Transcriptional control of gene expression during the G2-M phase of the cell cycle

The division of eukaryotic cells into two daughter cells is coordinately regulated. This division process takes from between 1 hour in yeasts to up to 24 hours in humans. Over this period a series of events is initiated in a process called the cell cycle whereby one event leads to another, and ultimately culminates in cell division and then reinitiation of the whole process. To regulate this process, there are numerous control points in the cell cycle. One of the major control points is following DNA replication and before the cells commit to division (the G2-M control point). This control point is at least partially regulated at the level of transcription, the process by which genes are turned on and their code deciphered and converted to first RNA and the protein. This project aims to study how transcription at this key control point is regulated. We will use the model organism S. Cerevisiae (budding yeast) which shows a remarkable conservation of cell cycle control mechanisms with higher organisms such as humans. We know the key regulators (transcription factors) that act at this control point, and how they themselves can be regulated through modification by a process known as phosphorylation. However, what we do not know is how the transcription factors regulate transcription. Furthermore, we do not know how subtly different activation events occur at this control point. ie not all genes are activated with the same temporal kinetics. We aim to study the molecular details of how transcription factors function to regulate target gene transcription and how these transcription factors are themselves controlled. We anticipate that the findings we make in our model system will be directly relevant to other systems. Indeed the deregulation of cell cycle control is a pre-requisite for tumourigenesis, therefore our studies may impact on our understanding of and ability to combat cancer.

The cell cycle is subjected to a number of stringent control measures to ensure that DNA duplication and separation is correctly executed with high fidelity. Many of the control mechanisms are conserved between eukaryotes as diverse as yeasts and humans. Part of this control is at the transcriptional level, where successive waves of transcription are initiated which give rise to the products required for progression through subsequent phases of the cell cycle. In this study, we will investigate the control of transcription around the G2-M border using the model organism S. cerevisiae. A peak of genes is coordinately regulated at this timepoint (the CLB2 cluster) and a transciption factor complex containing Mcm1p, Fkh2p and Ndd1p is thought to be involved in the coordinate regulation of all these genes. However, a comprehensive study of the entire cluster has not been performed and studies on a limited number of genes have indicated that while the core control mechanism is similar, subtle differences in control can be observed. The aim of this proposal is to establish the mechanisms by which these transcription factors activate transcription and how this is regulated. Importantly, we also wish to establish how the timing of peak expression of individual genes is subtly altered. To achieve this, we will establish the regulatory events taking place at CLB2 cluster promoters. Next we will study how additional regulators such as Yox1p, Cks1p and YDL203c/Ack1p impact on the control of CLB2 cluster promoters. In addition, we will investigate how cell cycle-dependent protein kinases such as Clbp-Cdk and Cdc5p impact on the activity of these promoters by targeting associated regulatory proteins.