Human phosducin-like protein (hPhLP) messenger RNA stability is regulated by cis-acting instability elements present in the 3'-untranslated region.
Phosducin (Pd) and phosducin-like protein (PhLP) have been shown to regulate G-protein signaling by binding G beta gamma subunits. To better define the function and regulation of PhLP, and to begin to investigate its potential role in human pathophysiological states, we have cloned the human PhLP (hPhLP) cDNA. The hPhLP shows 92% identity with the rat PhLP (rPhLP). However, unlike the rPhLP, no evidence of hPhLP isoforms were detected in the human tissues investigated. Additionally, unlike the rPhLP, alternative polyadenylation sites were detected in hPhLP cDNA clones which corresponded with two distinct mRNA transcripts, 1.2 kb and 3.1 kb, respectively. Interestingly, the predominantly expressed long transcript contains multiple AU-rich elements (AREs) in its 3'-untranslated region (3'-UTR) which have been shown to correlate with rapid mRNA turnover and translational control. This study shows that the hPhLP AREs are functional both in vitro and in vivo, with the long transcript exhibiting a much shorter mRNA half-life. We also demonstrate that subcloning of either the full-length 3'-UTR or the ARE-rich region of the long transcript immediately following the stop codon of luciferase reporter gene confers instability to the luciferase mRNA and results in a ninefold reduction of luciferase activity in the cell types investigated. Taken together, these findings suggest that the AREs present in the long hPhLP mRNA may play a critical role in the regulation of hPhLP gene expression.
Lazarov
M E
ME
Department of Chemistry and Biochemistry, Brigham Young University, Provo, UT 84602, USA.
Martin
M M
MM
Willardson
B M
BM
Elton
T S
TS
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry; D018076:DNA, Complementary / Q000302:isolation & purification; D003609:Dactinomycin; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D005786:Gene Expression Regulation; D015723:Gene Library; D006207:Half-Life; D020962:Heterotrimeric GTP-Binding Proteins; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry* / Q000302:isolation & purification; D012044:Regression Analysis; D014162:Transfection; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D002457:Cell Extracts; D018076:DNA, Complementary; C118981:G-protein Beta gamma; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D009419:Nerve Tissue Proteins; C098789:PDCL protein, human; D012333:RNA, Messenger; D003609:Dactinomycin; D008156:Luciferases; D019204:GTP-Binding Proteins; D020962:Heterotrimeric GTP-Binding Proteins