Nephron-specific deletion of ALMS1 (Alstrom syndrome 1) increases surface NKCC2 expression and NaCl reabsorption in the Thick Ascending Limb
Authors: Maskey Dipak , Ortiz Pablo 
Published: May 2023

Mutations in the ALMS1 gene in humans cause Alström syndrome, characterized by progressive metabolic alterations that include obesity, hypertension, and chronic kidney disease (CKD). SNPs in the ALMS1 gene in the general population are associated with lower GFR, hypertension and metabolic syndrome. ALMS1 is widely expressed where it localizes to endosomes, centrosomes and the base of cilia. Our lab has previously shown that ALMS1 is expressed in renal tubules of the Thick Ascending Limb (TAL) where NKCC2 is the primary apical NaCl transporter. Whole animal gene deletion of ALMS1 increases blood pressure, and inhibits NKCC2 endocytosis, increasing its surface expression and renal NaCl reabsorption. However, whole animal ALMS1 KO rats or mice develop obesity, affecting the interpretation of organ-specific data. We hypothesized that ALMS1 is a part of protein complex that binds apical NKCC2 in TALs and promotes its endocytosis. To study the role of ALMS1 in the nephron without influence of obesity we generated a loxP flanked exon 7 ALMS1 transgenic mouse line, that we crossed with nephron specific inducible Pax8-rtTA Cre mice (Pax8-ALMS1 KO). 4 weeks after inducing Cre expression with doxycycline (2mg/ml in 2% sucrose), ALMS1 expression in TAL suspensions decreased by 75% (p<0.01). To study the role of ALMS1 in TAL function, we measured surface to total NKCC2 ratio, which was increased by 53±15% in TALs from Pax8-ALMS1 KO compared to control floxed ALMS1 (p<0.01). Recombinant ALMS1 (GST-Carboxyl terminus-ALMS1), or immunoprecipitation of ALMS1, pulled down NKCC2 from WT TAL lysates (n=3), suggesting that the mechanism by which ALMS1 regulates surface NKCC2 involves protein-protein interactions. 4 weeks after inducing Cre expression with doxycycline we measured bumetanide-induced natriuresis (4h) as an index of NKCC2-mediated NaCl absorption. We found that bumetanide induced UNa excretion was higher in Pax8-ALMS1 KO, compared to control (control: 115±19 vs Pax8-ALMS1 KO: 146±30 μmols Na/4h, p<0.05). Since whole animal ALMS1 deletion induced hypertension, we measured systolic blood pressure (SBP) by tail cuff one month after inducible deletion of ALMS1 or in floxed ALMS1 mice treated with doxycycline as controls. Baseline SBP in normal Na diet (0.4%) was similar between strains. However, SBP increased by 12±3 mmHg in Pax8-ALMS1 KO, but not in control floxed ALMS1 littermates after 9 days of high Na diet (WT: 100±10, Pax8-ALMS1 KO: 115±11 mmHg, p<0.05 from baseline, n=4). We conclude that ALMS1, binds NKCC2 to regulate its surface expression. Depletion of ALMS1 in the nephron enhances surface NKCC2, TAL NaCl reabsorption, and induces salt-sensitivity of blood pressure. Our data indicate that regulation of the ALMS1-NKCC2 interaction in the TAL is important for renal NaCl reabsorption and regulation of blood pressure.

Funding: NIH R56DK131114, Henry Ford Fund

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.