Down-Regulation of Soluble Guanylyl Cyclase Expression by Cyclic AMP Is Mediated by mRNA-Stabilizing Protein HuR

Abstract

We analyzed whether the cyclic AMP induced down-regulation of the nitric oxide (NO) receptor soluble guanylyl cyclase (sGC) is mediated by the mRNA-protecting protein HuR. Exposure (up to 24 h) of isolated rat aortic segments to the activator of adenylyl cyclase, forskolin (10 M), and to both activators of cAMP-stimulated protein kinase (PKA), 5,6-dichloro-1-␤-D-ribofuranosylbenzimidazole-3Ј,5Ј-cyclic monophosphorothioate, Sp-isomer (Sp-5,6-DCl-cBIMPS; 400 nM), and N 6 -phenylcAMP (10 M), strongly reduced sGC␣ 1 ␤ 1 and HuR protein and mRNA expression in a time-dependent and actinomycin D (10 M)-sensitive fashion. In vitro degradation of sGC␣ 1 and ␤ 1 poly(A) ϩ mRNA by native rat aortic protein was markedly increased by pretreatment of intact aortas with forskolin. Native protein extract from rat aorta shifted the electrophoretic mobility of biotin-labeled riboprobes from the 3Ј-untranslated region of sGC␣ 1 and ␤ 1 mRNA, and these bands was supershifted by a monoclonal antibody directed against the mRNA-stabilizing protein HuR. Forskolin decreased the HuR-sGC␣ 1 and ␤ 1 mRNA interaction and HuR protein expression in rat aorta, and this was prevented by the PKA inhibitory cAMP analog 3Ј,5Ј-cyclic monophosphorothioate, Rp-isomer (Rp-cAMPS). In cultured smooth muscle cells from rat aorta, forskolin induced a rapid increase in Fos/p-Fos protein levels and activator protein 1 (AP-1) binding activity. Inhibition of this transcription factor by an AP-1 decoy prevented the forskolin-induced down-regulation of HuR. We conclude that forskolin/cAMP decrease the expression of heterodimeric sGC in rat aortic smooth muscle cells via activation of Fos/AP-1, which decreases the expression of HuR and thus destabilizes the sGC␣ 1 and ␤ 1 mRNA.

NMDA induces post-transcriptional regulation of α2-guanylyl-cyclase-subunit expression in cerebellar granule cells

“…Furthermore, we identified several AREs in this region that are targeted by proteins of the elav family and AUF1 proteins, involved in stabilising/destabilising mRNA. This structural characteristic has previously not been studied in mRNAs encoding the ␣ 2 subunit, although it has already been shown that mRNAs encoding the ␣ 1 and ␤ 1 subunits have a long 3Ј UTR to which stabilising proteins bind (Klöss et al, 2003;Klöss et al, 2004). Furthermore, our results demonstrate that expression of the ␣ 2 subunit of NO-sensitive guanylyl cyclase is regulated in granule cells by a mechanism that affects mRNA stability.…”

“…Whereas different mechanisms have been proposed to regulate the expression of the ␣ 1 and ␤ 1 subunits of guanylyl cyclase in different cell types (Ferrero et al, 2000;Ferrero and Torres, 2002;Klöss et al, 2003;Klöss et al, 2004), no studies on the regulation of ␣ 2 expression have been carried out to date. This is surprising, given that this subunit seems to be responsible for the synaptic localisation of guanylyl cyclase (Russwurm et al, 2001), which it is highly expressed in different areas of the central nervous system (Gibb and Garthwaite, 2001;Mergia et al, 2003), and that its expression profile coincides with neuronal differentiation (Jurado et al, 2003).…”

“…To date, such sequences have been found in many different mRNAs, including several components of the NO-cGMP pathway, the activities of which are known to be regulated by altering mRNA stability, e.g. ␣ 1 and ␤ 1 subunits of guanylyl cyclase in rat aorta (Klöss et al, 2003;Klöss et al, 2004), NO synthase II in human mesangial cells (Pérez-Sala et al, 2001), and in human intestinal epithelium (DLD1) cells (Rodríguez-Pascual Activation of N-methyl-D-aspartate (NMDA) glutamate receptors commonly affects gene expression in different neurons. We reported previously that chronic treatment of rat cerebellar granule cells with NMDA (24 hours) upregulates the expression of mRNA encoding the ␣ ␣ 2 subunit of the nitric-oxide-sensitive guanylyl cyclase.…”

Molecular cloning and characterization of α1-soluble guanylyl cyclase gene promoter in rat pituitary cells

“…In human mesangial cells, collagen type I can also reduce the steady-state b1-sGC mRNA levels and such reduction occurs at transcriptional level (De Frutos et al 2005). On the other hand, the AU-rich elements identified in the 3 0 -untranslated region of a1-and b1-sGC mRNAs provide sites for posttranscriptional regulation by cAMP (Kloss et al 2004). Differential expression of mRNAs for a and b-sGC also influences the formation of functional heterodimers.…”

Loss of Vascular Regulation by Soluble Guanylate Cyclase Is Emerging as a Key Target of the Hypertensive Disease Process

“…Previous work from this group showed how the HuR protein functions to stabilize the mRNA for sGC 2 and that the expression of HuR is controlled by the availability of cGMP, whereas cGMP appears to inhibit the expression of this mRNA-stabilizing protein. 3 Because hypertension is thought to generally decrease sGC expression and the stimuli for sGC activation, 4,5 the decrease in HuR expression is likely to be through a mechanism that is independent of the previously identified interactions of cGMP with this system. A model describing the potential influence of hypertension on relationships between these systems in controlling the expression and activity of sGC is shown in the Figure. The decrease in HuR has many implications for controlling processes potentially involved in the progression of hypertension that are discussed in the article by Klöss et al 1 However, the influence of HuR on the expression of sGC may be a primary factor in the progression of hypertension.…”