Regulation of aldosterone secretion by Ca(v)1.3

This work is supported by NIHR Senior Investigator grant NF-SI-0512-10052 awarded to M.J.B.; the Austin Doyle Award (Servier Australia) and the Tunku Abdul Rahman Centenary Fund (St Catharine's College, Cambridge, UK) awarded to E.A.B.A.; Gates Cambridge Scholarship awarded to C.B.X.; L.H.S., S.G. and C.M. are supported by the British Heart Foundation PhD studentship FS/11/35/28871, FS/14/75/31134 and FS/14/12/30540 respectively; J.Z. was supported by the Cambridge Overseas Trust Scholarship and the Sun Hung Kai Properties-Kwoks’ Foundation; A.E.D.T. is funded by the Agency for Science, Technology & Research (A*STAR) Singapore and Wellcome Trust Award 085686/Z/08/A; LHS, JZ and EABA were further supported by the NIHR Cambridge Biomedical Research Centre; the Human Research Tissue Bank is supported by the NIHR Cambridge Biomedical Research Centre. The Cav1.3 constructs were kindly gifted by Dr. Joerg Striessnig and Dr Petronel Tuluc

Physiological and Pathological Roles in Human Adrenal of the Glomeruli-Defining Matrix Protein NPNT (Nephronectin)

Primary aldosteronism is a common cause of hypertension, which becomes refractory if undiagnosed, but potentially curable when caused by an aldosterone-producing adenoma (APA). The discovery of somatic mutations and differences in clinical presentations led to recognition of small but common zona glomerulosa (ZG)-like adenomas, distinct from classical large zona fasciculata-like adenomas. The inverse correlation between APA size and aldosterone synthase expression prompted us to undertake a systematic study of genotype-phenotype relationships. After a microarray comparing tumor subtypes, in which NPNT ($\textit{nephronectin}$) was the most highly (>12-fold) upregulated gene in ZG-like APAs, we aimed to determine its role in physiological and pathological aldosterone production. NPNT was identified by immunohistochemistry as a secreted matrix protein expressed exclusively around aldosterone-producing glomeruli in normal adrenal ZG and in aldosterone-dense ZG-like APAs; the highest expression was in ZG-like APAs with gain-of-function $\textit{CTNNB1}$ mutations, whose removal cured hypertension in our patients. NPNT was absent from normal zona fasciculata, zona fasciculata-like APAs, and ZG adjacent to an APA. NPNT production was regulated by canonical Wnt pathway, and $\textit{NPNT}$ overexpression or silencing increased or reduced aldosterone, respectively. NPNT was proadhesive in primary adrenal and APA cells but antiadhesive and antiapoptotic in immortalized adrenocortical cells. The discovery of $\textit{NPNT}$ in the adrenal helped recognition of a common subtype of APAs and a pathway by which Wnt regulates aldosterone production. We propose that this arises through NPNT's binding to cell-surface integrins, stimulating cell-cell contact within glomeruli, which define ZG. Therefore, NPNT or its cognate integrin could present a novel therapeutic target.This research was funded by grants from the National Institute for Health Research (NIHR) Senior Investigator award (NF-SI-0512-10052) to M.J. Brown. A.E.D. Teo is supported by the Agency for Science, Technology and Research (A*STAR) Singapore. This study is also supported by Wellcome Trust Translational Medicine and Therapeutics award to M.J. Brown (085686/Z/08/A). S. Garg is supported by the British Heart Foundation (FS/14/75/31134). J. Zhou is supported by the Cambridge Overseas Trust. Additional support was provided by the NIHR Cambridge Biomedical Research Centre (Cardiovascular and Metabolic, and Human Tissue Bank)

Somatic mutations of GNA11 and GNAQ in CTNNB1-mutant aldosterone-producing adenomas presenting in puberty, pregnancy or menopause.

Most aldosterone-producing adenomas (APAs) have gain-of-function somatic mutations of ion channels or transporters. However, their frequency in aldosterone-producing cell clusters of normal adrenal gland suggests a requirement for codriver mutations in APAs. Here we identified gain-of-function mutations in both CTNNB1 and GNA11 by whole-exome sequencing of 3/41 APAs. Further sequencing of known CTNNB1-mutant APAs led to a total of 16 of 27 (59%) with a somatic p.Gln209His, p.Gln209Pro or p.Gln209Leu mutation of GNA11 or GNAQ. Solitary GNA11 mutations were found in hyperplastic zona glomerulosa adjacent to double-mutant APAs. Nine of ten patients in our UK/Irish cohort presented in puberty, pregnancy or menopause. Among multiple transcripts upregulated more than tenfold in double-mutant APAs was LHCGR, the receptor for luteinizing or pregnancy hormone (human chorionic gonadotropin). Transfections of adrenocortical cells demonstrated additive effects of GNA11 and CTNNB1 mutations on aldosterone secretion and expression of genes upregulated in double-mutant APAs. In adrenal cortex, GNA11/Q mutations appear clinically silent without a codriver mutation of CTNNB1