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Our recent studies suggest that 1,25-dihydroxyvitamin D3 functions as an endocrine suppressor of renin biosynthesis. Genetic disruption of the vitamin D receptor (VDR) results in overstimulation of the renin-angiotensin system (RAS), leading to high blood pressure and cardiac hypertrophy. Consistent with the higher heart-to-body weight ratio, the size of left ventricular cardiomyocytes in VDR knockout (KO) mice was markedly increased compared with wild-type (WT) mice. As expected, levels of atrial natriuretic peptide (ANP) mRNA and circulating ANP were also increased in VDRKO mice. Treatment of VDRKO mice with captopril reduced cardiac hypertrophy and normalized ANP expression. To investigate the role of the cardiac RAS in the development of cardiac hypertrophy, the expression of renin, angiotensinogen, and AT-1a receptor in the heart was examined by real-time RT-PCR and immunostaining. In VDRKO mice, the cardiac renin mRNA level was significantly increased, and this increase was further amplified by captopril treatment. Consistently, intense immunostaining was detected in the left ventricle of captopril-treated WT and VDRKO mice by use of an anti-renin antibody. Levels of cardiac angiotensinogen and AT-1a receptor mRNAs were unchanged in the mutant mice. These data suggest that the cardiac hypertrophy seen in VDRKO mice is a consequence of activation of both the systemic and cardiac RAS and support the notion that 1,25-dihydroxyvitamin D(3) regulates cardiac functions, at least in part, through the RAS.

Previously, we showed that vitamin D receptor gene knockout leads to hyperreninemia independent of calcium metabolism; however, the contribution of parathyroid hormone to renin upregulation remained unclear. Here we separated the role of vitamin D and parathyroid hormone in the regulation of renin expression in vivo by generating transgenic mice that overexpressed the human vitamin D receptor in renin-producing cells using the 4.1 kb Ren-1c gene promoter. Targeting of human vitamin D receptor to the juxtaglomerular cells of the mice was confirmed by immunohistochemistry. Renal renin mRNA levels and plasma renin activity were decreased in these transgenic mice by about 50% and 30%, respectively, with no significant change in blood pressure, calcium, or parathyroid hormone levels. Moreover using vitamin D receptor knockout mice, we found that expression of the human receptor in their juxtaglomerular cells reduced renin expression in these mice without affecting calcium or parathyroid hormone status. Our study shows that suppression of renin expression by 1,25-dihydroxyvitamin D in vivo is independent of parathyroid hormone and calcium.

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Material Supplementary 9.DC1http://www.jimmunol.org/content/suppl/2010/07/06/jimmunol.090336

SUMMARY E3 ubiquitin ligase Cbl-b has emerged as a gatekeeper that controls the activation threshold of the T cell antigen receptor and maintains the balance between tolerance and autoimmunity. Here, we report that the loss of Cbl-b facilitates T helper 2 (Th2) and Th9 cell differentiation in vitro. In a mouse model of asthma, the absence of Cbl-b results in severe airway inflammation and stronger Th2 and Th9 responses. Mechanistically, Cbl-b selectively associates with Stat6 upon IL-4 ligation and targets Stat6 for ubiquitination and degradation. These processes are heightened in the presence of T cell receptor (TCR)/ CD28 costimulation. Furthermore, we identify K108 and K398 as Stat6 ubiquitination sites. Intriguingly, introducing Stat6 deficiency into Cblb−/− mice abrogates hyper-Th2 responses but only partially attenuates Th9 responses. Therefore, our data reveal a function for Cbl-b in the regulation of Th2 and Th9 cell differentiation.

Summary E3 ubiquitin ligase Cbl-b plays a crucial role in T cell activation and tolerance induction. However, the molecular mechanism by which Cbl-b inhibits T cell activation remains unclear. Here we report that Cbl-b does not inhibit PI3-K, but rather suppresses TCR/CD28-induced inactivation of Pten. The elevated Akt activity in Cbl-b−/− T cells is therefore due to heightened Pten inactivation. Suppression of Pten inactivation in T cells by Cbl-b is achieved by impeding the association of Pten with Nedd4, which targets Pten K13 for K63-linked polyubiquitination. Consistent with this finding, introducing Nedd4 deficiency into Cbl-b−/− mice abrogates hyper-T cell responses caused by the loss of Cbl-b. Hence, our data are the first to demonstrate that Cbl-b inhibits T cell activation by suppressing Pten inactivation independently of its ubiquitin ligase activity.

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