Normal Calcium-Activated Anion Secretion in a Mouse Selectively Lacking TMEM16A in Intestinal Epithelium

Abstract

Calcium-activated anion secretion is expected to ameliorate cystic fibrosis, a genetic disease that carries an anion secretory defect in exocrine tissues. Human patients and animal models of the disease that present a mild intestinal phenotype have been postulated to bear a compensatory calcium-activated anion secretion in the intestine. TMEM16A is calcium-activated anion channel whose presence in the intestinal epithelium is contradictory. We aim to test the functional expression of TMEM16A using animal models with <i>Cftr</i> and/or <i>Tmem16a</i> intestinal silencing. Expression of TMEM16A was studied in a wild type and intestinal <i>Tmem16a</i> knockout mice by mRNA-seq, mass-spectrometry, q-PCR, Western blotting and immunolocalization. Calcium-activated anion secretion was recorded in the ileum and proximal colon of these animals including intestinal <i>Cftr</i> knockout and double mutants with dual <i>Tmem16a</i> and <i>Cftr</i> intestinal ablation. Mucus homeostasis was studied by immune-analysis of Mucin-2 (Muc2) and survival curves were recorded. <i>Tmem16a</i> transcript was found in intestine. Nevertheless, protein was barely detected in colon samples. Electrophysiological measurements demonstrated that the intestinal deletion of <i>Tmem16a</i> did not change calcium-activated anion secretion induced by carbachol or ATP in ileum and proximal colon. Muc2 architecture was not altered by <i>Tmem16a</i> silencing as was observed when <i>Cftr</i> was deleted from mouse intestine. <i>Tmem16a</i> silencing neither affected animal survival nor modified the lethality observed in the intestinal <i>Cftr</i>-null mouse. Our results demonstrate that TMEM16A function in the murine intestine is not related to electrogenic calcium-activated anion transport and does not affect mucus homeostasis and survival of animals.