TY - JOUR
T1 - Gene-specific selection against experimental fanconi anemia gene inactivation in human cancer
AU - Gallmeier, Eike
AU - Hucl, Tomas
AU - Calhoun, Eric S.
AU - Cunningham, Steven C.
AU - Bunz, Fred
AU - Brody, Jonathan R.
AU - Kern, Scott E.
N1 - Funding Information:
ATR, BRCA2, FANCC, FANCD2, FANCG, Fanconi anemia (FA) is a rare, recessively inherited disorder consisting of congenital fanconi anemia, pancreatic cancer skeletal abnormalities, progressive bone marrow failure, and increased cancer risk.1 The disease comprises at least 13 complementation groups (FANCA, FANCB, FANCC, BRCA2/FANCD1, FANCD2, FANCE, FANCF, FANCG, FANCI, BRIP1/FANCJ, FANCL, FANCM and PALB2/FANCN). All of these genes have been identified except for FANCI.2-8The FA genes appear to act in a common pathway, distal parts of which interact with regulators of cell cycle control and DNA repair, especially the repair of DNA interstrand-crosslinks and double-strand breaks.1There are at least three distinguish-We would like to thank L. A. Morsberger from able compartments of the FA pathway. The upstream element consists of a protein core the Johns Hopkins Cytogenetics core facility complex, assembled by FANCA, FANCB, FANCC, FANCE, FANCF, FANCG, FANCL, for the cytogenetic analyses, B. Vogelstein for FANCM, and a yet unidentified 100kDa protein FAAP100.9 This complex enables the providing the selection cassette, and M. Kohli, monoubiquitination of the central protein FANCD2, which subsequently translocates C. Rago and C. Lengauer for helpful discus-to nuclear DNA repair foci and colocalizes with various DNA repair proteins including sions. This work was funded by NIH grant BRCA2.10-17 Monoubiquitinated FANCD2 interacts directly with BRCA2 and promotes CA 62924. E.G. was supported by grants from loading of BRCA2 into chromatin complexes.17 While defects in any of the proximal the Deutsche Forschungsgemeinschaft (DFG FA core proteins preclude FANCD2 monoubiquitination, BRCA2 is not required for
PY - 2007/5
Y1 - 2007/5
N2 - The Fanconi anemia (FA) gene family comprises at least 12 genes interacting in a common pathway involved in DNA repair. To gain insight into the role of FA gene inactivation occurring in tumors among the general population, we endogenously targeted in cancer cells four FA genes that act at different stages of the FA pathway. After successful monoallelic deletion of all genes, the sequential homozygous deletion was achieved only for FANCC and FANCG, acting upstream, but not for BRCA2 or FANCD2, acting downstream in the FA pathway. Targeting of the second allele in rgeting BRCA2 and FANCD2 heterozygote clones resulted in redeletion exclusively of the already defective allele in multiple instances (13x concerning BRCA2, 25x concerning FANCD2), strongly suggesting a detrimental phenotype. Unlike complete FANCD2 disruption, the mere reduction of FANCD2 protein levels had no discernible effect. In addition, we confirmed that human cancer cells harboring the Seckel ATR mutation display impaired FANCD2 monoubiquitination and FANCD2 nuclear focus formation, as well as an increased sensitivity to DNA interstrand-crosslinking agents. Nevertheless, these cells were viable, indicating an ATR-independent function of FANCD2, distinct from its major known functions, to be responsible for the detrimental effects of FANCD2 loss. In conclusion, we established the downstream FA genes FANCD2 and BRCA2 to represent particularly vulnerable parts of the FA pathway, providing direct evidence for the paradoxical assumption that their inactivation could be predominantly selected against in cancer cells. This would explain why certain FA gene defects, despite an apparent selection for FA pathway inactivation in cancer, are rarely observed in tumors among the general population.
AB - The Fanconi anemia (FA) gene family comprises at least 12 genes interacting in a common pathway involved in DNA repair. To gain insight into the role of FA gene inactivation occurring in tumors among the general population, we endogenously targeted in cancer cells four FA genes that act at different stages of the FA pathway. After successful monoallelic deletion of all genes, the sequential homozygous deletion was achieved only for FANCC and FANCG, acting upstream, but not for BRCA2 or FANCD2, acting downstream in the FA pathway. Targeting of the second allele in rgeting BRCA2 and FANCD2 heterozygote clones resulted in redeletion exclusively of the already defective allele in multiple instances (13x concerning BRCA2, 25x concerning FANCD2), strongly suggesting a detrimental phenotype. Unlike complete FANCD2 disruption, the mere reduction of FANCD2 protein levels had no discernible effect. In addition, we confirmed that human cancer cells harboring the Seckel ATR mutation display impaired FANCD2 monoubiquitination and FANCD2 nuclear focus formation, as well as an increased sensitivity to DNA interstrand-crosslinking agents. Nevertheless, these cells were viable, indicating an ATR-independent function of FANCD2, distinct from its major known functions, to be responsible for the detrimental effects of FANCD2 loss. In conclusion, we established the downstream FA genes FANCD2 and BRCA2 to represent particularly vulnerable parts of the FA pathway, providing direct evidence for the paradoxical assumption that their inactivation could be predominantly selected against in cancer cells. This would explain why certain FA gene defects, despite an apparent selection for FA pathway inactivation in cancer, are rarely observed in tumors among the general population.
KW - ATR
KW - BRCA2
KW - FANCC
KW - FANCD2
KW - FANCG
KW - Fanconi anemia
KW - Pancreatic cancer
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U2 - 10.4161/cbt.6.5.3978
DO - 10.4161/cbt.6.5.3978
M3 - Article
C2 - 17387268
AN - SCOPUS:34548247158
SN - 1538-4047
VL - 6
SP - 654
EP - 660
JO - Cancer Biology and Therapy
JF - Cancer Biology and Therapy
IS - 5
ER -