Last Updated: 5/6/2009

Research Interests
Cell cycle checkpoints represent positions of control that ensure the completion of essential events in the cell division cycle, provide more time for repair of DNA damage before replication, and stabilize replication forks under conditions of stress. Several familial cancer syndromes are due to inherited mutations in cell cycle checkpoint genes, and defects in checkpoint gene function lead to enhanced growth and genetic instability, two key features of malignant neoplasms. Research in the Kaufmann laboratory is concerned with determining the mechanisms whereby cell cycle checkpoints suppress human carcinogenesis.

Highlights of our work in recent years include identification of a new G2 checkpoint that couples the onset of mitosis to completion of chromatid decatenation following DNA replication, elucidation of the mechanisms of S checkpoint response to DNA damage caused by ultraviolet light, and identification of stereotypic patterns of changes in gene expression in response to damage to DNA.

We have recently characterized a checkpoint that functions during G2 to increase the fidelity of chromosomal segregation. Intertwined daughter chromatids that are produced during DNA replication must be separated by DNA topoisomerase II prior to chromatid separation in mitosis. Inhibition of topoisomerase II function in mitotic cells interferes with sister chromatid separation and induces polyploidy. We have found that G2 cells monitor the process of chromatid decatenation by topoisomerase II and actively inhibit mitosis until achieving a level of decatenation that is sufficient for successful mitosis. This decatenation checkpoint was shown to require the activity of the checkpoint kinases ATR and Chk1 which acted with the breast and ovarian cancer tumor suppressor gene BRCA1 to block the activation of Plk1. Plk1 is a protein kinase that phosphorylates cyclin B1 and induces the accumulation within nuclei of the mitosis-promoting factor MPF. MPF initiates and drives the process of mitosis. Thus the decatenation checkpoint delays the onset of mitosis when chromatids are insufficiently decatenated by blocking the intranuclear accumulation of MPF. Defects in decatenation checkpoint function can be expected to cause cells to enter mitosis with insufficient decatenation and thereby to enhance chromosomal missegregation leading to aneuploidy and polyploidy. We and others have shown that cancer cell lines display defects in the decatenation G2 checkpoint and a recent report showed that human stem cells express a defect this checkpoint. Defects in the decatenation G2 checkpoint may contribute to genomic instability in cancer stem cells.

Solar radiation is a ubiquitous environmental carcinogen causing in the USA about one million new cases of skin cancer each year. UVB wavelengths induce the formation of mutagenic and clastogenic (chromosome-breaking) pyrimidine dimers in DNA. Pyrimidine dimers arrest DNA synthesis by the normal replicative DNA polymerases, and mutations and chromosomal aberrations are thought to occur due to error-prone mechanisms of bypass of dimers.

The mechanisms of human S checkpoint response to UV-induced DNA damage have been elucidated. Unlike the S checkpoint response to ionizing radiation-induced DNA damage, which requires the checkpoint kinase ATM, the UV-induced S checkpoint was independent of ATM and rather required the ATM-related checkpoint kinase, ATR. ATR was shown to signal from sites of UV damage to phosphorylate and activate the transducer checkpoint kinase Chk1. The ATR- and Chk1-dependent intra-S checkpoint response to UV was shown not to involve degradation of Cdc25A nor inhibition of cyclin E/Cdk2 kinase, but rather appeared to affect the action of the Dbf4/Cdc7 kinase that is required for replicon initiation. Activation of Chk1 by ATR also was shown to require the Timeless-Tipin replication fork protection complex. This complex appears to recruit Chk1 to stalled replication forks via interaction with replication protein A. The Timeless-Tipin complex mediated not only the inhibition of replicon initiation, it also regulated the rate of DNA chain elongation in active replicons. This work established a new component of the S checkpoint response to sumlight-induced DNA damage.

Ionizing radiation is used in therapy for cancer where it induces DNA double strand breaks to kill cells. We used microarray technology to chart the global changes in gene expression in diploid human fibroblasts after treatment with a low therapeutic dose of ionizing radiation. Fibroblast lines displayed stereotypic responses to ionizing radiation that included rapid induction of targets of p53 transactivation and delayed repression of hundreds of cell cycle-regulated genes. Proliferating cells were transiently arrested behind a p53-dependent checkpoint in a state resembling G0 quiescence. This work demonstrated for the first time that the pattern of gene expression in ionizing radiation-damaged fibroblasts was that of quiescence and not senescence.

