Role of Myosin1a in regulated exocytosis of CFTR in villus enterocytes

Abstract

Insertion of CFTR channels into the intestinal brush border membrane (BBM) is necessary for anion secretion, but the mechanisms of CFTR delivery into the BBM are poorly understood. BB myosins move cargo and regulate membrane trafficking. Myosin1a (Myo1a) regulates CFTR BBM localization and function in villus enterocytes. cAMP and cGMP regulate CFTR traffic into the enterocyte BBM from subapical endosomes. We hypothesized that Myo1a regulates cAMP/cGMP traffic of CFTR into the BBM. Myo1a KO and control mouse intestine were treated with cAMP or cGMP agonists. CFTR distribution was examined by confocal microscopy and surface biotinylation. Immunolocalization revealed absence of BBM CFTR and its accumulation in subapical endosomes in Myo1aKO villus enterocytes. Surface biotinylation confirmed no change in surface CFTR in stimulated Myo1aKO tissues. Specificity of Myo1a was confirmed by lack of changes in BBM localization of ALP, SLC26A6 (PAT‐1), and NHE3. Surface biotinylation of cAMP‐stimulated polarized Caco2BB‐e cells lacking Myo1a expression (shRNA silencing) confirmed CFTR specificity. Surface CFTR was reduced by 98% in Myo1a knockdown (KD) cells compared to control, while surface PAT1 was unchanged in control and Myo1a KD cells. These data support Myo1a as the motor for regulated exocytosis of CFTR. Future studies will seek to dissect the molecular mechanisms regulating Myo1a‐dependent CFTR traffic.