LncRNA-induced lysosomal localization of NHE1 promotes increased lysosomal pH in macrophages leading to atherosclerosis.

ANRIL, also referred to as CDKN2B-AS1, is an lncRNA gene implicated in the pathogenesis of multiple human diseases including atherosclerotic coronary artery disease; however, definitive in vivo evidence is lacking and the underlying molecular mechanism is largely unknown. In this study, we show that ANRIL overexpression causes atherosclerosis in vivo as transgenic mouse overexpression of full-length ANRIL (NR_003529) increases inflammation and aggravates atherosclerosis under ApoE background (ApoEANRIL mice). Mechanistically, ANRIL reduces the expression of miR-181b-5p, which leads to increased TMEM106B expression. TMEM106B is significantly upregulated in the atherosclerotic lesions of both human CAD patients and ApoEANRIL mice. TMEM106B interacts and colocalizes with Na-H exchanger NHE1, which results in the mislocalization of NHE1 from cell membranes to lysosomal membranes, leading to increased lysosomal pH in macrophages. Large truncation and point mutation analyses define the critical amino acids for TMEM106B-NHE1 interaction and lysosomal pH regulation as F115 and F117 on TMEM106B and I537, C538, and G539 on NHE1. Topological analysis suggests that both N terminus and C terminus of NHE1 are located inside lysosomal lumen, consistent with our finding that NHE1 is an important new proton efflux channel involved in raising lysosomal pH. A short TMEM106B peptide (YGRKKRRQRRR-LAVFFLF) disrupting the TMEM106B-NHE1 interaction normalized lysosomal pH in macrophages with ANRIL overexpression. Our data demonstrate that ANRIL promotes atherosclerosis in vivo and identify the ANRIL-miR-181b-5p-TMEM106B-NHE1-lysosomal pH axis as the underlying molecular pathogenic mechanism for the chromosome 9p21.3 genetic locus for coronary artery disease.