MODULATION OF AUUUA RESPONSE ELEMENT-BINDING BY HETEROGENEOUS NUCLEARRIBONUCLEOPROTEIN A1 IN HUMAN T-LYMPHOCYTES - THE ROLES OF CYTOPLASMIC LOCATION, TRANSCRIPTION, AND PHOSPHORYLATION

The heterogeneous nuclear ribonucleoprotein Al (hnRNP A1) shuttles bet ween the cytoplasm and nucleus and plays important roles in RNA metabo lism, Whereas nuclear hnRNP Al has been shown to bind intronic sequenc es and modulate splicing, cytoplasmic hnRNP Al is associated with poly (A)(+) RNA, indicating different RNA ligand specificity. Previous stud ies indicated that cytoplasmic hnRNP Al is capable of high-affinity bi nding of reiterated AUUUA sequences (ARE) that have been shown to modu late mRNA turnover and translation. Through a combination of two-dimen sional gel and proteolysis studies, we establish hnRNP Al (or structur ally related proteins that are post-translationally regulated in an id entical manner) as the dominant cytoplasmic protein in human T lymphoc ytes capable of interacting with the ARE contained within the context of full-length granulocyte-macrophage colony-stimulating factor mRNA. We additionally demonstrate that cytoplasmic hnRNP Al preferentially b inds ARE relative to pre-mRNAs in both cross-linking and mobility shif t experiments. RNA polymerase II inhibition increased the binding of A RE (AUBP activity) and poly(U)-Sepharose by cytoplasmic hnRNP Al, whil e nuclear hnRNP Al binding was unaffected. Nuclear and cytoplasmic hnR NP Al could be distinguished by the differential sensitivity of their RNA binding to diamide and N-ethyhmaleimide. The increase in AUBP acti vity of cytoplasmic hnRNP Al following RNA polymerase II inhibition co rrelated with serine-threonine dephosphorylation, as determined by inh ibitor and metabolic labeling studies, Thus, cytoplasmic and nuclear h nRNP Al exhibit different RNA binding profiles, perhaps transduced thr ough serine-threonine phosphorylation. These findings are relevant to the specific ability of hnRNP Al to serve distinct roles in post-trans criptional regulation of gene expression in both the nucleus and cytop lasm.