D-type cyclin expression is decreased and p21 and p27 CDK inhibitor expression is increased when tsBN462 CCG1/TAF₁₁250 mutant cells arrest in G1 at the restrictive temperature

Background: The tsBN462 temperature-sensitive mutant hamster cell line exhibits cell cycle arrest and apoptosis at the restrictive temperature of 39.5 °C, due to a point mutation in the CCG1/ TAF₁₁250 gene, which encodes a component of the general transcription factor TFIID. Results: We now report that CCG1/TAF_II250 persisted as a complex with TBP and associated proteins (TAFs) in tsBN462 cells at the restrictive temperature. FACScan analysis revealed that the tsBN462 mutation resulted in a failure to progress out of G0 into G1. Using two-dimensional gel electrophoresis we observed a decrease in the synthesis of several proteins, starting in the middle of the G1 phase, becoming very pronounced during late G1. The expression of the immediate early genes c-fos, c-jun and c-myc was normally induced by seruin treatment of quiescent cells at the restrictive temperature, whereas expression of cyclins A, Dl and D3 was reduced. Expression of the cyclin-dependent kinase (CDK) inhibitor proteins p21 and p27 was enhanced. Consistent with the decreased cyclin D and increased p21/p27 expression, we found that phosphorylation of Rb was decreased at 39.5 °C. Cyclin A-, E- and Cdk2-associated historic HI kinase activity was reduced concomitantly with the increase in p21 protein. Conclusion: Decreased cyclin/Cdk kinase activity and decreased Rb phosphorylation are possible causes of G1 cell cycle arrest in tsBN462 cells at the restrictive temperature.