Retinoblastoma Protein Modulates the Inverse Relationship between Cellular Proliferation and Elastogenesis

Cardiovascular Research, The Hospital for Sick Children, University of Toronto Toronto M5G 1X8, Canada.
Journal of Biological Chemistry (Impact Factor: 4.57). 08/2011; 286(42):36580-91. DOI: 10.1074/jbc.M111.269944
Source: PubMed

ABSTRACT The mechanism that leads to the inverse relationship between heightened cellular proliferation and the cessation of elastic fibers production, observed during formation of the arterial occlusions and dermal scars, is not fully understood. Because the retinoblastoma protein (Rb), responsible for cell cycle initiation, has also been implicated in insulin-like growth factor-I-mediated signaling stimulating elastin gene activation, we explored whether differential phosphorylation of Rb by various cyclin·cyclin-dependent kinase complexes would be responsible for promoting either elastogenic or pro-proliferative signals. We first tested cultures of dermal fibroblasts derived from Costello syndrome patients, in which heightened proliferation driven by mutated oncogenic H-Ras coincides with inhibition of elastogenesis. We found that Costello syndrome fibroblasts display elevated level of Rb phosphorylation on serine 780 (Ser(P)-780-Rb) and that pharmacological inhibition of Ras with radicicol, Mek/Erk with PD98059, or cyclin-dependent kinase 4 with PD0332991 not only leads to down-regulation of Ser(P)-780-Rb levels but also enhances Rb phosphorylation on threonine-821 (Thr(P)-821-Rb), which coincides with the recovery of elastin production. Then we demonstrated that treatment of normal skin fibroblasts with the pro-proliferative PDGF BB also up-regulates Ser(P)-780-Rb levels, but treatment with the pro-elastogenic insulin-like growth factor-I activates cyclinE-cdk2 complex to phosphorylate Rb on Thr-821. Importantly, we have established that elevation of Thr(P)-821-Rb promotes Rb binding to the Sp1 transcription factor and that successive binding of the Rb-Sp1 complex to the retinoblastoma control element within the elastin gene promoter stimulates tropoelastin transcription. In summary, we provide novel insight into the role of Rb in mediating the inverse relationship between elastogenesis and cellular proliferation.

Download full-text


Available from: Aleksander Hinek, Sep 28, 2015
32 Reads
  • Source
    • "The deposition of durable elastin is also modulated by the 67-kDa elastin binding protein (EBP) [16] identified as a molecular chaperone that binds and escorts intracellular tropoelastin through the secretory pathway, thus assuring its orderly assembly into extracellular elastic fibers [17] [18]. While previous in vitro studies from our laboratory have shown that production of new elastic fibers could be potently stimulated by several natural and pharmacological factors [19] [20] [21] [22] [23] [24], our current results demonstrated for the first time that cultures of human cardiac fibroblasts treated with tanshinone IIA contained significantly fewer collagen fibers than their untreated counterparts, and this phenomenon coincided with the remarkable up-regulation in elastic fiber production. "
    International journal of cardiology 08/2015; 202. DOI:10.1016/j.ijcard.2015.08.191 · 4.04 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Diabetes mellitus accelerates atherosclerotic progression, peripheral angiopathy development, and arterial hypertension, all of which are associated with elastic fiber disease. However, the potential mechanistic links between insulin deficiency and impaired elastogenesis in diabetes have not been explored. Results of the present study reveal that insulin administered in therapeutically relevant concentrations (0.5 to 10 nmol/L) selectively stimulates formation of new elastic fibers in cultures of human aortic smooth muscle cells. These concentrations of insulin neither up-regulate collagen type I and fibronectin deposition nor stimulate cellular proliferation. Further, the elastogenic effect of insulin occurs after insulin receptor activation, which triggers the PI3K downstream signaling pathway and activates elastin gene transcription. In addition, the promoter region of the human elastin gene contains the CAAATAA sequence, consistent with the FoxO-recognized element, and the genomic effects of insulin occur after removal of the FoxO1 transcriptional inhibitor from the FoxO-recognized element in the elastin gene promoter. In addition, insulin signaling facilitates the association of tropoelastin with its specific 67-kDa elastin-binding protein/spliced form of β-galactosidase chaperone, enhancing secretion. These results are crucial to understanding of the molecular and cellular mechanisms of diabetes-associated vascular disease, and, in particular, endorse use of insulin therapy for treatment of atherosclerotic lesions in patients with type 1 diabetes, in which induction of new elastic fibers would mechanically stabilize the developing plaques and prevent arterial occlusions.
    American Journal Of Pathology 02/2012; 180(2):715-26. DOI:10.1016/j.ajpath.2011.10.022 · 4.59 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Williams-Beuren Syndrome (WBS) and Supravalvular Aortic Stenosis (SVAS) are genetic syndromes marked by the propensity to develop severe vascular stenoses. Vascular lesions in both syndromes are caused by haploinsufficiency of the elastin gene. We used these distinct genetic syndromes as models to evaluate the feasibility of using engineered zinc finger protein transcription factors (ZFPs) to achieve compensatory expression of haploinsufficient genes by inducing augmented expression from the remaining wild-type allele. Targeting the elastin gene, we show that transcriptional activation by engineered ZFPs can increase elastin expression in wild-type cells, induce compensatory expression from the wild-type allele in both WBS and SVAS cells, induce expression of the major elastin splice variants, and recapitulate their natural stoichiometry. Further, we establish that transcriptional activation of the mutant allele in SVAS does not overcome nonsense-mediated decay and thus ZFP-mediated transcriptional activation is not likely to induce production of a mutant protein, a crucial consideration. Finally, we show in bioengineered blood vessels that ZFP-mediated induction of elastin expression is capable of stimulating functional elastogenesis. These findings have significant implications for WBS and SVAS, and establish that haploinsufficiency can be overcome by targeted transcriptional activation without inducing protein expression from the mutant allele.
    Human gene therapy 08/2012; 23(11). DOI:10.1089/hum.2011.201 · 3.76 Impact Factor
Show more