Dynamic subcellular localization of sodium-bicarbonate cotransporter NBCn1/SLC4A7 to plasma membrane, centrosomes, spindle, and primary cilia

Abstract

Finely tuned regulation of transport protein localization is vital for epithelial function. Sodium-bicarbonate co-transporter NBCn1 (SLC4A7) is a key contributor to epithelial pH homeostasis, yet the regulation of its subcellular localization is not understood. Here, we show that a predicted N-terminal {beta}-sheet and short C-terminal -helical motif are essential for NBCn1 plasma membrane localization in epithelial cells. This localization was abolished by cell-cell contact disruption, and co-immunoprecipitation (co-IP) and proximity ligation (PLA) revealed NBCn1 interaction with E-cadherin and DLG1, linking the transporter to adherens junctions and the Scribble complex. NBCn1 also interacted with RhoA and localized to lamellipodia and filopodia in migrating cells. Finally, analysis of localization of native and GFP-tagged NBCn1, subcellular fractionation, co-IP of NBCn1 with Arl13B and CEP164, and PLA of NBCn1 and tubulin in mitotic spindles led to the surprising conclusion that NBCn1 additionally localizes to the centrosome and primary cilium in non-dividing, polarized epithelial cells, and to spindle, centrosome and midbodies during mitosis. We propose that NBCn1 traffics between lateral junctions, leading edge, and cell division machinery in Rab11 endosomes, adding new insight to the role of NBCn1 in cell cycle progression. Summary statementWe unravel molecular determinants of plasma membrane localization of the Na+,HCO3- cotransporter NBCn1 and discover that NBCn1 also localizes to centrosomes, spindle, midbody and primary cilia, likely cycling between these compartments.