The RGG domain in hnRNP A2 affects subcellular localization

The heterogeneous nuclear ribonucleoproteins (hnRNP) associate with pre-mRN A in the nucleus and play an important role in RNA processing and splice si te selection. In addition, hnRNP A proteins function in the export of mRNA to the cytoplasm, Although the hnRNP A proteins are predominantly nuclear, hnRNP A1 shuttles rapidly between the nucleus and the cytoplasm, HnRNP A2, whose cytoplasmic overexpression has been identified as an early biomarker of lung cancer, has been less well studied. Cytosolic hnRNP A2 overexpressi on has also been noted in brain tumors, in which it has been correlated wit h translational repression of Glucose Transporter-1 expression, We now exam ine the role of arginine methylation on the nucleocytoplasmic localization of hnRNP A2 in the HEK-293 and NIH-3T3 mammalian cell lines. Treatment of e ither cell line with the methyltransferase inhibitor adenosine dialdehyde d ramatically shifts hnRNP A2 localization from the nuclear to the cytoplasmi c compartment, as shown both by immunoblotting and by immunocytochemistry, In vitro radiolabeling with [H-3]AdoMet of GST-tagged hnRNP A2 RGG mutants, using recombinant protein arginine methyltransferase (PRMT1), shows (i) th at hnRNP A2 is a substrate for PRMT1 and (ii) that methylated residues are found only in the RGG domain. Deletion of the RGG domain (R191-G253) of hnR NP A2 results in a cytoplasmic localization phenotype, detected both by imm unoblotting and by immunocytochemistry, These studies indicate that the RGG domain of hnRNP A2 contains sequences critical for cellular localization o f the protein. The data suggest that hnRNP A2 may contain a novel nuclear l ocalization sequence, regulated by arginine methylation, that lies in the R 191-G253 region and may function independently of the M9 transportin-1-bind ing region in hnRNP A2. (C) 2000 Academic Press.