Characterization of the Human α1β1 Soluble Guanylyl Cyclase Promoter

Abstract

Soluble guanylyl cyclase (sGC) is the principal receptor for NO and plays a ubiquitous role in regulating cellular function. This is exemplified in the cardiovascular system where sGC governs smooth muscle tone and growth, vascular permeability, leukocyte flux, and platelet aggregation. As a consequence, aberrant NO-sGC signaling has been linked to diseases including hypertension, atherosclerosis, and stroke. Despite these key (patho)physiological roles, little is known about the expressional regulation of sGC. To address this deficit, we have characterized the promoter activity of human ␣ 1 and ␤ 1 sGC genes in a cell type relevant to cardiovascular (patho)physiology, primary human aortic smooth muscle cells. Luciferase reporter constructs revealed that the 0.3-and 0.5-kb regions upstream of the transcription start sites were optimal for ␣ 1 and ␤ 1 sGC promoter activity, respectively. Deletion of consensus sites for c-Myb, GAGA, NFAT, NF-B(p50), and CCAAT-binding factor(s) (CCAAT-BF) revealed that these are the principal transcription factors regulating basal sGC expression. In addition, under pro-inflammatory conditions, the effects of the strongest ␣ 1 and ␤ 1 sGC repressors were enhanced, and enzyme expression and activity were reduced; in particular, NF-B(p50) is pivotal in regulating enzyme expression under such conditions. NO itself also elicited a cGMP-independent negative feedback effect on sGC promoter activity that is mediated, in part, via CCAAT-BF activity. In sum, these data provide a systematic characterization of the promoter activity of human sGC ␣ 1 and ␤ 1 subunits and identify key transcription factors that govern subunit expression under basal and pro-inflammatory (i.e. atherogenic) conditions and in the presence of ligand NO.

RNA splicing in regulation of nitric oxide receptor soluble guanylyl cyclase

“…Consistent with the reports indicating that sGC expression is regulated during embryonic development [71–75], TF binding sites involved in morphogenesis and differentiation, such as c-Myb, AP1, GATA1, Nkx-2.5 and Lyf-1, have been identified in the α1 and β1 sGC promoters [66, 67, 69]. Additionally, human and rodent α1 sGC promoters possess a variety of binding sites for stress response TFs, including NFAT, NF-kB, AP1 and SP1.…”

“…The core promoter regions of α1 and β1 sGC genes are TATA-less and contain a number of similar transcription factor (TF) binding sites. Importantly, each has a CCAAT-binding site, which is associated with the CBF (NF-Y) transcriptional factor, and shown to be essential for house-keeping promoter activity in all species investigated to date [66, 67, 69, 70]. The presence of these common elements suggests a potential for coordinated regulation of α1 and β1 sGC genes transcription.…”

Estrogen and the cardiovascular system

“…Interestingly, there is work demonstrating that NFκB inhibits expression of sGC, and increased NFκB activity has been found in the aging cardiovascular system. (Marro et al, 2008; Stice et al, 2011; Csiszar, Wang, Lakatta, & Ungvari, 2008) Increased NFκB activation can lead to increased TNF. Provocatively, Etanercept (the truncated TNF receptor used to reduce TNF activity in rheumatoid arthritis) treatment resulted in significant improvement in vascular relaxation in aged rats, and this correlated with a reduction in the elevated TNF serum levels post ovx, suggesting that TNF may be an important contributor to post-menopausal vascular disease.…”

Genetic modification of hypertension by sGCα1

“…Extensive characterization of the promoter region revealed the presence of transcription factor binding sites, repressor elements, and enhancer elements that govern expression of (human) sGCα 1 and sGCβ 1 (Marro et al, 2008). Some of these elements appear to be mediating previously identified inhibitory effects of NO on sGC expression and therefore have implications for blood pressure regulation.…”

“…Downstream of NO, decreased expression of sGC may result in impaired NO-cGMP signaling. For example, sGCα 1 and sGCβ 1 expression is repressed under pro-inflammatory conditions, an observation that has important implications in cardiovascular diseases characterized by chronic vascular inflammation such as atherosclerosis where sGC may exert anti-atherogenic effects (Marro et al, 2008). Alternatively, sGC can be converted to an NO-insensitive state as a consequence of increased oxidative stress (Stasch et al, 2006) (e.g., resulting from NOS3 uncoupling), a known risk factor for cardiovascular disease.…”