Functional domain of NBS1 for telomere replication

Full text: Nijmegen breakage syndrome (NBS), characterized by high sensitivity to ionizing radiation and predisposition to lymphoid cancer, is phenotypically similar to ataxia telangiectasia (AT). The gene mutated in NBS, NBS1, functions in both DNA repair and telomere maintenance. NBS1 has three functional domains known as FHA /BRCT domain, phosphorylation domain and MRE11 binding domain. To examine how these functional domains are participating in telomere maintenance, we analyzed the telomere length and the interaction of NBS1 with telomere binding protein TRF2 by using NBS cell lines transfected with several mutants of NBS1 cDNA. First, the terminal restriction fragment length (TRF) in the primary cells from an NBS patient and an AT patient at different population doubling levels were determined by southern blot analysis. NBS primary cells showed short TRF length comparable with that of AT primary cells. On the other hand, SV40-transformed NBS cell lines, maintaining the minimum size of telomere, elongated TRF length after transfection with full length NBS1 cDNA and S278/343A (mutated in phosphorylation domain) NBS1 cDNA, whereas the TRF length remained short in all subclones transfected with truncated NBS1 cDNA. These results suggested that phosphorylation of Ser278/Ser343 is not associated with the telomere maintenance of NBS1. Our results also showed that NBS1 interacted with TRF2 particularly in G1/early S phase. This cell cycle dependent interaction was detected in both AT cells and NBS cells transfected with S278/343A NBS1 cDNA. These evidences indicate that NBS1 functions in telomere maintenance through ATM-independent pathway. We are now analyzing the regulatory factor for this cell cycle dependent interaction of NBS1 with TRF2