Cell cycle-dependent gene expression in V point-arrested BALB/c-3T3 cells

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Abstract
Density‐arrested BALB/c‐3T3 cells stimulated to proliferate in an amino acid‐deficient medium arrest in mid‐G1 at a point termed the V point. Cells released from V point arrest require 6 hr to traverse late G1 and enter S phase. As data presented here show that mRNA synthesis is needed for 2–3 hr after release of cells from the V point, after which inhibition of mRNA synthesis does not prevent entry into S phase, we used this mid‐G1 arrest protocol to analyze gene expression in late G1. We found that although stimulation of cells in amino acid‐deficient medium did not inhibit the induction of genes expressed in early G1, genes normally expressed in late G1 were expressed only after release from the V point. The expression of late G1 genes in cells released from the V point was temporally similar, in respect to G1 location, as was seen in stimulation of quiescent Go cells. As this protocol effectively divides gene expression into early (pre‐V point) and late (post‐V point) categories, it should be useful in studies of growth factor‐modulated events that regulate traverse of late G1 and commitment to DNA synthesis. In addition, we used c‐myb antisense oligonucleotides to show that c‐myb expression, which occurs in late G1, is required for BALB/c‐3T3 fibroblasts to traverse late G1 and initiate DNA synthesis.