Nicholas Rhind, Ph.D.

Academic Role: Assistant Professor

Faculty Appointment(s) In:
   Biochemistry and Molecular Pharmacology

Other Affiliation(s):
   Cell Biology
   Interdisciplinary Graduate Program

Checkpoint Regulation of the Fission Yeast Cell Cycle

The main interest of my lab is checkpoint regulation of the cell cycle. Checkpoints are mechanisms that cells use to deal with problems during the cell cycle, such as DNA damage, replication errors, and misattachment of chromosomes to the mitotic spindle. By actively responding to these problems, cells can fix most of them. In contrast, cells that lack proper checkpoints are very sensitive to DNA damage and show increased rates of mutation and other chromosomal abnormalities. Loss of checkpoints is an important step in the development of cancer.

Most of the work in my lab focuses on the fission yeast Schizosaccharomyces pombe (Figure 1). S. pombe is a great organism for studying checkpoint regulation because it has a simple, well-understood cell cycle and is amenable to genetic, molecular and biochemical approaches. It has the added attraction that the mechanisms of cell cycle and checkpoint control in pombe are very similar to those used by human cells. In fact, much of what is known about human cell cycle checkpoints was first discovered in pombe. My lab is currently pursuing three general areas of checkpoint regulation in pombe.

The G2 DNA damage and DNA replication checkpoints are two closely related checkpoints that prevent cells from entering mitosis if DNA is damaged or incompletely replicated (Figure 2). By preventing mitosis, the checkpoints allow the cell to repair the damage or complete replication before dividing. Attempting mitosis with broken or unreplicated chromosomes would be most deleterious. While these checkpoints are understood at a broad level, the molecular mechanisms remain to be worked out. One particularly interesting aspect is the regulation of the mitotic inhibitor Mik1. Mik1 is a kinase that phosphorylates and inhibits Cdc2, the master regulator of mitosis. Mik1 plays important roles in both checkpoints, and is regulated by the checkpoints in at least three ways: at the level of transcript accumulation, at the level of protein accumulation, and at the level of enzyme activation. I am particularly interested to understand the transcriptional and post-translation regulation of Mik1 abundance and how this regulation relates to general checkpoint regulation of protein level.

The S-phase DNA damage checkpoint slows the rate of replication in response to DNA damage. The simple model is that this checkpoint prevents damaged DNA from being replicated before it is repaired. However, the checkpoint is clearly more subtle than that. Recent results from pombe and human cells suggest that this checkpoint may coordinate recombinational repair and replication during S-phase. We are characterizing the requirements for recombinational machinery in the checkpoint and using physical techniques to investigate the effect of the checkpoint on replication origin use and fork elongation.

The G1 size control checkpoint prevents cells from entering S-phase when they are too small. This raises the interesting question of how cells know how big they are. Several genetic projects are underway to identify the genes involved in cell size measurement and regulation, and to test various models of cell size control.

Figures

Figure 1.   The fission yeast Schizosaccharomyces pombe is a simple, single cell eukaryote that has proven to be an excellent model for cell cycle and checkpoint regulation. It divides by medial fission, distinguishing it from the budding yeast Saccharomyces cerevisiae, another popular lab yeast.

Figure 2.   A cartoon of the G2 DNA damage and replication checkpoint pathways. The ultimate target of these checkpoints is the tyrosine phosphorylation of Cdc2, the master regulator of mitosis. If Cdc2 is phosphorylated on Y15, the cell remains in G2. When Cdc2 Y15 is dephosphorylated, the cell enters mitosis and divides. Phosphorylation of Cdc2 Y15 is regulated by the Wee1 and Mik1 kinases and by Cdc25 phosphatase. The Checkpoint Rad proteins respond to DNA damage or incomplete replication and activate Chk1 or Cds1, two checkpoint kinases that prevent mitosis by inhibiting Cdc25 and by activating Mik1. The regulation of Cdc25 by the checkpoints has been well studied, but regulation of Mik1 is still poorly understood.

