Regulation of NKCC2 and NCC by Protein Protein interactions: From salt losing tubulopathies to salt sensitive hypertension

点击翻译按钮获取中文摘要

Hypertension in one of the most common diseases affecting human population. It is a major risk factor of cardiovascular morbidity and mortality. Analyses of monogenic syndromes, has allowed to link hypertension to elevated renal Na reabsorption, in particular through the increase in the activity of the sodium channel (ENaC) in the collecting duct. However, this focused attention on ENaC and its activator aldosterone has somehow masked the pivotal significance of the thick ascending limb (TAL) and the distal convoluted tubule (DCT) for NaCl transport and hence for blood pressure regulation. The apically located Na-K-2Cl and NaCl co-transporters (NKCC2 and NCC) reabsorb one third of the filtered NaCl and therefore are the pacemakers of sodium chloride reabsorption in TAL and DCT, respectively. Mutations in NKCC2 and NCC cause Bartter's (BS) and Gitelman's syndromes (GS), inherited disorders featuring low blood pressure due to renal NaCl wasting. Our recent finding that mutations in MAGE-D2 cause a very severe form of antenatal BS (BS type V) with extreme prematurity and hence high mortality due to functional impairment of NKCC2 and NCC (Laghmani et al, NEJM 2016) clearly underlines the importance of the regulatory proteins of the transporters for their functioning. MAGE-D2 is expressed in the TAL and DCT in fetal and adult kidney. Intriguingly, all symptoms of BS type V completely resolve postnatally, indicating that not only regulatory factors but also the context (physiological prenatal hypoxia versus postnatal normoxia) determine the functioning of NKCC2 and NCC. Despite the obvious significance of regulatory proteins and perhaps also the cellular environment very little is known about their impact on NKCC2 and NCC expression. Hence, we propose to: 1) Unravel the protein networks and key pathways involved in the regulation, in vitro and in vivo, of NKCC2 and NCC. 2) Elucidate the molecular pathways differentially regulating expression and function of mutated NKCC2 proteins. 3) Investigate the molecular mechanisms responsible for the transient nature of BS in carriers of MAGE-D2 mutations. 4) Identify new regulatory pathways and/or genes responsible for antenatal BS. To this end, we propose a multidisciplinary project combining several experimental approaches including: molecular and cell biology, genetics in patients with unidentified causes of BS, In vivo and in vitro gene silencing techniques (using primary cell culture, stable cell lines, as well as drosophila and mice as a whole animal model), in vivo physiology using metabolic cages and in vitro microperfusion of nephron segments. A better understanding of the regulatory pathways acting on NKCC2 and NCC expression and activity will help to gain insights into the pathophysiology of salt retention. Our efforts will ultimately identify new drugable targets to prevent and/or treat kidney disorders related to sodium balance.