Loss of PALS1 expression leads to tight junction and polarity defects.
Prior work in our laboratory established a connection between the PALS1/PATJ/CRB3 and Par6/Par3/aPKC protein complexes at the tight junction of mammalian epithelial cells. Utilizing a stable small interfering RNA expression system, we have markedly reduced expression of the tight junction-associated protein PALS1 in MDCKII cells. The loss of PALS1 resulted in a corresponding loss of expression of PATJ, a known binding partner of PALS1, but had no effect on the expression of CRB3. However, the absence of PALS1 and PATJ expression did result in the decreased association of CRB3 with members of the Par6/Par3/aPKC protein complex. The consequences of the loss of PALS1 and PATJ were exhibited by a delay in the polarization of MDCKII monolayers after calcium switch, a decrease in the transepithelial electrical resistance, and by the inability of these cells to form lumenal cysts when grown in a collagen gel matrix. These defects in polarity determination may be the result of the lack of recruitment of aPKC to the tight junction in PALS1-deficient cells, as observed by confocal microscopy, and subsequent alterations in downstream signaling events.
Straight
Samuel W
SW
University of Michigan, Department of Biological Chemistry, Ann Arbor, Michigan 48109, USA.
Shin
Kunyoo
K
Fogg
Vanessa C
VC
Fan
Shuling
S
Liu
Chia-Jen
CJ
Roh
Michael
M
Margolis
Ben
B
D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003094:Collagen / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004285:Dogs; D004594:Electrophysiology; D004847:Epithelial Cells / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D005136:Eye Proteins / Q000502:physiology*; D015151:Immunoblotting; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000502:physiology*; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011233:Precipitin Tests; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D062725:Tight Junction Proteins; D019108:Tight Junctions; D013997:Time Factors; D014162:Transfection
C472002:CRB3 protein, human; D005136:Eye Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C107903:PATJ protein, human; D010455:Peptides; D034741:RNA, Small Interfering; D062725:Tight Junction Proteins; D003094:Collagen; D004247:DNA; D002118:Calcium