Regulation of Cellular Senescence and p16 Expression by Id1 and E47 Proteins in Human Diploid Fibroblast

Abstract

Id1, a member of Id family of helix-loop-helix transcriptional regulatory proteins, is implicated in cellular senescence by repressing p16INK4a expression, but the mechanisms and cellular effects in human diploid fibroblasts remain unknown. Here we analyzed the patterns of p16INK4a and Id1 expression during the lifespan of 2BS cells and presented the inverse correlation between these two proteins. Immunoprecipitation assays demonstrated the presence of endogenous interaction of Id1 and E47 proteins that was strong in young 2BS cells and weakened during replicative senescence and, thereby, influenced the transcription activation of p16INK4a by E47. Furthermore, we found that E47 protein could bind to the E-box-containing region in p16 INK4a promoter in senescent cells by chromatin immunoprecipitation analyses, suggesting that E47 is indeed ultimately involved in the regulation of p16INK4a transcription in vivo. Silencing Id1 expression in young cells by RNA interference induced an increased p16INK4a level and premature cellular senescence, whereas silencing E47 expression inhibited the expression of p16INK4a and delayed the onset of senescent phenotype. The present study demonstrated not only the capacity of Id1 to regulate p16INK4a gene expression by E47, but also the phenotypic consequence of the regulation on cellular senescence, moreover, raised the possibility of Id1-specific gene silencing for human cancer therapy.

Id1 cooperates with oncogenic Ras to induce metastatic mammary carcinoma by subversion of the cellular senescence response

“…Several studies have implicated Id1 in the repression of p16 INK4A expression during replicative or oncogene induced senescence in vitro (20)(21). However, we have shown that increased p16 INK4A expression is not critical to the senescence response of mammary epithelia and that Id1 does not significantly regulate p16 INK4A expression.…”

Id3 is a novel regulator of p27kip1 mRNA in early G1 phase and is required for cell-cycle progression

“…They play a crucial role in the coordinated regulation of gene expression during cell growth, differentiation and tumorigenesis (Norton, 2000;Benezra et al, 2001). The involvement of Ids was also recently shown in cellular senescence (Alani et al, 2001;Ohtani et al, 2001;Zheng et al, 2004) and in the fate of specialized cells such as lymphocytes, vascular endothelial cells and neurons (Lyden et al, 1999;Benezra et al, 2001;Engel and Murre, 2001).…”

“…In addition, it has also been shown that E2A and Ids proteins participate to the transcriptional regulation of the cyclin-dependent kinase inhibitors (CKIs) p21 ink4a (Id1) (Zheng et al, 2004) in different biological models including senescence. Recent studies have shown that besides p21 and p16, Id1 and Id3 are also involved in the regulation of p27 protein, notably in Xenopus neural crest progenitors (Kee and Bronner-Fraser, 2005) and in Epstein-Barr virus infected cells (Everly et al, 2004); however, the molecular basis of this inhibition has never been elucidated.…”

Smad3 mediates immediate early induction of Id1 by TGF-β

“…They mostly act as negative transcriptional regulators in many biological processes such as cell differentiation, cell senescence, neurogenesis, apoptosis and angiogenesis [6][7][8][9][10][11][12][13]. As all of them lack a DNA-binding domain, they heterodimerize with and inhibit the function of basic HLH (bHLH) transcription factors [6,14], for examples, myogenic regulatory factors, such as MyoD, myogenin, Myf5 and MRF4/Myf6 [15,16], or class A of E proteins, such as E12, E47, E2-2 and ITF-2 [15,[17][18][19]. Inhibitor of differentiation (Id) proteins also interact with non-HLH proteins such as Ets2 [20] and MIDA1 [21] to regulate their activities.…”