CHIP promotes Runx2 degradation and negatively regulates osteoblast differentiation.
Ontology highlight
ABSTRACT:
SUBMITTER: Li X
PROVIDER: S-EPMC2426947 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Cyclophilin A (CypA) is a ubiquitously expressed and highly conserved protein with peptidyl-prolyl cis-trans isomerase activity that is involved in various biological activities by regulating protein folding and trafficking. Although CypA has been reported to positively regulate osteoblast differentiation, the mechanistic details remain largely unknown. In this study, we aimed to elucidate the mechanism of CypA-mediated regulation of osteoblast differentiation. Overexpression of CypA promoted osteoblast differentiation in bone morphogenic protein 4 (BMP4)-treated C2C12 cells, while knockdown of CypA inhibited osteoblast differentiation in BMP4-treated C2C12. CypA and Runx2 were shown to interact based on immunoprecipitation experiments and CypA increased Runx2 transcriptional activity in a dose-dependent manner. Our results indicate that this may be because CypA can increase the DNA binding affinity of Runx2 to Runx2 binding sites such as osteoblast-specific cis-acting element 2. Furthermore, to identify factors upstream of CypA in the regulation of osteoblast differentiation, various kinase inhibitors known to affect osteoblast differentiation were applied during osteogenesis. Akt inhibition resulted in the most significant suppression of osteogenesis in BMP4-induced C2C12 cells overexpressing CypA. Taken together, our results show that CypA positively regulates osteoblast differentiation by increasing the DNA binding affinity of Runx2, and Akt signaling is upstream of CypA.
| S-EPMC9409320 | biostudies-literature
Project description:The maintenance of eukaryotic telomeres requires telomerase, which is minimally composed of a telomerase reverse transcriptase (TERT) and an associated RNA component. Telomerase activity is tightly regulated by expression of human (h) TERT at both the transcriptional and post-translational levels. The Hsp90 and p23 molecular chaperones have been shown to associate with hTERT for the assembly of active telomerase. Here, we show that CHIP (C terminus of Hsc70-interacting protein) physically associates with hTERT in the cytoplasm and regulates the cellular abundance of hTERT through a ubiquitin-mediated degradation. Overexpression of CHIP prevents nuclear translocation of hTERT and promotes hTERT degradation in the cytoplasm, thereby inhibiting telomerase activity. In contrast, knockdown of endogenous CHIP results in the stabilization of cytoplasmic hTERT. However, it does not affect the level of nuclear hTERT and has no effect on telomerase activity and telomere length. We further show that the binding of CHIP and Hsp70 to hTERT inhibits nuclear translocation of hTERT by dissociating p23. However, Hsp90 binding to hTERT was not affected by CHIP overexpression. These results suggest that CHIP can remodel the hTERT-chaperone complexes. Finally, the amount of hTERT associated with CHIP peaks in G(2)/M phases but decreases during S phase, suggesting a cell cycle-dependent regulation of hTERT. Our data suggest that CHIP represents a new pathway for modulating telomerase activity in cancer.
| S-EPMC3009929 | biostudies-literature
Project description:GATA4 is a zinc-finger transcription factor that is a pioneer factor in various tissues and regulates tissue-specific gene regulation. In vivo deletion of Gata4 using Cre-recombinase under the control of the Col1a1 2.3 kb promoter showed significantly reduced values for trabecular bone properties by microCT analysis of femur and tibia of 14-week-old male and female mice, suggesting GATA4 is necessary for maintaining normal adult bone phenotype. Quantitative PCR analysis revealed higher expression of Gata4 in trabecular bone compared with cortical bone, suggesting a role for GATA4 in maintaining normal trabecular bone mass. In vivo and in vitro, reduction of Gata4 correlates with reduced Runx2 gene expression, along with reduced osteoblast mineralization. To determine if Runx2 is a direct target of GATA4, chromatin immunoprecipitation (ChIP) was performed, and it demonstrated that GATA4 is recruited to the two Runx2 promoters and an enhancer region. Furthermore, when Gata4 is knocked down, the chromatin at the Runx2 region is not open, as detected by DNase assays and ChIP with antibodies to the open chromatin marks H3K4me2 (histone 3 lysine 4 dimethylation) and H3K27ac (histone 3 lysine 27 acetylation) and the closed chromatin mark H3K27me2 (histone 3 lysine 27 trimethylation). Together, the data suggest that GATA4 binds near the Runx2 promoter and enhancer and helps maintain open chromatin to regulate Runx2 expression leading to bone mineralization.
