Cytoplasmic Protein mRNA Interaction Mediates cGMP-modulated Translational Control of the Asialoglycoprotein Receptor

Abstract

Expression of the asialoglycoprotein receptor by the human hepatocellular carcinoma cell line HuH-7 in response to intracellular cGMP concentrations was previously shown to be regulated at the translational level. In a cell-free system, initiation of asialoglycoprotein receptor mRNA translation was dependent on the presence of the 7-methylguanylate cap site and was independent of 8-bromo-cGMP levels in which the cells were grown prior to RNA isolation. Stable transfection of COS-7 cells with deletion constructs of the asialoglycoprotein receptor H2b subunit localized the cGMP-responsive cisacting element to the mRNA 5 -untranslated region (UTR). Addition of biotin (an activator of guanylate cyclase) induced the expression of ␤-galactosidase present as a chimeric plasmid containing the H2b 187-nucleotide 5 -UTR. An RNA gel retardation assay identified a 37-nucleotide cognate sequence within this 187-nucleotide region. Titration of the 5 -UTR with a cytosolic fraction isolated from HuH-7 grown in the presence or absence of 8-bromo-cGMP or biotin provided direct evidence for an RNA-binding protein responsive to intracellular levels of cGMP. Based on these findings, it seems reasonable to propose that reduction of intracellular levels of cGMP by biotin deprivation results in a negative trans-acting factor associating with the 5 -UTR of asialoglycoprotein receptor mRNAs, thereby inhibiting translation.

Reactive Oxygen and Nitrogen Intermediates Increase Transforming Growth Factor– β 1 Release from Human Epithelial Alveolar Cells through Two Different Mechanisms

“…that cGMP-dependent kinase might affect the phosphorylation status of a transacting factor that binds to specific sequences within the 5Ј untranslated region of TGF-␤1 mRNA. This mechanism has been suggested to regulate the expression of various proteins at a translational level (51).…”

Biotin-dependent regulation of gene expression in human cells

“…These results led to the finding that transcriptional regulation of genes from the biotin cycle is mediated by a signal transduction pathway involving a soluble guanylate cyclase (sGC) and a cGMP-dependent protein kinase (PKG) [13,14]. This pathway was previously shown to be responsible for the biotin-dependent posttranscriptional regulation of ASGPR expression by Stockert et al [7,8,12]. To continue the characterization of biotindependent gene expression, we used cells from patients with neonatal MCD.…”

Selective accumulation of biotin in arterial chemoreceptors: requirement for carotid body exocytotic dopamine secretion

“…Moreover, biotin is known to activate the guanyl cyclase/cyclic GMP/protein kinase G pathway (Stockert et al . ; Stocker & Ren, ), which could indirectly lead to VMAT2 modulation. Indeed, phosphorylation and nitration of VMAT2 residues have been reported (Watabe & Nakaki, ; Ramamoorthy et al .…”