COMPLEMENTATION OF THE NIJMEGEN BREAKAGE SYNDROME DEFECT

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Nijmegen Breakage syndrome (NBS) is a rare inherited human disorder characterized by immune deficiencies, microcephaly, developmental delay and cancer susceptibility. Patients affected by NBS do not display cerebellar ataxia or telangiectasia, two prominent symptoms in Ataxia telangiectasia patients. However at the cellular level NBS and AT cells are indistinguishable showing marked hypersensitivity to ionizing radiation with no measurable defect in DNA double-strand break repair; radioresistant DNA synthesis and chromosomal instability. Two lines of evidence support the assumption that the gene for NBS is distinct from the AT-gene: Microcell-mediated transfer of human chromosome 11 (carrying the AT-gene) into NBS-cells fails to complement the phenotype [19]/ Genetic evidence from five different families with two affected siblings each who inherited different alleles from their parents for the region on chromosome 11q containing the AT gene, also excludes the ATM gene as the primary defect in NBS [24,25]. Our overall goal in this proposal is to further characterize cells from patients with NBS families is restricted by the rarity of this disease. Hence we will create hybrids with NBS cells retaining mouse chromosome fragments, which will be selected for radioresistance under low dose rate irradiation. Subsequently the complementing chromosomal region can be mapped by FISH and PCR. Anonymous DNA markers spaced in 3.5 centiMorgan (about 3.5 Megabases) distances will be tested for the presences or absence in radioresistant and radiosensitive hybrids. This type of analysis allows a statistical deduction of the chromosome region containing the NBS gene. By pinpointing the localization of the NBS gene to a minimal region (less than one Megabase) we should be able to identify candidate cDNAs using either a traditional cloning approach or by utilizing the accumulating information of the Human Genome Project. In addition, we will further characterize the DNA damage response following ionizing radiation and will study the kinetics of chromosome repair in NBS cells. Identification of the NBS gene will add to our understanding of the pathways involved in the response of cells to ionizing radiation and to the factors governing radiation sensitivity.