Mammalian lin-7 stabilizes polarity protein complexes.
Mammalian Lin-7 forms a complex with several proteins, including PALS1, that have a role in polarity determination in epithelial cells. In this study we have found that loss of Lin-7 protein from the polarized epithelial cell line Madin-Darby canine kidney II by small hairpin RNA results in defects in tight junction formation as indicated by lowered transepithelial electrical resistance and mislocalization of the tight junction protein ZO-1 after calcium switch. The knock down of Lin-7 also resulted in the loss of expression of several Lin-7 binding partners, including PALS1 and the polarity protein PATJ. The effects of Lin-7 knock down were rescued by the exogenous expression of murine Lin-7 constructs that contained the L27 domain, but not the PDZ domain alone. Furthermore, exogenously expressed PALS1, but not other Lin-7 binding partners, also rescued the effects of Lin-7 knock down, including the restoration of PATJ protein in rescued cell lines. Finally, the effects of Lin-7 knock down appeared to be due to instability of PALS1 protein in the absence of Lin-7, as indicated by an increased rate of PALS1 protein degradation. Taken together, these results indicate that Lin-7 functions in tight junction formation by stabilizing its membrane-associated guanylate kinase binding partner PALS1.
Straight
Samuel W
SW
Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA.
Pieczynski
Jay N
JN
Whiteman
Eileen L
EL
Liu
Chia-Jen
CJ
Margolis
Ben
B
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology; D004285:Dogs; D004355:Drug Stability; D006801:Humans; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D017434:Protein Structure, Tertiary; D012313:RNA / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019108:Tight Junctions / Q000378:metabolism
C119588:LIN-7 protein, mammalian; D008565:Membrane Proteins; D011994:Recombinant Proteins; D012313:RNA