Post-translational arginylation of calreticulin: a new isospecies of calreticulin component of stress granules.
Post-translational arginylation consists of the covalent union of an arginine residue to a Glu, Asp, or Cys amino acid at the N-terminal position of proteins. This reaction is catalyzed by the enzyme arginyl-tRNA protein transferase. Using mass spectrometry, we have recently demonstrated in vitro the post-translational incorporation of arginine into the calcium-binding protein calreticulin (CRT). To further study arginylated CRT we raised an antibody against the peptide (RDPAIYFK) that contains an arginine followed by the first 7 N-terminal amino acids of mature rat CRT. This antibody specifically recognizes CRT obtained from rat soluble fraction that was arginylated in vitro and also recognizes endogenous arginylated CRT from NIH 3T3 cells in culture, indicating that CRT arginylation takes place in living cells. Using this antibody we found that arginylation of CRT is Ca2+-regulated. In vitro and in NIH 3T3 cells in culture, the level of arginylated CRT increased with the addition of a Ca2+ chelator to the medium, whereas a decreased arginine incorporation into CRT was found in the presence of Ca2+. The arginylated CRT was observed in the cytosol, in contrast to the non-arginylated CRT that is in the endoplasmic reticulum. Under stress conditions, arginylated CRT was found associated to stress granules. These results suggest that CRT arginylation occurs in the cytosolic pool of mature CRT (defined by an Asp acid N-terminal) that is probably retrotranslocated from the endoplasmic reticulum.
Decca
María B
MB
Centro de Investigaciones en Química Biológica de Córdoba, CIQUIBIC, (UNC-CONICET), Departamento de Química Biológica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, 5000-Córdoba, Argentina.
Carpio
Marcos A
MA
Bosc
Christophe
C
Galiano
Mauricio R
MR
Job
Didier
D
Andrieux
Annie
A
Hallak
Marta E
ME
D000818:Animals; D001120:Arginine / Q000737:chemistry*; D001921:Brain / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D037282:Calreticulin / Q000737:chemistry* / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D047468:Immunoprecipitation; D051379:Mice; D041681:NIH 3T3 Cells; D010455:Peptides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D051381:Rats; D013997:Time Factors
D037282:Calreticulin; D010455:Peptides; D001120:Arginine; D002118:Calcium