Biochem. J. (2008) 414
(195–203) (Printed in Great Britain)
Assembly of a membrane receptor complex: roles of the uroplakin II prosequence in regulating uroplakin bacterial receptor oligomerization
Chih-Chi Andrew HU*†, Thomas BACHMANN‡, Ge ZHOU‡, Feng-Xia LIANG†, Jorge GHISO§∥, Gert KREIBICH‡** and Tung-Tien SUN†‡¶**1
*Department of Pharmacology, New York University School of Medicine, New York, NY 10016, U.S.A., †Epithelial Biology Unit, The Ronald O. Perelman Department of Dermatology, New York University School of Medicine, New York, NY 10016, U.S.A., ‡Department of Cell Biology, New York University School of Medicine, 550 1st Avenue, New York, NY 10016, U.S.A., ‡Department of Pathology, New York University School of Medicine, New York, NY 10016, U.S.A., ∥Department of Psychiatry, New York University School of Medicine, New York, NY 10016, U.S.A., ¶Department of Urology, New York University School of Medicine, New York, NY 10016, U.S.A., and **NYU Cancer Institute, New York University School of Medicine, New York, NY 10016, U.S.A.
The apical surface of the mammalian urothelium is almost completely covered by two-dimensional protein crystals (known as urothelial plaques) of hexagonally packed 16 nm particles consisting of two UP (uroplakin) heterodimers, i.e. UPs Ia/II and Ib/III pairs. UPs are functionally important as they contribute to the urothelial permeability barrier function, and UPIa may serve as the receptor for the uropathogenic Escherichia coli that causes over 90% of urinary tract infections. We study here how the UP proteins are assembled and targeted to the urothelial apical surface, paying special attention to the roles of the prosequence of UPII in UP oligomerization. We show that (i) the formation of the UPIa/UPII heterodimer, necessary for ER (endoplasmic reticulum) exit, requires disulfide formation in the prosequence domain of proUPII (the immature form of UPII still containing its prosequence); (ii) differentiation-dependent N-glycosylation of the prosequence leads to UP stabilization; (iii) a failure to form tetramers in cultured urothelial cells, in part due to altered glycosylation of the prosequence, may block two-dimensional crystal formation; and (iv) the prosequence of UPII remains attached to the mature protein complex on the urothelial apical surface even after it has been cleaved by the trans-Golgi-network-associated furin. Our results indicate that proper secondary modifications of the prosequence of UPII play important roles in regulating the oligomerization and function of the UP protein complex.
Key words: disulfide formation, glycosylation, integral membrane protein, prosequence, protein assembly, uroplakin.
Abbreviations used: asterisks indicate the glycosylated forms of each protein; AUM, asymmetric unit membrane; BN-PAGE, blue native PAGE; DMEM, Dulbecco's modified Eagle's medium; DSS, disuccinimidyl suberate; DTT, dithiothreitol; endo F, endoglycosidase F; endo H, endoglycosidase H; ER, endoplasmic reticulum; FBS, fetal bovine serum; TGN, trans-Golgi network; UP, uroplakin; proUPII, the immature form of UPII still containing its prosequence; pro-UPII, proUPII in which the prosequence has been cleaved by furin, but remains attached to UPII; UPIa, UPIb, UPII and UPIIIa, the four principal UP proteins.
1To whom correspondence should be addressed, at Department of Cell Biology, New York University School of Medicine, 550 1st Avenue, New York, NY 10016, U.S.A. (email Tungtien.sun@gmail.com).
Received 11 March 2008/9 May 2008; accepted 16 May 2008
Published as BJ Immediate Publication 16 May 2008, doi:10.1042/BJ20080550
© The Authors Journal compilation © 2008 Biochemical Society
Editorial Advisory Panel
RSS feeds
Table of contents by email
My BJ toolbox
