Targeted deletion of the Tsg101 gene results in cell cycle arrest at G1/S and p53-independent cell death.
The tumor susceptibility gene 101 (Tsg101) was originally discovered in a screen for potential tumor suppressors using insertional mutagenesis in immortalized fibroblasts. To investigate essential functions of this gene in cell growth and neoplastic transformation, we derived primary mouse embryonic fibroblasts from Tsg101 conditional knockout mice. Expression of Cre recombinase from a retroviral vector efficiently down-regulated Tsg101. The deletion of Tsg101 caused growth arrest and cell death but did not result in increased proliferation and cellular transformation. Inactivation of p53 had no influence on the deleterious phenotype, but Tsg101(-/-) cells were rescued through expression of exogenous Tsg101. Fluorescence-activated cell sorting, proliferation assays, and Western blot analysis of crucial regulators of the cell cycle revealed that Tsg101 deficiency resulted in growth arrest at the G(1)/S transition through inactivation of cyclin-dependent kinase 2. As a consequence, DNA replication was not initiated in Tsg101-deficient cells. Our results clearly demonstrate that Tsg101 is not a primary tumor suppressor in mouse embryonic fibroblasts. However, the protein is crucial for cell proliferation and cell survival.
Krempler
Andrea
A
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Nebraska Medical Center, Omaha, Nebraska 68198-6805, USA.
Henry
MaLinda D
MD
Triplett
Aleata A
AA
Wagner
Kay-Uwe
KU
D000818:Animals; D002453:Cell Cycle / Q000502:physiology*; D016923:Cell Death / Q000502:physiology*; D002455:Cell Division; D002470:Cell Survival; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D004622:Embryo, Mammalian; D056827:Endosomal Sorting Complexes Required for Transport; D005347:Fibroblasts / Q000166:cytology; D016193:G1 Phase / Q000502:physiology; D016350:Leucine Zippers; D051379:Mice; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000378:metabolism; D016196:S Phase / Q000502:physiology; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D000099177:Tsg101 Protein
D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D011994:Recombinant Proteins; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D000099177:Tsg101 Protein