| S-EPMC6053063 | biostudies-literature
Project description:VGLL4 has been identified as a YAP inhibitor. However, the exact function of VGLL4 in bone development and bone homeostasis remains unclear. In this study, we demonstrated that VGLL4 breaks TEADs-mediated transcriptional inhibition of RUNX2 to promote osteoblast differentiation and bone development. We found that knockout of VGLL4 in mesenchymal stem cells and preosteoblasts showed osteoporosis and a cleidocranial dysplasia-like phenotype due to osteoblast differentiation disorders. Mechanistically, we showed that the TEAD transcriptional factors severely inhibited osteoblast differentiation in a YAP binding-independent manner. TEADs interacted with RUNX2 to repress RUNX2 transcriptional activity. Furthermore, VGLL4 relieved the transcriptional inhibition of TEADs by directly competing with RUNX2 to bind TEADs through its two TDU domains. Collectively, our studies demonstrate that VGLL4 plays an important role in regulating osteoblast differentiation and bone development, and that TEADs regulate the transcriptional activity of RUNX2, which may shed light on treatment of cleidocranial dysplasia and osteoporosis.
| S-EPMC7608831 | biostudies-literature
Project description:Bone regeneration is the ultimate goal of periodontal therapies, in which osteogenic differentiation of human periodontal ligament stem cells plays a critical role. The tripartite motif (TRIM)16, an E3 ubiquitin ligase, is downregulated in periodontal tissues of patients with periodontitis, while the role of TRIM16 in the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) is largely unknown. Firstly, we found that TRIM16 was increased throughout the osteogenic media induced differentiation of hPDLSCs. Then overexpression plasmids and specific short-hairpin RNAs (shRNAs) were constructed to manipulate the expression of target molecules. TRIM16 significantly promoted alkaline phosphatase activity, mineralized nodule formation, and positively regulated the expression of osteo-specific markers RUNX2, COL1A1 and OCN except the mRNA of RUNX2. Mechanistically, TRIM16 serves as a pivotal factor that stabilizes RUNX2 protein levels by decreasing CHIP-mediated K48-linked ubiquitination degradation of the RUNX2 protein. This study identified a novel mechanism of TRIM16 in regulating stability of the RUNX2 protein, which promoted the osteogenic differentiation of hPDLSCs. TRIM16 may be a potential target of stem cell based-bone regeneration for periodontal therapies.
| S-EPMC7817816 | biostudies-literature
Project description:RelA-mediated NF-?B canonical signaling promotes mesenchymal progenitor cell (MPC) proliferation, but inhibits differentiation of mature osteoblasts (OBs) and thus negatively regulates bone formation. Previous studies suggest that NF-?B RelB may also negatively regulate bone formation through noncanonical signaling, but they involved a complex knockout mouse model, and the molecular mechanisms involved were not investigated. Here, we report that RelB(-/-) mice develop age-related increased trabecular bone mass associated with increased bone formation. RelB(-/-) bone marrow stromal cells expanded faster in vitro and have enhanced OB differentiation associated with increased expression of the osteoblastogenic transcription factor, Runt-related transcription factor 2 (Runx2). In addition, RelB directly targeted the Runx2 promoter to inhibit its activation. Importantly, RelB(-/-) bone-derived MPCs formed bone more rapidly than wild-type cells after they were injected into a murine tibial bone defect model. Our findings indicate that RelB negatively regulates bone mass as mice age and limits bone formation in healing bone defects, suggesting that inhibition of RelB could reduce age-related bone loss and enhance bone repair.
| S-EPMC3961566 | biostudies-literature
Project description:The LIM-homeodomain transcription factor Lmx1b plays a key role in body pattern formation during development. Although Lmx1b is essential for the normal development of multiple tissues, its regulatory mechanism in bone cells remains unclear. Here, we demonstrated that Lmx1b negatively regulates bone morphogenic protein 2 (BMP2)-induced osteoblast differentiation. Overexpressed Lmx1b in the osteoblast precursor cells inhibited alkaline phosphatase (ALP) activity and nodule formation, as well as the expression of osteoblast maker genes, including runt-related transcription factor 2 (Runx2), alkaline phosphatase (Alpl), bone sialoprotein (Ibsp), and osteocalcin (Bglap). Conversely, the knockdown of Lmx1b in the osteoblast precursors enhanced the osteoblast differentiation and function. Lmx1b physically interacted with and repressed the transcriptional activity of Runx2 by reducing the recruitment of Runx2 to the promoter region of its target genes. In vivo analysis of BMP2-induced ectopic bone formation revealed that the knockdown of Lmx1b promoted osteogenic differentiation and bone regeneration. Our data demonstrate that Lmx1b negatively regulates osteoblast differentiation and function through regulation of Runx2 and provides a molecular basis for therapeutic targets for bone diseases.
