Synthesis of DNA-binding proteins during the cell cycle of WI-38 cells

Abstract

The DNA-binding proteins synthesized by WI-38 cells at specific periods during the cell cycle were chromatographed on calf thymus DNA-cellulose and analyzed by SDS-polyacrylamide gel electrophoresis. The DNA-binding proteins which were predominantly labeled in G 1 had approximate mol wts of 20 000, 30 000 and 68 000. During the S phase, in addition to the G 1 proteins, some large proteins and histones were labeled. Proteins of approx. 45 000 D seemed to be unique to the S period. The profile of DNA-binding proteins synthesized in G 2 is very similar to that of G 1. The labeling of DNA-binding proteins in the early G1 was markedly reduced in the presence of 5 μg/ml of actinomycin D indicating that synthesis of these G 1 proteins is dependent on new transcription. In order to determine the relationship between the DNA-binding proteins prepared from the whole cell sonicates and nuclear proteins, DNA-binding proteins of cytoplasm and nuclear fractions were examined. DNA-binding proteins are found to exist both in nuclear and cytoplasmic fractions of WI-38 cells. Nucleocytoplasmic migration of DNA-binding proteins was also studied by pulse labeling G 1 cells with 3H-leucine and determining the amount of radioactivity incorporated into cytoplasmic and nuclear DNA-binding protein fractions after chasing for various periods of time. Furthermore, incubation of unlabeled isolated nuclei with the 3H-DNA-binding proteins in vitro revealed that fractions of these cytoplasmic DNA-binding proteins are transported in nuclei. Present experiments suggest that DNA-binding proteins synthesized in the late G 1 are correlated with the onset of S and that a fraction of DNA-binding proteins synthesized in the cytoplasm migrates into the nuclei.