Functional capacity of XRCC1 protein variants identified in DNA repair-deficient Chinese hamster ovary cell lines and the human population.
XRCC1 operates as a scaffold protein in base excision repair, a pathway that copes with base and sugar damage in DNA. Studies using recombinant XRCC1 proteins revealed that: a C389Y substitution, responsible for the repair defects of the EM-C11 CHO cell line, caused protein instability; a V86R mutation abolished the interaction with POLbeta, but did not disrupt the interactions with PARP-1, LIG3alpha and PCNA; and an E98K substitution, identified in EM-C12, reduced protein integrity, marginally destabilized the POLbeta interaction, and slightly enhanced DNA binding. Two rare (P161L and Y576S) and two frequent (R194W and R399Q) amino acid population variants had little or no effect on XRCC1 protein stability or the interactions with POLbeta, PARP-1, LIG3alpha, PCNA or DNA. One common population variant (R280H) had no pronounced effect on the interactions with POLbeta, PARP-1, LIG3alpha and PCNA, but did reduce DNA-binding ability. When expressed in HeLa cells, the XRCC1 variants-excluding E98K, which was largely nucleolar, and C389Y, which exhibited reduced expression-exhibited normal nuclear distribution. Most of the protein variants, including the V86R POLbeta-interaction mutant, displayed normal relocalization kinetics to/from sites of laser-induced DNA damage: except for E98K and C389Y, and the polymorphic variant R280H, which exhibited a slightly shorter retention time at DNA breaks.
Berquist
Brian R
BR
Laboratory of Molecular Gerontology, National Institute on Aging, NIH, Baltimore, MD 21224, USA.
Singh
Dharmendra Kumar
DK
Fan
Jinshui
J
Kim
Daemyung
D
Gillenwater
Elizabeth
E
Kulkarni
Avanti
A
Bohr
Vilhelm A
VA
Ackerman
Eric J
EJ
Tomkinson
Alan E
AE
Wilson
David M
DM
D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000378:metabolism; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D000076105:X-ray Repair Cross Complementing Protein 1
D004268:DNA-Binding Proteins; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human; D004247:DNA
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