Cell-cycle arrest in g0/g1 phase by contact inhibition and
transforming growth factor-[beta]1 in mink mv1lu lung epithelial
cells.
Wu, Frank, Sue Buckley, Kim Chi Bui, Ann Yee, Hua-Yang Wu, Jian Liu,
and David Warburton.
Departments of Surgery and Pediatrics, Childrens Hospital of Los
Angeles, University of Southern California School of Medicine, Los
Angeles, CA 90027
APStracts 2:0210L, 1995.
Cell-cell contact and transforming growth factor-[beta]1 (TGF-b1)
inhibit Mv1Lu cell proliferation by arresting cell cycle progression
in G0/G1 phase. We, therefore, postulated that contact inhibition and
TGF-[beta]1 may target the same regulatory molecules to exert their
inhibitory effects. To test this hypothesis, Western analysis was
performed to determine the expression patterns of a 45 kD protein
(p45) that is cell cycle-regulated and of various cyclins and cyclin
-dependent protein kinases (Cdks) in synchronized Mv1Lu cells. Both
p45 and cyclin D2 were expressed at relatively high levels in late G1
phase of Mv1Lu cells pharmacologically synchronized with mimosine.
However, both contact inhibition and TGF-[beta]1 significantly
suppressed p45 expression and also arrested Mv1Lu cell cycle in G0/G1
phase. On the other hand, neither contact inhibition nor TGF-[beta]1
treatment affected the expression of cyclins D1, E, and A or of Cdk2
and Cdk5, while the expression of cyclin B1, Cdk4, and Cdc2, Cdc2
-associated kinase activity, and phosphorylation of the retinoblastoma
tumor suppressor protein (pRb) were all significantly inhibited by
both contact inhibition and TGF-[beta]1. Kinetic studies showed that
p45 expression in Mv1Lu cells reappeared either by 12 h after release
from contact inhibition or by 6-8 h after release from TGF-[beta]1
block. Release of Mv1Lu cells from contact inhibition in the presence
of TGF-[beta]1 prevented both the re-expression of p45 and cell cycle
progression. In contrast, while similar levels of cyclin D2
expression were observed in growth arrested Mv1Lu cells as compared
to asynchronous control Mv1Lu cells, increased levels of cyclin D2
expression were detected by 3-6 h after release from contact
inhibition, and this elevation was prevented by TGF-[beta]1 addition
early in G1 phase. Additionally, cyclin D2 phosphorylation and its
associated kinase activity were greatly inhibited by contact
inhibition and TGF-[beta]1, but cyclin D2 de novo synthesis was not
affected under these conditions. Our results indicate that inhibition
of G1 progression in Mv1Lu cells is mediated, at least in part, by
suppression of p45, cyclin D2/Cdk4, and cyclin B1/Cdc2 expression
and/or their activities, and hypophosphorylation of pRb. Taken
together, these results may define a common mechanism through which
negative growth signals converge on similar molecular targets to
exert G0/G1 control over Mv1Lu cell proliferation.
Received 21 December 1995; accepted in final form 25 October
1995.
APS Manuscript Number L366-4.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 30 November 95