Chaperonin containing TCP-1 (CCT) is a large multisubunit complex that mediates protein folding in eukaryotic cells. CCT participates in the folding of newly synthesized polypeptides, including actin, tubulin, and several cell cycle regulators; therefore, CCT plays an important role in cytoskeletal organization and cell division. Here we identify the chaperonin CCT as a novel physiological substrate for p90 ribosomal S6 kinase (RSK) and p70 ribosomal S6 kinase (S6K). RSK phosphorylates the beta subunit of CCT in response to tumor promoters or growth factors that activate the Ras-mitogen-activated protein kinase (MAPK) pathway. CCTbeta Ser-260 was identified as the RSK site by mass spectrometry and confirmed by site-directed mutagenesis. RSK-dependent Ser-260 phosphorylation was sensitive to the MEK inhibitor UO126 and the RSK inhibitor BID-1870. Insulin weakly activates RSK but strongly activates the phosphoinositide 3-kinase (PI3K)-mammalian target of rapamycin (mTOR) pathway and utilizes S6K to regulate CCTbeta phosphorylation. Thus, the Ras-MAPK and PI3K-mTOR pathways converge on CCTbeta Ser-260 phosphorylation in response to multiple agonists in various mammalian cells. We also show that RNA interference-mediated knockdown of endogenous CCTbeta causes impaired cell proliferation that can be rescued with ectopically expressed murine CCTbeta wild-type or phosphomimetic mutant S260D, but not the phosphorylation-deficient mutant S260A. Although the molecular mechanism of CCTbeta regulation remains unclear, our findings demonstrate a link between oncogene and growth factor signaling and chaperonin CCT-mediated cellular activities.
"Therefore , CCT6a seems to be differentially regulated by MMP-3, possibly by a modification at the transcriptional or/and post-translational level. CCT6a is a member of the eukaryotic cytosolic chaperonin containing TCP-1 (CCT) complex that mediates folding of various proteins such as actin and tubulin (Fig. 3; Abe et al., 2009). To investigate whether CCT6a is the only subunit that has undergone MMP-3- dependent modifications during development of the cerebellum, Western blotting was also performed for several other CCT subunits, more specifically CCT3, 4, 5, 6b, and 7. Yet, no genotypic differences in expression level or in molecular weight could be observed for any of these CCT subunits [Figure 4 "
[Show abstract][Hide abstract] ABSTRACT: Matrix metalloproteinase-3 (MMP-3) deficiency in mice was previously reported to result in a transiently retarded granule cell migration at postnatal day 8 (P8) and a sustained disturbed arborization of Purkinje cell dendrites from P8 on, concomitant with a delayed synapse formation between granule cells and Purkinje cells and resulting in mild deficits in motor performance in adult animals. However, the molecular mechanisms by which MMP-3 contributes to proper development of the cerebellar cortex during the first postnatal weeks remains unknown. In this study, we used a functional proteomics approach to investigate alterations in protein expression in postnatal cerebella of wild-type versus MMP-3 deficient mice, and to further elucidate MMP-3-dependent pathways and downstream targets in vivo. At P8, two-dimensional difference gel electrophoresis (2-D DIGE) and mass spectrometry identified 20 unique proteins with a different expression between the two genotypes. Subsequent ‘Ingenuity Pathway Analysis’ (IPA) analysis and Western blotting indicate that the chaperonin containing T-complex polypeptide 1, subunit 6A (CCT6a) and the MAP kinase signaling pathway play a key role in the MMP-3-dependent regulation of neurite outgrowth and neuronal migration in the developing brain. This article is protected by copyright. All rights reserved
"CCT is composed of eight different subunits (1, 2, 3, 4, 5, 6, 7, and 8) and their functions are poorly understood . Previous studies showed that CCT2 expression was important for normal cell proliferation , . Moreover, CCT2 is overexpressed in certain malignant tumors and its overexpression is closely correlated with poor prognosis . "
[Show abstract][Hide abstract] ABSTRACT: Dental pulp stem cells (DPSCs), precursor cells of odontoblasts, are ideal seed cells for tooth tissue engineering and regeneration. Our previous study has demonstrated that stem cells exist in dental pulp with deep caries and are called carious dental pulp stem cells (CDPSCs). The results indicated that CDPSCs had a higher proliferative and stronger osteogenic differentiation potential than DPSCs. However, the molecular mechanisms responsible for the biological differences between DPSCs and CDPSCs are poorly understood. The aim of this study was to define the molecular features of DPSCs and CDPSCs by comparing the proteomic profiles using two-dimensional fluorescence difference gel electrophoresis (2-D DIGE) in combination with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Our results revealed that there were 18 protein spots differentially expressed between DPSCs and CDPSCs in a narrow pH range of 4 to 7. These differently expressed proteins are mostly involved in the regulation of cell proliferation, differentiation, cell cytoskeleton and motility. In addition, our results suggested that CDPSCs had a higher expression of antioxidative proteins that might protect CDPSCs from oxidative stress. This study explores some potential proteins responsible for the biological differences between DPSCs and CDPSCs and expands our understanding on the molecular mechanisms of mineralization of DPSCs in the formation of the dentin-pulp complex.
PLoS ONE 05/2014; 9(5):e97026. DOI:10.1371/journal.pone.0097026 · 3.23 Impact Factor
"Forced expression of HER2 or HER3 alone was sufficient to restore p-AKT expression levels in the cells with miR-199a and miR-125b overexpression. P70S6K1 is a downstream target of AKT and mTOR . P70S6K1 may regulate HIF-1α expression and tumor angiogenesis . "
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs (miRNAs) have been shown to be involved in different aspects of cancer biology including tumor angiogenesis. In this study, we identified that two miRNAs, miR-199a and miR-125b were downregulated in ovarian cancer tissues and cell lines. Overexpression of miR-199a and miR-125b inhibited tumor-induced angiogenesis associated with the decrease of HIF-1α and VEGF expression in ovarian cancer cells. Moreover, the levels of miR-199a and miR-125b were negatively correlated with VEGF mRNA levels in ovarian tissues. We further showed that direct targets of miR-199a and miR-125b HER2 and HER3 were functionally relevant. Forced expression of HER2 and HER3 rescued miR-199a- and miR-125b-inhibiting angiogenesis responses and Akt/p70S6K1/HIF-1α pathway. This study provides a rationale for new therapeutic approach to suppress tumor angiogenesis using miR-199a, miR-125b, or their mimics for ovarian cancer treatment in the future.
PLoS ONE 02/2013; 8(2):e56647. DOI:10.1371/journal.pone.0056647 · 3.23 Impact Factor
Gabriel Almroth, Johanna Lönn, Fredrik Uhlin, Lars Brudin, Bengt Andersson, Mirjana Hahn-Zoric
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