A patient-derived mutant form of the Fanconi anemia protein, FANCA, is defective in nuclear accumulation.
Fanconi anemia (FA) is an autosomal recessive cancer susceptibility syndrome with at least eight complementation groups (A-H). Three FA genes, corresponding to complementation groups A, C, and G, have been cloned, but the function of the encoded FA proteins remains unknown. We recently demonstrated that the FANCA and FANCC proteins bind and form a nuclear complex. In the current study, we identified a homozygous mutation in the FANCA gene (3329A>C) in an Egyptian FA patient from a consanguineous family. This mutant FANCA allele is predicted to encode a mutant FANCA protein, FANCA(H1110P), in which histidine 1110 is changed to proline. Initially, we characterized the FANCA(H1110P) protein, expressed in an Epstein Barr virus (EBV)-immortalized lymphoblast line derived from the patient. Unlike wild-type FANCA protein expressed in normal lymphoblasts, FANCA(H1110P) was not phosphorylated and failed to bind to FANCC. To test directly the effect of this mutation on FANCA function, we used retroviral-mediated transduction to express either wild-type FANCA or FANCA(H1110P) protein in the FA-A fibroblast line, GM6914. Unlike wild-type FANCA, the mutant protein failed to complement the mitomycin C sensitivity of these cells. In addition, the FANCA(H1110P) protein was defective in nuclear accumulation in the transduced cells. The characteristics of this mutant protein underscore the importance of FANCA phosphorylation, FANCA/FANCC binding, and nuclear accumulation in the function of the FA pathway.
Kupfer
G
G
Dana-Farber Cancer Institute, and Department of Pediatrics, Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Naf
D
D
Garcia-Higuera
I
I
Wasik
J
J
Cheng
A
A
Yamashita
T
T
Tipping
A
A
Morgan
N
N
Mathew
C G
CG
D'Andrea
A D
AD
D019943:Amino Acid Substitution / Q000235:genetics; D018797:Cell Cycle Proteins; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D005199:Fanconi Anemia / Q000235:genetics* / Q000378:metabolism; D052218:Fanconi Anemia Complementation Group C Protein; D051856:Fanconi Anemia Complementation Group Proteins; D015870:Gene Expression; D005816:Genetic Complementation Test; D006801:Humans; D015151:Immunoblotting; D008214:Lymphocytes / Q000737:chemistry; D009687:Nuclear Proteins; D010766:Phosphorylation; D017354:Point Mutation; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C497651:FANCC protein, human; D052218:Fanconi Anemia Complementation Group C Protein; D051856:Fanconi Anemia Complementation Group Proteins; D009687:Nuclear Proteins; D011506:Proteins