Mechanisms in cardiac fibroblast growth: an obligate role for Skp2 and FOXO3a in ERK1/2 MAPK-dependent regulation of p27^kip1

Abstract

Cardiac fibroblast hyperplasia associated with enhanced matrix deposition is a major determinant of tissue remodeling in several disease states of the heart. However, mechanisms controlling cell cycle progression in cardiac fibroblasts remain unexplored. Identification of cell cycle regulatory elements in these cells is important to develop strategies to check adverse cardiac remodeling under pathological conditions. This study sought to probe the mechanisms underlying ERK1/2-mediated p27(Kip1) regulation in mitogenically stimulated cardiac fibroblasts. Addition of 10% fetal calf serum to quiescent cultures of adult rat cardiac fibroblasts promoted ERK1/2 activation, as evidenced by its phosphorylation status. Reduction in [(3)H]thymidine incorporation into DNA increased population doubling time, flow cytometry, and Western blot analysis showing reduced levels of cyclins D and A, p27(Kip1) induction, and retinoblastoma protein (Rb) hypophosphorylation in ERK1/2-inhibited cells indicated ERK1/2 dependence of G1-S transition in cardiac fibroblasts. Lack of p27(Kip1) protein in serum-stimulated, ERK1/2-active cells was associated with increased levels of Skp2, an E3 ubiquitin ligase for p27(Kip1), whose knockdown by RNA interference induced p27(Kip1) expression. Further, forced expression of Skp2 in ERK1/2-inhibited cells downregulated p27(Kip1). Transcriptional upregulation of p27(Kip1) mRNA in ERK1/2-inhibited cells, demonstrated by real-time PCR, correlated with forkhead box O 3a (FOXO3a) transcription factor activation, shown by gel shift assay. FOXO3a knockdown attenuated p27(Kip1) mRNA and protein expression in ERK1/2-inhibited cells. We provide evidence for the first time that, in cardiac fibroblasts, activated ERK1/2 regulates p27(Kip1) expression transcriptionally and posttranslationally via FOXO3a- and Skp2-dependent mechanisms. Additionally, this study uncovers interesting interactions between critical cell cycle regulatory elements that are only beginning to be understood.

Egg antigen p40 of Schistosoma japonicum promotes senescence in activated hepatic stellate cells via SKP2/P27 signaling pathway

“…G1-S phase transition has recently been recognized to be mediated by the induction of P27 in ERK-inhibited fibroblasts 23 . P27 signaling pathway is served as an important aging-associated signaling pathway 24 .…”

“…Cellular senescence refers to an irreversible cell cycle arrest state that could limit cell proliferation and growth 23 . It is generally known that cell senescence is regarded as an effective method of tumor suppression 29 , however, recent researches demonstrate that it also plays an important suppressive role in other non-neoplastic diseases, such as liver fibrosis.…”

Inhibition of farnesyl pyrophosphate synthase improves pressure overload induced chronic cardiac remodeling

“…Fibronectin, collagen I accumulation, overall interstitial fibrosis and the myofibroblast population were lower in the H-Ras knockout mice than in the wild-type mice27. Inhibiting ERK1/2 activity represses cell proliferation and collagen gene expression in activated cardiac fibroblasts28. Li et al .…”

Post-translational modifications of FOXO family proteins

“…In cardiac fibroblasts, activation of ERK1/2 can directly phosphorylate FOXO3a and regulate its activity, and ERK1/2 can also phosphorylate Skp2 in the same manner. Notable, a report previously demonstrated that Skp2 can inhibit the activity of FOXO3a and promote its degradation (48,49). SGK is another link between MAPK and FOXO proteins.…”