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Free keywords:
LARGE-T-ANTIGEN; CASEIN-KINASE-I; SIMIAN-VIRUS-40 DNA-REPLICATION;
WILD-TYPE P53; PROTEIN-KINASE; BREAST-CANCER; BETA-CATENIN; TRANSGENIC
MICE; SV40-TRANSFORMED CELLS; MUTATIONAL ANALYSISBiology;
Abstract:
Simian virus 40 (SV40) is a powerful tool to study cellular
transformation in vitro, as well as tumor development and progression
in vivo. Various cellular kinases, among them members of the CK1
family, play an important role in modulating the transforming activity
of SV40, including the transforming activity of T-Ag, the major
transforming protein of SV40, itself. Here we characterized the effects
of mutant CK1 delta variants with impaired kinase activity on
SV40-induced cell transformation in vitro, and on SV40-induced mammary
carcinogenesis in vivo in a transgenic/bi-transgenic mouse model. CK1
delta mutants exhibited a reduced kinase activity compared to wtCK1
delta in in vitro kinase assays. Molecular modeling studies suggested
that mutation N172D, located within the substrate binding region, is
mainly responsible for impaired mutCK1 delta activity. When stably
over-expressed in maximal transformed SV-52 cells, CK1 delta mutants
induced reversion to a minimal transformed phenotype by
dominant-negative interference with endogenous wtCK1 delta. To
characterize the effects of CK1 delta on SV40-induced mammary
carcinogenesis, we generated transgenic mice expressing mutant CK1
delta under the control of the whey acidic protein (WAP) gene promoter,
and crossed them with SV40 transgenic WAP-T-antigen (WAP-T) mice. Both
WAP-T mice as well as WAP-mutCK1 delta/WAP-T bi-transgenic mice
developed breast cancer. However, tumor incidence was lower and life
span was significantly longer in WAP-mutCK1 delta/WAP-T bi-transgenic
animals. The reduced CK1 delta activity did not affect early lesion
formation during tumorigenesis, suggesting that impaired CK1 delta
activity reduces the probability for outgrowth of in situ carcinomas to
invasive carcinomas. The different tumorigenic potential of SV40 in
WAP-T and WAP-mutCK1 delta/WAP-T tumors was also reflected by a
significantly different expression of various genes known to be
involved in tumor progression, specifically of those involved in
wnt-signaling and DNA repair. Our data show that inactivating mutations
in CK1 delta impair SV40-induced cellular transformation in vitro and
mouse mammary carcinogenesis in vivo.