Representative Publications

S. Forsburg and N. Rhind (2006).  Basic methods for fission yeast. Yeast 23(3):173

P.K. Patel, B. Arcangioli, S.P. Baker, A. Bensimon and N. Rhind (2006).   DNA replication origins fire stochastically in fission yeast. Mol. Biol. Cell 17(1):308 

A. Sigova, N. Rhind and P. D. Zamore (2004). A single Argonaute protein mediates both transcriptional and post-transcriptional silencing in Schizosaccharomyces pombe. Genes and Development 18(19):2359

S. Sivakumar, M. Porter-Goff, P. K. Patel, K. Benoit and N. Rhind (2004). In vivo labeling of fission yeast DNA with thymidine and thymidine analogs.Methods 33(3):213

C. Chahwan, T. M. Nakamura, S. Sivakumar, P. Russell, N. Rhind (2003). The Fission Yeast Rad32(Mre11)-Rad50-Nbs1 Complex is Required for the S-phase DNA Damage Checkpoint. Mol. Cell. Biol. 23(18):6564

N. Rhind and P. Russell (2001). The Roles of the Mitotic Inhibitors Wee1 and Mik1 in the G2 DNA Damage and Replication Checkpoints.   Mol. Cell. Biol. 21(5): 1499

B.A. Baber-Furnari, N. Rhind, M.N. Boddy, P. Shanahan, A. Lopez-Girona, and P. Russell (2000). Damage checkpoint insurance: Regulation of mitotic inhibitor Mik1 helps to enforce the DNA damage checkpoint . Mol. Biol. Cell 11(1): 1

N. Rhind, B.A. Baber-Furnari, A. Lopez-Girona, M.N. Boddy, J.-M. Brondello, B. Moser, P. Shanahan, A. Blasina, C. McGowan, and P. Russell (2000). DNA Damage Checkpoint Control of Mitosis in Fission Yeast. Cold Spring Harbor Symposia on Quantitative Biology , Volume LXIV

N. Rhind and P. Russell (2000). Chk1 and Cds1: linchpins of the DNA damage and replication checkpoint pathways. J. Cell Sci. 113(22):3889

N. Rhind  and P. Russell (2000). Checkpoints: It takes more than time to heal some wounds . Curr. Biol. 10(24): R912

N. Rhind and P. Russell (1998). Cdc2 Activation is Inhibited by Tyrosine Phosphorylation During a Replication Checkpoint in Schizosaccharomyces pombe. Mol. Cell Biol. 18(7): 3782.

N. Rhind and P. Russell (1998). The Schizosaccharomyces pombe S-phase DNA Damage Checkpoint Differentiates Between Different Types of DNA Damage . Genetics 149: 1729

N. Rhind and P. Russell (1998). Mitotic DNA damage and replication checkpoints in yeast.   Curr. Opin. Cell Biol. 10(6): 749

N. Rhind, B. Furnari, and P. Russell (1997). Cdc2 Tyrosine Phosphorylation is Required for the DNA Damage Checkpoint in Fission Yeast.   Genes & Development 11(4): 504.

B. Furnari, N. Rhind, and P. Russell (1997). Cdc25 Mitotic Inducer Targeted by Chk1 DNA Damage Checkpoint Kinase . Science 277: 1495.

Potential Rotation Projects

1) Coordination of replication and recombination by the S-phase DNA damage checkpoint.

Cells slow replication in the presence of DNA damage.  In humans and fission yeast, this slowing requires the MRN recombinational repair nuclease, consisting of Mre11, Rad50 and Nbs1. This raises the possibility that the slowing of replication is caused by induced recombinational repair.  We are testing this hypothesis using a combination of genetics, to measure induction of recombination in response to DNA damage during S-phase, biochemistry, to investigate the roles and regulation MRN's DNA binding and nuclease activities, and physical techniques, to measure the effect of the checkpoint of replication fork rate and origin use.  One possible project would be to study how the nuclease activity of MRN is regulated by checkpoint activation.

2) Regulation of S-phase transcription by the replication checkpoint.

The replication checkpoint prevents mitosis until replication is complete.It also up regulates the expression of genes required to complete replication, such as enzymes involved in nucleotide synthesis and DNA polymerization.  Using micro-array analysis, we have shown that the checkpoint up regulates most of the genes normally expressed during S-phase, suggesting that the checkpoint acts through MBF, the S-phase transcription factor. We are currently investigating how the checkpoint regulates MBF.  Interestingly, the one class of genes not regulated by the checkpoint are the histones.  Somehow, a checkpoint independent mechanism monitors replication and regulates the histone so that they are only transcribed when replication is occurring.  An interesting rotation project would be to explore the mechanism of this regulation.

Academic Background

Nick Rhind received his Ph.D. from the Department of Molecular and Cell Biology at U.C. Berkeley in 1995, where he studied worm sex determination with Barbara Meyer. He was a Leukemia and Lymphoma Society post-doctoral fellow with Paul Russell at the Scripps Research Institute from 1996 to 2001.

Office: Research 904, Lab 940D&E
Phone: 508-856-8316
E-mail: Nick.Rhind@umassmed.edu
Keywords: Checkpoints, DNA Recombination, Cell Cycle, DNA Replication, Genetics

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