Periodic transcription as a means of regulating gene expression during the cell cycle: contrasting modes of expression of DNA ligase genes in budding and fission yeast.

Abstract

Using cultures synchronised by three independent procedures, we have shown that the CDC9 gene, coding for DNA ligase, is periodically expressed in the Saccharomyces cerevisiae cell cycle. The level of CDC9 transcript increases many fold in late G1 reaching a peak at about the G1/S phase boundary and preceding the peak in histone message by some 20 min. The level of DNA ligase itself also fluctuates, showing the expected pattern for a stable enzyme synthesised periodically. In contrast, the transcript from the DNA ligase gene (CDC17) of Schizosaccharomyces pombe is present at a constant level throughout the cell cycle, and no fluctuation in amount was detected, although the histone H2A showed the expected periodic synthesis. Furthermore, DNA ligase activity remains at a constant level during the S. pombe cell cycle showing that there is unlikely to be any form of translational control. These contrasting modes of expression of the DNA ligase genes in the two organisms suggests that when periodic transcription is observed from an essential cell cycle gene, it may have no particular significance for regulating progress through the cell cycle. Also, regulatory circuits may be less well conserved between organisms than the processes they control and thus different organisms may utilise quite different modes of control to achieve the same ends.

TheSchizosaccharomyces pombeHIRA-Like Protein Hip1 Is Required for the Periodic Expression of Histone Genes and Contributes to the Function of Complex Centromeres

“…The aqueous layer was subjected to two further phenol-chloroform extractions before the RNA was precipitated with 0.1 volume of sodium acetate (pH 5.2) and 0.6 volume of isopropanol. RNA pellets were washed in 70% ethanol and resuspended in H 2 O. RNA analysis was performed as described previously (66). Briefly, a 10-to 15-g sample of total RNA was denatured with glyoxal, separated on a 1.2% agarose gel prepared in 15 mM sodium phosphate (pH 6.5), and transferred to a GeneScreen hybridization membrane (Dupont NEN Research Products).…”

Gin4 of S. cerevisiae is a bud neck protein that interacts with the Cdc28 complex

“…H2A is cyclically transcribed and its transcription peaks in mid S-phase, whereas the adjacent Protein 1 gene, called TRT1, is expressed constitutively (Hereford et al 1981;White et al 1986). This periodic transcription of GIN4 with a peak at the G1/S boundary is reminiscent of the pattern observed with many other genes involved in the S phase control or the bud emergence regulation, in which the periodic transcriptions were regulated by a promoter element called MCB (ACGCGTNA, Johnston & Loundes 1992; Igual et al 1996).…”

P40SDB25, a putative CDK inhibitor, has a role in the M/G1 transition in Saccharomyces cerevisiae.

“…Total RNA was extracted from cells as described previously (White et al 1986). A 5-~g sample of total RNA, denatured with glyoxal, was separated on a 1.2% agarose gel, transferred to a GeneScreen hybridization membrane (Dupont, NEN Research Products, Boston, MA}.…”

“…Probes for RNA-DNA hybridization were internal fragments from the genes concerned, with the SDB25 probe being the 750-bp KpnI-HpaI fragment. DNA was labeled with [a-a2P]TTP by use of an oligo-labeling protocol (White et al 1986). RNA levels were quantitated with a Personal Densitometer PD-130 (Molecular Dynamics).…”