Publications
Chen,B, Simpson,DA, Zhou,Y, Mitra,A, Mitchell,DL, Cordeiro-Stone,M, and Kaufmann, WK, Human papilloma virus type16 E6 deregulates CHK1 and sensitizes human fibroblasts to environmental carcinogens independently of its effect on p53. Cell Cycle 2009, in press

Heard, J, Kaufmann, WK and Guan, X. A novel method for large tree visualization. (2009) Bioinformatics, PMID: 19129207

Tran, N, Qu, P, Simpson, DA, Lindsey-Boltz, L, Guan, X, Schmitt, CP, Ibrahim, J and Kaufmann, WK. (2009) In-silico construction of a protein interaction landscape for nucleotide excision repair. Cell Biochemistry and Biophysics, PMID: 19156361

Nikolaishvili-Feinberg N, Jenkins GS, Nevis KR, Staus DP, Scarlett CO, Unsal-Kamaz K, Kaufmann WK, Cordeiro-Stone M. (2008) Ubiquitylation of Proliferating Cell Nuclear Antigen and Recruitment of Human DNA Polymerase eta. Biochemistry. 47(13):4141-50

Kaufmann WK, Nevis KR, Qu P, Ibrahim JG, Zhou T, Zhou Y, Simpson DA, Helms-Deaton J, Cordeiro-Stone M, Moore DT, Thomas NE, Hao H, Liu Z, Shields JM, Scott GA and Sharpless, NE (2008) Defective cell cycle checkpoint functions in melanoma are associated with altered patterns of gene expression. J Invest Dermatol. 128:175-187.

Chou, J.W., Zhou, T., Kaufmann, W.K., Paules, R.S. and Bushel, P.R. (2007) Extracting gene expression patterns and identifying co-expressed genes from microarray data reveals biologically responsive processes. BMC Bioinformatics, 8, 427. PMCID: PMC2194742

Kesseler, K. J., W. K. Kaufmann, J. T. Reardon, T. C. Elston, and A. Sancar. (2007) A mathematical model for human nucleotide excision repair: Damage recognition by random order assembly and kinetic proofreading. J Theor Biol. 249(2):361-75

Zhou, T, Chou, J, Zhou, Y, Simpson, DA, Cao, F, Bushel, PR, Paules, RS, and Kaufmann, WK. (2007) ATM-dependent DNA damage checkpoint functions regulate gene expression in human fibroblasts, Molecular Cancer Research, 5:813-822.

Zhou, T, Chou, J, Mullen, T.E., Elkon, R, Zhou, Y, Simpson, D.A., Bushel, P.R., Paules, R.S., Lobenhofer, E.K., Hurban, P. and Kaufmann, W.K. (2007) Identification of primary transcriptional regulation of cell-cycle-regulated genes upon DNA damage. Cell Cycle, 6:972-981. PMCID: PMC2117899

nsal-Kamaz, K, Chastain, PD, Qu, P, Minoo, P, Cordeiro-Stone, M, Sancar, A, and Kaufmann, WK. (2007) The human Tim/Tipin complex coordinates an intra-S checkpoint response to UV that slows replication fork displacement. Mol. Cell. Biol. 27:3131-3142. PMCID: PMC1899931

Heffernan, TP, nsal-Kamaz, K, Heinloth, AN, Simpson, DA, Paules, RS, Sancar, A, Cordeiro-Stone, M, and Kaufmann, WK. (2007) Cdc7/Dbf4 and the Human S Checkpoint Response to UVC. J. Biol. Chem. 282:9458-9468. PMCID: PMC1839878

Shields, JM, Thomas, NE, Cregger, M, Berger, AJ, Leslie, M, Torrice, C, Hao, H, Penland, S, Arbiser, J, Scott, G, Zhou, T, Bar-Eli, M, Bear, JE, Der, CJ, Kaufmann, WK, Rimm DL, and Sharpless, NE. (2007) Lack of ERK mitogen-activated protein kinase signaling demonstrates a new type of melanoma, Cancer Research, 67:1502-1512.

E-mail: Bill_Kaufmann@med.unc.edu
Telephone: (919) 966-8209
FAX: (919) 966-9673
Address: 31-325 Lineberger Comprehensive Cancer Center, CB# 7295 Chapel Hill, NC
URL: www.med.unc.edu/patholog/kaufmann/

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