EFFECTS OF HISTONE-H4 DEPLETION ON THE CELL-CYCLE AND TRANSCRIPTION OF SACCHAROMYCES-CEREVISIAE

We have constructed a yeast strain (UKY403) in which the sole histone H4 gene is under control of the GAL1 promoter. This allows the activation of H4 mRNA synthesis on galactose and its repression on glucose. UKY403 cells, pre-synchronized in G1 with alpha-mating factor, have been used to show that glucose treatment results in the loss of approximately half the chromosomal nucleosomes. This depletion in only partially reversible when the H4 gene is reactivated on galactose. It was found that the resultant lethality manifests itself first in S phase, the period of nucleosome assembly, but leads to highly synchronous arrest in G2 and a virtually complete block in chromosomal segregation. Histone H4-depleted chromatin was analyzed for its efficiency as a template for all three RNA polymerases. Using pulse-labeling, we find no evidence for altered transcription by RNA polymerase I (25S, 18S, and 5.8S rRNAs) or RNA polymerase III (5S, rRNA, tRNAs). Northern blot analysis was used to measure levels of RNA polymerase II transcripts. There was little effect on the activation or repression of the CUP1 chelatin gene. While there may be some decrease in the level of certain mRNAs (e.g. HIS4, ARG4) other message levels (HIS3, TRP1) show little change upon glucose repression. Therefore, nucleosome loss certainly does not have a general effect on transcription.