| S-EPMC9103437 | biostudies-literature
Project description:Although CCAAT/enhancer-binding protein beta (C/EBPbeta) is involved in osteocalcin gene expression in osteoblast in vitro, the physiological importance of and molecular mechanisms governing C/EBPbeta in bone formation remain to be elucidated. In particular, it remains unclear whether C/EBPbeta acts as a homodimer or a heterodimer with other proteins during osteoblast differentiation. Here, deletion of the C/EBPbeta gene from mice resulted in delayed bone formation with concurrent suppression of chondrocyte maturation and osteoblast differentiation. The expression of type X collagen as well as chondrocyte hypertrophy were suppressed in mutant bone, providing new insight into the possible roles of C/EBPbeta in chondrocyte maturation. In osteoblasts, luciferase reporter, gel shift, DNAP, and ChIP assays demonstrated that C/EBPbeta heterodimerized with activating transcription factor 4 (ATF4), another basic leucine zipper transcription factor crucial for osteoblast maturation. This complex interacted and transactivated osteocalcin-specific element 1 (OSE1) of the osteocalcin promoter. C/EBPbeta also enhanced the synergistic effect of ATF4 and Runx2 on osteocalcin promoter transactivation by enhancing their interaction. Thus, our results provide evidence that C/EBPbeta is a crucial cofactor in the promotion of osteoblast maturation by Runx2 and ATF4.
| S-EPMC2592674 | biostudies-literature
Project description:The adaptor protein CrkII is involved in several biological activities, including mitogenesis, phagocytosis, and cytoskeleton reorganization. Previously, we demonstrated that CrkII plays an important role in osteoclast differentiation and function through Rac1 activation both in vitro and in vivo. In this study, we investigated whether CrkII also regulates the differentiation and function of another type of bone cells, osteoblasts. Overexpression of CrkII in primary osteoblasts inhibited bone morphogenetic protein (BMP) 2-induced osteoblast differentiation and function, whereas knockdown of CrkII expression exerted the opposite effect. Importantly, CrkII strongly enhanced c-Jun-N-terminal kinase (JNK) phosphorylation, and the CrkII overexpression-mediated attenuation of osteoblast differentiation and function was recovered by JNK inhibitor treatment. Furthermore, transgenic mice overexpressing CrkII under control of the alpha-1 type I collagen promoter exhibited a reduced bone mass phenotype. Together, these results indicate that CrkII negatively regulates osteoblast differentiation and function through JNK phosphorylation. Given that CrkII acts as a negative and positive regulator of osteoblast and osteoclast differentiation, respectively, the regulation of CrkII expression in bone cells may help to develop new strategies to enhance bone formation and inhibit bone resorption.
| S-EPMC6802640 | biostudies-literature
Project description:The circadian clock regulates various physiological processes, including bone metabolism. The nuclear receptors Reverbs, comprising Rev-erb? and Rev-erb?, play a key role as transcriptional regulators of the circadian clock. In this study, we demonstrate that Rev-erbs negatively regulate differentiation of osteoclasts and osteoblasts. The knockdown of Rev-erb? in osteoclast precursor cells enhanced receptor activator of nuclear factor-?B ligand (RANKL)-induced osteoclast formation, as well as expression of nuclear factor of activated T cells 1 (NFATc1), osteoclast-associated receptor (OSCAR), and tartrate-resistant acid phosphatase (TRAP). The overexpression of Rev-erb? leads to attenuation of the NFATc1 expression via inhibition of recruitment of c-Fos to the NFATc1 promoter. The overexpression of Rev-erb? in osteoblast precursors attenuated the expression of osteoblast marker genes including Runx2, alkaline phosphatase (ALP), bone sialoprotein (BSP), and osteocalcin (OC). Rev-erb? interfered with the recruitment of Runx2 to the promoter region of the target genes. Conversely, knockdown of Reverb? in the osteoblast precursors enhanced the osteoblast differentiation and function. In addition, Rev-erb? negatively regulated osteoclast and osteoblast differentiation by suppressing the p38 MAPK pathway. Furthermore, intraperitoneal administration of GSK4112, a Rev-erb agonist, protects RANKL-induced bone loss via inhibition of osteoclast differentiation in vivo . Taken together, our results demonstrate a molecular mechanism of Rev-erbs in the bone remodeling, and provide a molecular basis for a potential therapeutic target for treatment of bone disease characterized by excessive bone resorption.
| S-EPMC6999712 | biostudies-literature