In all eukaryotic organisms, replication origins fire at specified times during S phase. Some fire early, some in middle S phase, and some late. The biological significance of origin timing is not yet clear, but previous investigations have revealed a strong correlation between replication timing and transcription. That is, genes located in early-replicated regions tend to be transcriptionally active, while genes located in late-replicated regions tend to be inactive. Thus replication timing may be one of the mechanisms by which cells control gene transcription.
At this point, nothing is known about the mechanisms of replication timing control, and until recently nothing was known about the features that distinguish early-firing from late-firing origins. Now we (C. Yompakdee and J.A. Huberman, JBC 2004) have identified a short, G-rich late consensus sequence (LCS), which (when present in clusters of three or more) can force late replication timing on nearby origins. We are currently trying to identify the protein(s) that bind to the LCS. Once we have found LCS-binding protein(s), we will have a handle on the mechanism that enforces late replication for at least a subset of origins in fission yeast, but likely also for at least a subset of origins in all eukaryotic organisms.
Like other eukaryotic cells, fission yeast cells slow down their procession through S phase in response to DNA damage. This retardation of S phase depends on DNA damage checkpoint proteins such as Rad3 (similar to human ATM) and Cds1 (similar to human CDS1/CHK2), and the checkpoint is mediated by selective inhibition of the firing of late replication origins. We previously identified several of the proteins that cooperate with Rad3 to activate the Cds1 kinase (Marchetti et al., 2002), and we (Kumar and Huberman, JBC 2004) have recently identified the proteins in this pathway that function downstream of Cds1. Now we are identifying additional proteins that participate in this pathway, and we are studying the mechanisms by which these proteins regulate the rate of DNA replication.
The availability of the complete nucleotide sequence of the fission yeast genome now permits us to study replication timing at a genome-wide level. We are collaborating with the laboratory of Janet Leatherwood at SUNY Stony Brook to determine the replication time of every portion of the fission yeast genome and to determine how mutations in genes that affect replication, checkpoints and chromatin structure modulate that timing.
We are also collaborating with the laboratory of Bill Burhans at Roswell Park Cancer Institute to explore the connections between initiation of DNA replication, cell-cycle checkpoints, and apoptosis. Recent findings (see list of recent publications) suggest that multiple independent pathways lead from defects in the initiation of DNA replication to apoptosis in both budding and fission yeasts. Defects in similar apoptosis-inducing pathways may contribute to the development of cancer in vertebrates.
This page was updated on March 27, 2006.
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