosclerosis with ultrastructural changes, including foot process fusion and both mesangial and subendothelial deposits. QLα12LacZ+/Cre+ mice showed no changes in podocyte number, apoptosis, proliferation or Rho/Src activation. Real-time PCR revealed no significant changes in Nphs1, Nphs2, Cd2ap or Trpc6 expression, but Col4a2 message was increased in younger and older mice, while Col4a5 was decreased in older mice. Confocal microscopy revealed disordered collagen IVα1/2 staining in older mice and loss of α5 without changes in other collagen IV subunits. Taken together, these studies suggest that Gα12 activation promotes glomerular injury without podocyte depletion through a novel mechanism regulating collagen (α)IV expression, and supports the notion that glomerular damage may accrue through persistent GPCR activation in podocytes.


Glomerulosclerosis (GS) is a common pathological finding in patients with progressive chronic kidney disease (CKD) and often leads to end-stage renal disease. Numerous conditions predispose patients to GS, including diabetes, hypertension, IgA nephropathy, FSGS (focal segmental GS) and immune-mediated injury. In adults over 60 years old, the prevalence of CKD Stage III (glomerular filtration rate, 30–59 ml/min) is estimated to be >25%.[1] Although risk factors such as hypertension and diabetes are linked to CKD, little is known about the signaling mechanisms that lead to progression with aging. Post-mortem and nephrectomy samples in otherwise 'healthy' adults reveal variable amounts of GS and interstitial fibrosis, suggestive of age-associated damage.[2,3] Recent studies show that primary podocyte injury is sufficient to induce GS.[4,5] Podocytes are exposed to filtered reactive oxygen species (ROS), lipid mediators, cytokines and hormones that could contribute to injury. Many of these molecules activate G-protein-coupled receptors (GPCRs), which couple to multiple Gα subunits. Each of the 16 Gα subunits (four main families: Gαs, Gαi/o, Gαq and Gα12/13) couples to many different GPCRs;[6] thus, defining specific pathways in vivo has been difficult.

Gα12/13 are expressed in podocytes and couple to angiotensin II, thrombin, endothelin and LPA receptors, which are important in renal injury.[7] Gα12/13 can activate Rho or Src to regulate the actin cytoskeleton,[8] in addition to proliferation, transformation,[9] tight junction assembly,[10–12] cell–cell adhesion,[13,14] directed cell migration,[15] apoptosis[16] and cell attachment.[17] RhoGDIα knockout mice develop proteinuria and renal failure,[18] and many mutations in hereditary FSGS affect proteins linked to the actin cytoskeleton (reviewed in ref.[19]). Gα12 also upregulates TGFβ,[20,21] and several gene profiling studies found upregulated Gα12 in proteinuric kidneys and post-transplant CKD (via Nephromine[22,23]).

Targeting activated Gα subunits to specific cells in vivo permits identification of downstream effector pathways independent of receptor activation, and thus permits insight into disease mechanisms otherwise impossible to study in vivo. Herein, we confirm the expression of endogenous Gα12 in the major podocyte processes. Constitutively activated Gα12 (QLα12) was expressed in podocytes using a transgenic model that results in mosaic expression and mimics the focal nature of GS pathology. QLα12LacZ+/Cre+ mice develop proteinuria and focal GS without differences in podocyte number, apoptosis, proliferation or Rho/Src signaling over time. Col4a was dysregulated and correlated with altered localization and ultrastructural changes. These findings indicate that Gα12 activation in podocytes leads to dysregulated collagen α(IV) expression and support a model of altered glomerular structure and function resulting from time-dependent stimulation of GPCR-Gα12 signaling pathways.


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