cAMP-mediated stimulation of tyrosine hydroxylase mRNA translation is mediated by polypyrimidine-rich sequences within its 3'-untranslated region and poly(C)-binding protein 2.
Tyrosine hydroxylase (TH) plays a critical role in maintaining the appropriate concentrations of catecholamine neurotransmitters in brain and periphery, particularly during long-term stress, long-term drug treatment, or neurodegenerative diseases. Its expression is controlled by both transcriptional and post-transcriptional mechanisms. In a previous report, we showed that treatment of rat midbrain slice explant cultures or mouse MN9D cells with cAMP analog or forskolin leads to induction of TH protein without concomitant induction of TH mRNA. We further showed that cAMP activates mechanisms that regulate TH mRNA translation via cis-acting sequences within its 3'-untranslated region (UTR). In the present report, we extend these studies to show that MN9D cytoplasmic proteins bind to the same TH mRNA 3'-UTR domain that is required for the cAMP response. RNase T1 mapping demonstrates binding of proteins to a 27-nucleotide polypyrimidine-rich sequence within this domain. A specific mutation within the polypyrimidine-rich sequence inhibits protein binding and cAMP-mediated translational activation. UV-cross-linking studies identify a approximately 44-kDa protein as a major TH mRNA 3'-UTR binding factor, and cAMP induces the 40- to 42-kDa poly(C)-binding protein-2 (PCBP2) in MN9D cells. We show that PCBP2 binds to the TH mRNA 3'-UTR domain that participates in the cAMP response. Overexpression of PCBP2 induces TH protein without concomitant induction of TH mRNA. These results support a model in which cAMP induces PCBP2, leading to increased interaction with its cognate polypyrimidine binding site in the TH mRNA 3'-UTR. This increased interaction presumably plays a role in the activation of TH mRNA translation by cAMP in dopaminergic neurons.
Xu
Lu
L
Department of Pharmacology and Physiology, University of Rochester Medical Center, Rochester, NY 14642, USA.
Sterling
Carol R
CR
Tank
A William
AW
D020413:3' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D000242:Cyclic AMP / Q000502:physiology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D014176:Protein Biosynthesis / Q000502:physiology*; D011743:Pyrimidines / Q000737:chemistry*; D012333:RNA, Messenger / Q000235:genetics*; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D014446:Tyrosine 3-Monooxygenase / Q000737:chemistry / Q000235:genetics*
D020413:3' Untranslated Regions; D011743:Pyrimidines; D012333:RNA, Messenger; D000242:Cyclic AMP; D014446:Tyrosine 3-Monooxygenase; C030986:pyrimidine