A novel role of the C-terminus of b^0,+AT in the ER-Golgi trafficking of the rBAT-b^0,+AT heterodimeric amino acid transporter

The heterodimeric complex composed of rBAT (related to b^0,+ amino acid transporter), a single-membrane-spanning glycosylated heavy chain, and b^0,+ AT, a putative 12-membrane-spanning non-glycosylated light chain, is an amino acid transporter that mediates the activity of system b^0,+, a major apical transport system for cystine and dibasic amino acids in renal proximal tubule and small intestine. The C-terminus of b^0,+ AT has been proposed to play an important role in the functional expression of the heterodimeric transporters. In the present study, to reveal the roles of the C-terminus, we analysed b^0,+ AT mutants whose C-termini were sequentially deleted or replaced by site-directed mutagenesis in polarized MDCKII (Madin-Darby canine kidney II), non-polarized HEK-293 (human embryonic kidney-293) and HeLa cells. Although the deletion of C-terminus of b^0,+ AT did not affect the formation of a heterodimer with rBAT, it resulted in the loss of apparent transport function, owing to the failure of the plasma-membrane targeting of rBAT-b^0,+ AT heterodimeric complex associated with incomplete glycosylation of rBAT. A motif-like sequence Val⁴⁸⁰-Pro⁴⁸¹-Pro⁴⁸² was identified in the C-terminus of b^0,+ AT to be responsible for the C-terminus action in promoting the trafficking of rBAT-b^0,+ AT heterodimeric complex from the ER (endoplasmic reticulum) to Golgi apparatus. This is, to our knowledge, the first demonstration of the active contribution of the C-terminus of a light-chain subunit to the intracellular trafficking of heterodimeric transporters. Because the motif-like sequence Val⁴⁸⁰-Pro⁴⁸¹-Pro⁴⁸² is well conserved among the C-termini of light-chain subunits, common regulatory mechanisms could be proposed among heterodimeric amino acid transporters.