Rad18-mediated translesion synthesis of bulky DNA adducts is coupled to activation of the Fanconi anemia DNA repair pathway.
Fanconi anemia (FA) is a cancer susceptibility syndrome characterized by sensitivity to DNA-damaging agents. The FA proteins (FANCs) are implicated in DNA repair, although the precise mechanisms by which FANCs process DNA lesions are not fully understood. An epistatic relationship between the FA pathway and translesion synthesis (TLS, a post-replication DNA repair mechanism) has been suggested, but the basis for cross-talk between the FA and TLS pathways is poorly understood. We show here that ectopic overexpression of the E3 ubiquitin ligase Rad18 (a central regulator of TLS) induces DNA damage-independent mono-ubiquitination of proliferating cell nuclear antigen (PCNA) (a known Rad18 substrate) and FANCD2. Conversely, DNA damage-induced mono-ubiquitination of both PCNA and FANCD2 is attenuated in Rad18-deficient cells, demonstrating that Rad18 contributes to activation of the FA pathway. WT Rad18 but not an E3 ubiquitin ligase-deficient Rad18 C28F mutant fully complements both PCNA ubiquitination and FANCD2 activation in Rad18-depleted cells. Rad18-induced mono-ubiquitination of FANCD2 is not observed in FA core complex-deficient cells, demonstrating that Rad18 E3 ligase activity alone is insufficient for FANCD2 ubiquitylation. Instead, Rad18 promotes FA core complex-dependent FANCD2 ubiquitination in a manner that is secondary to PCNA mono-ubiquitination. Taken together, these results demonstrate a novel Rad18-dependent mechanism that couples activation of the FA pathway with TLS.
Song
Ihn Young
IY
Graduate Program in Genetics and Genomics, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Palle
Komaraiah
K
Gurkar
Aditi
A
Tateishi
Satoshi
S
Kupfer
Gary M
GM
Vaziri
Cyrus
C
D019943:Amino Acid Substitution; D002460:Cell Line; D018736:DNA Adducts / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005199:Fanconi Anemia / Q000235:genetics / Q000378:metabolism*; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020125:Mutation, Missense; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
D018736:DNA Adducts; D004268:DNA-Binding Proteins; C499905:FANCD2 protein, human; D052236:Fanconi Anemia Complementation Group D2 Protein; D018809:Proliferating Cell Nuclear Antigen; C485954:RAD18 protein, human; D044767:Ubiquitin-Protein Ligases