Characterization of a mutant calcineurin A alpha gene expressed by EL4 lymphoma cells.
The calmodulin-stimulated phosphatase calcineurin plays a critical role in calcium-dependent T-lymphocyte activation pathways. Here, we report the identification of a missense mutation in the calcineurin A alpha gene expressed by EL4 T-lymphoma cells. This mutation changes an evolutionarily conserved aspartic acid to asparagine within the autoinhibitory domain of the calcineurin A alpha protein. A comparison of wild-type and mutant autoinhibitory peptides indicates that this amino acid substitution greatly reduces inhibition of calcineurin phosphatase activity. Additional peptide inhibition studies support a pseudosubstrate model of autoinhibitory function, in which the conserved aspartic acid residue may serve as a molecular mimic of either phosphoserine or phosphothreonine. Expression of the mutant calcineurin appears to affect cellular signal transduction pathways, as EL4 cells can be activated by suboptimal concentrations of calcium ionophore in the presence of phorbol esters. Moreover, this phenotype can be transferred to Jurkat T cells by transfection of the mutated calcineurin gene. These findings implicate a conserved aspartic acid in the mechanism of calcineurin autoinhibition and suggest that mutation of this residue is associated with aberrant calcium-dependent signaling in vivo.
Fruman
D A
DA
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Pai
S Y
SY
Burakoff
S J
SJ
Bierer
B E
BE
D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000235:genetics; D001224:Aspartic Acid / Q000235:genetics; D001483:Base Sequence; D019703:Calcineurin; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000494:pharmacology; D002148:Calmodulin-Binding Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D007376:Interleukin-2 / Q000096:biosynthesis; D015759:Ionomycin / Q000494:pharmacology; D016399:Lymphoma, T-Cell / Q000235:genetics*; D051379:Mice; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000235:genetics; D013329:Structure-Activity Relationship; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D007376:Interleukin-2; D010446:Peptide Fragments; D011994:Recombinant Proteins; D001224:Aspartic Acid; D015759:Ionomycin; D001216:Asparagine; D019703:Calcineurin; D010749:Phosphoprotein Phosphatases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium