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PMID: 8657125 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Ku86 defines the genetic defect and restores X-ray resistance and V(D)J recombination to complementation group 5 hamster cell mutants.

Molecular and cellular biology ·Vol. 16 ·No. 4 ·1996-04-00 ·Pages 1519-26

Errami A, Smider V, Rathmell WK, He DM, Hendrickson EA, Zdzienicka MZ, Chu G

Abstract

X-ray-sensitive hamster cells in complementation groups 4, 5, 6, and 7 are impaired for both double-strand break repair and V(D)J recombination. Here we show that in two mutant cell lines (XR-V15B and XR-V9B) from group 5, the genetic defects are in the gene encoding the 86-kDa subunit of the Ku autoantigen, a nuclear protein that binds to the double-stranded DNA ends. These mutants express Ku86 mRNA containing deletions of 138 and 252 bp, respectively, and the encoded proteins contain internal, in-frame deletions of 46 and 84 amino acids. Two X-ray-resistant revertants of XR-V15B expressed two Ku86 transcripts, one with and one without the deletion, suggesting that reversion occurred by activation of a silent wild-type allele. Transfection of full-length cDNA encoding hamster Ku86 into XR-V15B cells resulted in a complete rescue of DNA-end-binding (DEB) activity and Ku70 levels, suggesting that Ku86 stabilizes the Ku70 polypeptide. In addition, cells expressing wild-type levels of DEB activity were fully rescued for X-ray resistance and V(D)J recombination, whereas cells expressing lower levels of DEB activity were only partially rescued. Thus, Ku is an essential component of the pathway(s) utilized for the resolution of DNA double-strand breaks induced by either X rays or V(D)J recombination, and mutations in the Ku86 gene are responsible for the phenotype of group 5 cells.

MeSH Terms
Amino Acid Sequence Animals Antigens, Nuclear Autoantigens/genetics Base Sequence Cell Survival/radiation effects Cricetinae Cricetulus DNA Helicases DNA Nucleotidyltransferases/genetics DNA-Binding Proteins/genetics Genetic Complementation Test Ku Autoantigen Molecular Sequence Data Mutation Nuclear Proteins/genetics Radiation Tolerance Sequence Alignment VDJ Recombinases
Chemicals
Antigens, Nuclear Autoantigens DNA-Binding Proteins Nuclear Proteins DNA Nucleotidyltransferases VDJ Recombinases DNA Helicases XRCC5 protein, human Xrcc6 protein, human Ku Autoantigen
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Errami A
MGC-Department of Radiation Genetics and Chemical Mutagenesis, Leiden University, The Netherlands.
Smider V
Rathmell W K
He D M
Hendrickson E A
Zdzienicka M Z
Chu G
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.16.4.1519
Published
1996-04-00
Pages
1519-26
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC231136
Subset
IM
Grants
NIAID NIH HHS · AI35763 · United States
Databases
GENBANK
U40570
External Links
PubMed source DOI: 10.1128/MCB.16.4.1519 PMC Full Text View journal details
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