The Fanconi anemia group C gene product is located in both the nucleus and cytoplasm of human cells

Maureen Hoatlin, Tracy A. Christianson, Winnie W. Keeble, Adam T. Hammond, Yu Zhi, Michael Heinrich, Paula A. Tower, Grover C. Bagby

Research output: Contribution to journalArticle

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Abstract

The Fanconi anemia (FA) complementation group C (FAC) protein gene encodes a cytoplasmic protein with a predicted M(r) of 63,000. The protein's function is unknown, but it has been hypothesized that it either mediates resistance to DNA cross-linking agents or facilitates repair after exposure to such factors. The protein also plays a permissive role in the growth of colony-forming unit-granulocyte/macrophage (CFU-GM), burst-forming unit- erythroid (BFU-E), and CFU-erythroid (CFU-E). Attributing a specific function to this protein requires an understanding of its intracellular location. Recognizing that prior study has established the functional importance of its cytoplasmic location, we tested the hypothesis that FAC protein can also be found in the nucleus. Purified recombinant Escherichia coli-derived FAC antigens were used to create antisera able to specifically identify an M(r) = 58,000 protein in lysates from human Epstein-Barr virus (EBV)-transformed cell lines by immunoblot analysis. Subcellular fractionation of the cell lysates followed by immunoblot analysis revealed that the majority of the FAC protein was cytoplasmic, as reported previously; however, approximately 10% of FAC protein was reproducibly detected in nuclear fractions. These results were reproducible by two different fractionation methods, and included markers to control for contamination of nuclear fractions by cytoplasmic proteins. Moreover, confocal image analysis of human 293 cells engineered to express FAC clearly demonstrated that FAC protein is located in both cytoplasmic and nuclear compartments, consistent with data obtained from fractionation of the FA cell lines. Finally, complementation of the FAC defect using retroviral-mediated gene transfer resulted in a substantial increase in nuclear FAC protein. Therefore, while cytoplasmic localization of this protein appears to be functionally important, it may also exert some essential nuclear function.

Original languageEnglish (US)
Pages (from-to)1418-1425
Number of pages8
JournalBlood
Volume91
Issue number4
StatePublished - Feb 15 1998
Externally publishedYes

Fingerprint

Fanconi Anemia
Cytoplasm
Genes
Cells
Proteins
Fractionation
Fanconi Anemia Complementation Group C Protein
Cell Fractionation
Granulocyte-Macrophage Progenitor Cells
Erythroid Precursor Cells
Transformed Cell Line
Gene transfer
Macrophages
Nuclear Proteins
Human Herpesvirus 4
Viruses
Immune Sera
Image analysis
Escherichia coli
Contamination

ASJC Scopus subject areas

  • Hematology

Cite this

The Fanconi anemia group C gene product is located in both the nucleus and cytoplasm of human cells. / Hoatlin, Maureen; Christianson, Tracy A.; Keeble, Winnie W.; Hammond, Adam T.; Zhi, Yu; Heinrich, Michael; Tower, Paula A.; Bagby, Grover C.

In: Blood, Vol. 91, No. 4, 15.02.1998, p. 1418-1425.

Research output: Contribution to journalArticle

Hoatlin, M, Christianson, TA, Keeble, WW, Hammond, AT, Zhi, Y, Heinrich, M, Tower, PA & Bagby, GC 1998, 'The Fanconi anemia group C gene product is located in both the nucleus and cytoplasm of human cells', Blood, vol. 91, no. 4, pp. 1418-1425.
Hoatlin M, Christianson TA, Keeble WW, Hammond AT, Zhi Y, Heinrich M et al. The Fanconi anemia group C gene product is located in both the nucleus and cytoplasm of human cells. Blood. 1998 Feb 15;91(4):1418-1425.
Hoatlin, Maureen ; Christianson, Tracy A. ; Keeble, Winnie W. ; Hammond, Adam T. ; Zhi, Yu ; Heinrich, Michael ; Tower, Paula A. ; Bagby, Grover C. / The Fanconi anemia group C gene product is located in both the nucleus and cytoplasm of human cells. In: Blood. 1998 ; Vol. 91, No. 4. pp. 1418-1425.
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abstract = "The Fanconi anemia (FA) complementation group C (FAC) protein gene encodes a cytoplasmic protein with a predicted M(r) of 63,000. The protein's function is unknown, but it has been hypothesized that it either mediates resistance to DNA cross-linking agents or facilitates repair after exposure to such factors. The protein also plays a permissive role in the growth of colony-forming unit-granulocyte/macrophage (CFU-GM), burst-forming unit- erythroid (BFU-E), and CFU-erythroid (CFU-E). Attributing a specific function to this protein requires an understanding of its intracellular location. Recognizing that prior study has established the functional importance of its cytoplasmic location, we tested the hypothesis that FAC protein can also be found in the nucleus. Purified recombinant Escherichia coli-derived FAC antigens were used to create antisera able to specifically identify an M(r) = 58,000 protein in lysates from human Epstein-Barr virus (EBV)-transformed cell lines by immunoblot analysis. Subcellular fractionation of the cell lysates followed by immunoblot analysis revealed that the majority of the FAC protein was cytoplasmic, as reported previously; however, approximately 10{\%} of FAC protein was reproducibly detected in nuclear fractions. These results were reproducible by two different fractionation methods, and included markers to control for contamination of nuclear fractions by cytoplasmic proteins. Moreover, confocal image analysis of human 293 cells engineered to express FAC clearly demonstrated that FAC protein is located in both cytoplasmic and nuclear compartments, consistent with data obtained from fractionation of the FA cell lines. Finally, complementation of the FAC defect using retroviral-mediated gene transfer resulted in a substantial increase in nuclear FAC protein. Therefore, while cytoplasmic localization of this protein appears to be functionally important, it may also exert some essential nuclear function.",
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