2 years ago

2013 Scientific Report

  • Text
  • Report
  • Institute
  • Clinical
  • Molecular
  • Scientific
  • Tumor
  • Laboratory
  • Signaling

Michael Weinreich, Ph.D.

Michael Weinreich, Ph.D. Laboratory of Genome Integrity and Tumorigenesis Dr. Weinreich received his Ph.D. in biochemistry from the University of Wisconsin–Madison, after which he was a postdoctoral fellow in the laboratory of Bruce Stillman, director of Cold Spring Harbor Laboratory, New York. Dr. Weinreich joined VARI in March 2000 and is currently an Associate Professor. From left: Weinreich, Chang, Minard, Chen, Kenworthy, Tiwari Staff FuJung Chang, M.S. Jessica Kenworthy, B.S. Michelle Minard Kanchan Tiwari, M.S. Students Ying-Chou Chen, M.S. Nanda Kumar Sasi, B.S. Sandya Subramanian Raymond Yeow 63

Van Andel Research Institute | Scientific Report Research Interests The goal of our research is to understand how cells stably and accurately maintain their genetic information. Many diseases, including cancer, are caused by mutations in DNA, and it is now clear that the development of cancer requires multiple independent mutations. Early mutations often impair cellular surveillance mechanisms (checkpoints) that maintain genetic stability, and, in the absence of such checkpoints, additional mutations and genetic alterations become more frequent. This cumulative burden can ultimately lead to cancer as cells escape the normal growth and proliferation controls. Genetic instability also explains why cancer treatments often fail: tumors have such high mutation rates that they can readily develop resistance to chemotherapeutic drugs. The two-subunit Dbf4-dependent kinase (DDK) that we study (also known as Cdc7-Dbf4 protein kinase) is critical for the accurate replication and segregation of chromosomes. DDK is required for the initiation of DNA replication at multiple independent origins throughout the genome. It accomplishes this by phosphorylating and activating the MCM helicase, previously loaded in an inactive form at all origins during G1 phase. It is clear that DDK also affects replication fork stability and DNA repair processes during S phase, although the mechanisms for these activities are poorly understood. We recently reported that Dbf4 interacts with the yeast Polo-like kinase, Cdc5, to maintain the spindle position checkpoint. Polo kinases are master regulators of mitotic events. For example, Cdc5 promotes the loss of chromosome cohesion during metaphase, entry into anaphase, spindle elongation, exit from mitosis, and cytokinesis. Because of its essential role during mitosis, Cdc5 is the target of multiple checkpoint mechanisms to ensure the accurate segregation of chromosomes. We found that DDK inhibits Cdc5 when the mitotic spindle apparatus is not properly aligned between mother and daughter cells. Loss of this regulation can cause a significant increase in chromosome mis-segregation events and cell death. The DNA damage and replication checkpoints are critical regulators of chromosome stability. The checkpoints facilitate repair of DNA damage, suppress late-origin firing, and also prevent premature entry into mitosis, which would be catastrophic with damaged or incompletely replicated chromosomes. The Rad53 protein kinase of yeast, the ortholog of the human tumor suppressor Chk2, is an essential regulator of these checkpoints and directly interacts with Dbf4. Rad53 phosphorylates Dbf4 to prevent the activation of late origins when replication forks stall, and our genetic data imply that Rad53 and DDK also cooperate in another (unknown) pathway that is essential for cell survival. We have recently investigated the basis of the molecular interaction between Dbf4 and Rad53. Rad53 likely binds Dbf4 using multiple protein-protein contacts in the Dbf4 N-terminus. Interestingly, loss of the Rad53-Dbf4 regulation leads to activation of late-origin firing during periods of replication stress. It is unknown how Rad53 phosphorylation prevents late-origin activation, since we have shown that Rad53 phosphorylation does not disrupt the Dbf4-Cdc7 interaction and results in only a modest decrease in DDK activity. The Rad53 protein binds to the Dbf4 N-terminus but phosphorylates critical residues in the Dbf4 C-terminus to prevent late-origin activation. In summary, work over the last several years has shown that Dbf4 acts as a molecular scaffold to bind three separate protein kinases: Cdc7, Cdc5, and Rad53 (Figure 1). Binding of Cdc7 occurs through essential middle and C-terminal Dbf4 residues. Binding of Cdc5 and Rad53 occurs through Dbf4 N-terminal residues that have evolved a checkpoint effector role to mediate the response to DNA damage, replication fork stalling, and chromosome segregation defects. Many different types of tumors show increased levels of DDK, and inhibiting DDK causes the death of many types of tumor cells, but not normal cells. Because the ability of DDK to control multiple aspects of chromosome metabolism is likely conserved, it is crucial to understand these pathways in order to further the development of highly effective chemotherapeutic agents and interventions. 64

Publications by Year