Identification of an RNA element that confers post-transcriptional repression of connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene: Similarities to retroviral RNA-protein interactions

Kubota, Satoshi; Kondo, Seiji; Eguchi, Takanori; Hattori, Takako; Nakanishi, Tohru; Pomerantz, Roger J.; Takigawa, Masaharu

doi:10.1038/sj.onc.1203835

Cited by 51 publications

(57 citation statements)

References 45 publications

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“…The locations of the pentameric instability consensus signals and stem-loop structure causing premature termination of transcription are depicted by open asterisk and open box, respectively. Construction of the original plasmids (pGL3SA2, pGL3SA3, pGL3SA5) was described in a previous report (Kubota et al, 2000). Right panel: Analysis of the synthesized RNAs in 6% urea-denatured PAGE.…”

Section: Discussionmentioning

confidence: 99%

“…In this study, we showed that a hypoxic-inducible nuclear/cytoplasmic protein of approximately 35 kDa bound to the minimal sequence of ctgf/ccn2 3 0 -UTR mRNA. This 84 bp 3 0 -UTR sequence, termed CAESAR, was capable of forming a unique stem-loop secondary structure (Kubota et al, 2000). When linked in cis, CAESAR/SA5 was found to maintain the basal gene expression at a low level through the interference with translation process (Kubota et al, 2000, which can be defined as the constitutive function of CAESAR/SA5.…”

Section: Regulation Of Ctgf/ccn2 Gene Stability Under Hypoxia S Kondomentioning

confidence: 99%

“…This 84 bp 3 0 -UTR sequence, termed CAESAR, was capable of forming a unique stem-loop secondary structure (Kubota et al, 2000). When linked in cis, CAESAR/SA5 was found to maintain the basal gene expression at a low level through the interference with translation process (Kubota et al, 2000, which can be defined as the constitutive function of CAESAR/SA5. In addition to this classical function, we here uncovered another dynamic function of CAESAR as a post-transcriptional regulatory element, which was exerted by regulating mRNA stability.…”

Section: Regulation Of Ctgf/ccn2 Gene Stability Under Hypoxia S Kondomentioning

confidence: 99%

“…We further analysed the possible involvement of the posttranscriptional regulatory mechanism through the 3 0 -UTR. The ctgf/ccn2 3 0 -UTR contains a cis-acting element of structure-anchored repression (CAESAR; Figure 3a), which is a post-transcriptional RNA regulatory element of 84 bases, and has an important role in determining ctgf/ccn2 basal expression level in Cos7 cells (Kubota et al, 2000). Suspecting another dynamic function therein, we performed cell-free RNA decay assays using RNA transcripts of this region.…”

Section: Gene Expression Profile Of Hypoxic Hcs-2/8 Cellsmentioning

confidence: 99%

“…We had formerly constructed firefly luciferase-ctgf/ccn2 chimeric gene constructs (pGL3UTRS, pGL3SA2, pGL3SA3, pGL3SA5), as described by Kubota et al (1999Kubota et al ( , 2000. For the preparation of template plasmids for in vitro transcription of full-length ctgf/ccn2 3 0 -UTR, these pGL3 derivatives were double-digested with XbaI and EcoRI, and the 3 0 -UTR fragments were purified and subcloned between the corresponding sites in pGEM3Zf( þ ) (Promega, Madison, WI, USA).…”

Section: Preparation Of Probesmentioning

confidence: 99%

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Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3′-untranslated region interacting with a cellular protein in human chondrosarcoma cells

et al. 2005

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Connective tissue growth factor (CTGF/CCN2) can be induced by various forms of stress such as exposure to high glucose, mechanical load, or hypoxia. Here, we investigated the molecular mechanism involved in the induction of ctgf/ccn2 by hypoxia in a human chondrosarcoma cell line, HCS-2/8. Hypoxia increased the ctgf/ ccn2 mRNA level by altering the 3 0 -untranslated region (UTR)-mediated mRNA stability without requiring de novo protein synthesis. After a series of extensive analyses, we eventually found that the cis-repressive element of 84 bases within the 3 0 -UTR specifically bound to a cytoplasmic/nuclear protein. By conducting a UV crosslinking assay, we found the cytoplasmic/nuclear protein to be a 35 kDa molecule that bound to the ciselement in a hypoxia-inducible manner. These results suggest that a cis-element in the 3 0 -UTR of ctgf/ccn2 mRNA and trans-factor counterpart(s) play an important role in the post-transcriptional regulation by determining the stability of ctgf/ccn2 mRNA.

