Analysis of cells harboring a putative DNA repair gene reveals a lack of evidence for a second independent xeroderma pigmentosum group A correcting gene

Christopher J. Jones, Robert (Stephen) Lloyd, Richard D. Wood

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The UV hypersensitivity of xeroderma pigmentosum (XP) complementation group A cells is restored to near-normal by transfection of the XPA gene located on human chromosome 9. However, it has been reported that a cosmid related to a cDNA on chromosome 8 is also able to partially correct the UV sensitivity of XP-A cells. We describe here an investigation of a representative cosmid transfectant, denoted 2-0-A2. Whole cell extracts prepared from a 2-0-A2 cells carried out DNA repair synthesis in vitro that was in the normal range, consistent with their UV-resistant phenotype. Immunoblotting indicated that 2-0-A2 cells expressed full-length XPA protein. This was unexpected because the 2-0-A2 cell line was thought to have been isolated by transfection of a cell line derived from patient XP2OS, and a known homozygous mutation in XP2OS prevents expression of XPA gene product. This mutation creates an AlwNI restriction endonuclease cleavage site in XPA and was not present in 2-0-A2. These results prompted and RFLP analysis which revealed that the 2-0-A2 cell line was not derived from XP2OS but from another line that fails to express XPA protein, XP12BE. It appears that the significant UV-resistance and DNA repair capacity of 2-0-A2 can be ascribed to the re-expression of XPA in XP12BE, and it is unnecessary to postulate a second XP-A complementing gene to explain the results.

Original languageEnglish (US)
Pages (from-to)159-164
Number of pages6
JournalMutation Research Letters
Volume324
Issue number4
DOIs
StatePublished - 1994
Externally publishedYes

Fingerprint

Xeroderma Pigmentosum
varespladib methyl
DNA Repair
Cosmids
Repair
Genes
Cells
Chromosomes
DNA
DNA Restriction Enzymes
Cell Line
Proteins
Complementary DNA
Transfection
Chromosomes, Human, Pair 8
Chromosomes, Human, Pair 9
Mutation
Human Chromosomes
Cell Extracts
Immunoblotting

Keywords

  • DNA repair
  • Human
  • Ultraviolet light
  • Xeroderma pigmentosum
  • XPA

ASJC Scopus subject areas

  • Genetics
  • Toxicology

Cite this

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title = "Analysis of cells harboring a putative DNA repair gene reveals a lack of evidence for a second independent xeroderma pigmentosum group A correcting gene",
abstract = "The UV hypersensitivity of xeroderma pigmentosum (XP) complementation group A cells is restored to near-normal by transfection of the XPA gene located on human chromosome 9. However, it has been reported that a cosmid related to a cDNA on chromosome 8 is also able to partially correct the UV sensitivity of XP-A cells. We describe here an investigation of a representative cosmid transfectant, denoted 2-0-A2. Whole cell extracts prepared from a 2-0-A2 cells carried out DNA repair synthesis in vitro that was in the normal range, consistent with their UV-resistant phenotype. Immunoblotting indicated that 2-0-A2 cells expressed full-length XPA protein. This was unexpected because the 2-0-A2 cell line was thought to have been isolated by transfection of a cell line derived from patient XP2OS, and a known homozygous mutation in XP2OS prevents expression of XPA gene product. This mutation creates an AlwNI restriction endonuclease cleavage site in XPA and was not present in 2-0-A2. These results prompted and RFLP analysis which revealed that the 2-0-A2 cell line was not derived from XP2OS but from another line that fails to express XPA protein, XP12BE. It appears that the significant UV-resistance and DNA repair capacity of 2-0-A2 can be ascribed to the re-expression of XPA in XP12BE, and it is unnecessary to postulate a second XP-A complementing gene to explain the results.",
keywords = "DNA repair, Human, Ultraviolet light, Xeroderma pigmentosum, XPA",
author = "Jones, {Christopher J.} and Lloyd, {Robert (Stephen)} and Wood, {Richard D.}",
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T1 - Analysis of cells harboring a putative DNA repair gene reveals a lack of evidence for a second independent xeroderma pigmentosum group A correcting gene

AU - Jones, Christopher J.

AU - Lloyd, Robert (Stephen)

AU - Wood, Richard D.

PY - 1994

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N2 - The UV hypersensitivity of xeroderma pigmentosum (XP) complementation group A cells is restored to near-normal by transfection of the XPA gene located on human chromosome 9. However, it has been reported that a cosmid related to a cDNA on chromosome 8 is also able to partially correct the UV sensitivity of XP-A cells. We describe here an investigation of a representative cosmid transfectant, denoted 2-0-A2. Whole cell extracts prepared from a 2-0-A2 cells carried out DNA repair synthesis in vitro that was in the normal range, consistent with their UV-resistant phenotype. Immunoblotting indicated that 2-0-A2 cells expressed full-length XPA protein. This was unexpected because the 2-0-A2 cell line was thought to have been isolated by transfection of a cell line derived from patient XP2OS, and a known homozygous mutation in XP2OS prevents expression of XPA gene product. This mutation creates an AlwNI restriction endonuclease cleavage site in XPA and was not present in 2-0-A2. These results prompted and RFLP analysis which revealed that the 2-0-A2 cell line was not derived from XP2OS but from another line that fails to express XPA protein, XP12BE. It appears that the significant UV-resistance and DNA repair capacity of 2-0-A2 can be ascribed to the re-expression of XPA in XP12BE, and it is unnecessary to postulate a second XP-A complementing gene to explain the results.

AB - The UV hypersensitivity of xeroderma pigmentosum (XP) complementation group A cells is restored to near-normal by transfection of the XPA gene located on human chromosome 9. However, it has been reported that a cosmid related to a cDNA on chromosome 8 is also able to partially correct the UV sensitivity of XP-A cells. We describe here an investigation of a representative cosmid transfectant, denoted 2-0-A2. Whole cell extracts prepared from a 2-0-A2 cells carried out DNA repair synthesis in vitro that was in the normal range, consistent with their UV-resistant phenotype. Immunoblotting indicated that 2-0-A2 cells expressed full-length XPA protein. This was unexpected because the 2-0-A2 cell line was thought to have been isolated by transfection of a cell line derived from patient XP2OS, and a known homozygous mutation in XP2OS prevents expression of XPA gene product. This mutation creates an AlwNI restriction endonuclease cleavage site in XPA and was not present in 2-0-A2. These results prompted and RFLP analysis which revealed that the 2-0-A2 cell line was not derived from XP2OS but from another line that fails to express XPA protein, XP12BE. It appears that the significant UV-resistance and DNA repair capacity of 2-0-A2 can be ascribed to the re-expression of XPA in XP12BE, and it is unnecessary to postulate a second XP-A complementing gene to explain the results.

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