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Section: Discussionmentioning

confidence: 99%

Section: Regulation Of Ctgf/ccn2 Gene Stability Under Hypoxia S Kondomentioning

confidence: 99%

Section: Regulation Of Ctgf/ccn2 Gene Stability Under Hypoxia S Kondomentioning

confidence: 99%

Section: Gene Expression Profile Of Hypoxic Hcs-2/8 Cellsmentioning

confidence: 99%

Section: Preparation Of Probesmentioning

confidence: 99%

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Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3′-untranslated region interacting with a cellular protein in human chondrosarcoma cells

et al. 2005

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A coding RNA segment that enhances the ribosomal recruitment of chicken ccn1 mRNA

Mukudai

Kubota

Eguchi

et al. 2010

J of Cellular Biochemistry

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CCN1, a member of the CCN family of proteins, plays important physiological or pathological roles in a variety of tissues. In the present study, we initially found a highly guanine-cytosine (GC)-rich region of approximately 200 bp near the 5'-end of the open reading frame, which was always truncated by amplification of the corresponding cDNA region through the conventional polymerase chain reaction. An RNA in vitro folding assay and selective ribonuclease digestion of the corresponding segment of the ccn1 mRNA confirmed the involvement of a stable secondary structure. Subsequent RNA electromobility-shift assays demonstrated the specific binding of some cytoplasmic factor(s) in chicken embryo fibroblasts to the RNA segment. Moreover, the corresponding cDNA fragment strongly enhanced the expression of the reporter gene in cis at the 5'-end, but did not do so at the 3'-end. According to the results of a ribosomal assembly test, the effect of the mRNA segment can predominantly be ascribed to the enhancement of transport and/or entry of the mRNA into the ribosome. Finally, the minimal GC-rich mRNA segment that was predicted and demonstrated to form a secondary structure was confirmed to be a functional regulatory element. Thus, we here uncover a novel dual-functionality of the mRNA segment in the ccn1 open reading frame, which segment acts as a cis-element that mediates posttranscriptional gene regulation, while retaining the information for the amino acid sequence of the resultant protein.

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Role of CTGF/HCS24/ecogenin in skeletal growth control

Takigawa¹,

Nakanishi²,

Kubota³

et al. 2003

Journal Cellular Physiology

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173

185

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Connective tissue growth factor/hypertrophic chondrocyte-specific gene product 24 (CTGF/Hcs24) is a multifunctional growth factor for chondrocytes, osteoblasts, and vascular endothelial cells. CTGF/Hcs24 promotes the proliferation and maturation of growth cartilage cells and articular cartilage cells in culture and hypertrophy of growth cartilage cells in culture. The factor also stimulates the proliferation and differentiation of cultured osteoblastic cells. Moreover, CTGF/ Hcs24 promotes the adhesion, proliferation, and migration of vascular endothelial cells, as well as induces tube formation by the cells and strong angiogenesis in vivo. Because angiogenesis is critical for the replacement of cartilage with bone at the final stage of endochondral ossification and because gene expression of CTGF/ Hcs24 predominates in hypertrophic chondrocytes in the physiological state, a major physiological role for this factor should be the promotion of the entire process of endochondral ossification, with the factor acting on the above three types of cells as a paracrine factor. Thus, CTGF/Hcs24 should be called ''ecogenin: endochondral ossification genetic factor.'' In addition to hypertrophic chondrocytes, osteoblasts activated by various stimuli including wounding also express a significantly high level of CTGF/Hcs24. These findings in conjunction with in vitro findings about osteoblasts mentioned above suggest the involvement of CTGF/Hcs24 in intramembranous ossification and bone modeling/remodeling. Because angiogenesis is also critical for intramembranous ossification and bone remodeling, CTGF/ Hcs24 expressed in endothelial cells activated by various stimuli including wounding may also play important roles in direct bone formation. In conclusion, although the most important physiological role of CTGF/Hcs24 is ecogenin action, the factors also play important roles in skeletal growth and modeling/remodeling via its direct action on osteoblasts under both physiological and pathological conditions.

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Identification of an RNA element that confers post-transcriptional repression of connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene: Similarities to retroviral RNA-protein interactions

Cited by 51 publications

References 45 publications

Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3′-untranslated region interacting with a cellular protein in human chondrosarcoma cells

Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3′-untranslated region interacting with a cellular protein in human chondrosarcoma cells

A coding RNA segment that enhances the ribosomal recruitment of chicken ccn1 mRNA

Role of CTGF/HCS24/ecogenin in skeletal growth control

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