Topic
Small hairpin RNA
About: Small hairpin RNA is a research topic. Over the lifetime, 9279 publications have been published within this topic receiving 285471 citations.
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TL;DR: Targeting the hypoxic niche, and more specifically hypoxia-inducible factor-2, may represent a viable strategy to inhibit angiogenesis in multiple myeloma and progression of this disease.
Abstract: Background Multiple myeloma is an incurable malignancy of bone marrow plasma cells. Progression of multiple myeloma is accompanied by an increase in bone marrow angiogenesis. Studies from our laboratory suggest a role for the CXCL12 chemokine in this process, with circulating levels of CXCL12 correlating with bone marrow angiogenesis in patients with multiple myeloma. While the mechanisms responsible for aberrant plasma cell expression of CXCL12 remain to be determined, studies in other systems suggest a role for hypoxia and hypoxia-inducible transcription factors.Design and Methods The expression of hypoxia-inducible factor protein was examined in patients’ bone marrow biopsy specimens using immunohistochemistry. The hypoxic regulation of CXCL12 was examined in multiple myeloma plasma cell lines using polymerase chain reaction and western blotting. The role of hypoxia-inducible factors-1 and -2 in the regulation of CXCL12 expression was examined using over-expression and short hairpin RNA knockdown constructs, electrophoretic mobility shift assays and chromatin immunoprecipitation. The contribution of CXCL12 to hypoxia-induced angiogenesis was examined in vivo using a subcutaneous murine model of neovascularization.Results Strong hypoxia-inducible factor-2 protein expression was detected in CD138+ multiple myeloma plasma cells in patients’ biopsy specimens. Prolonged exposure to hypoxia strongly up-regulated CXCL12 expression in multiple myeloma plasma cells and hypoxia-inducible factor-2 was found to play a key role in this response. Promoter analyses revealed increased hypoxia-inducible factor-2 binding to the CXCL12 promoter under hypoxic conditions. Over-expression of hypoxia-inducible factor in multiple myeloma plasma cells strongly induced in vivo angiogenesis, and administration of a CXCL12 antagonist decreased hypoxia-inducible factor-induced angiogenesis.Conclusions Hypoxia-inducible factor-2 is a newly identified regulator of CXCL12 expression in multiple myeloma plasma cells and a major contributor to multiple myeloma plasma cell-induced angiogenesis. Targeting the hypoxic niche, and more specifically hypoxia-inducible factor-2, may represent a viable strategy to inhibit angiogenesis in multiple myeloma and progression of this disease.
80 citations
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TL;DR: It is confirmed that in the long bones of newborn mice, VEGF-D and VEGFR-3 are expressed in the osteoblasts of the growing plate and suggested that VEGf-D is a downstream effector of VEGFs in osteogenesis and plays a critical role in osteoblast maturation.
80 citations
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TL;DR: Overexpression of cyclin D1 enhanced the proliferation and invasive potential of human glioblastoma cells, while reducing apoptosis, and may provide a new gene therapy approach for patients with malignant glioma.
Abstract: Elevated cyclin D1 (CCND1) in human glioblastoma correlates with poor clinical prognosis. In this study, the human glioblastoma cell lines SHG-44 and U251 were stably transfected with short hairpin RNA (shRNA) targeting cyclin D1 or with ectogenic cyclin D1 by lentivirus-mediated transfection. Glioblastoma cells overexpressing or underexpressing cyclin D1 were then examined by in vitro growth assays, apoptosis assays, cell cycle analysis, and invasion assays. Cyclin D1 knockdown in SHG-44 cells inhibited cell proliferation, induced apoptosis, and attenuated migration across Matrigel, a model of invasive capacity. Western blot analysis and quantitative reverse-transcription polymerase chain reaction (RT-PCR) revealed that cells underexpressing CCND1 exhibited decreased multidrug resistance protein 1 (MDR1) and B-cell lymphoma-2 (Bcl-2) expression, but enhanced apoptosis effector caspase-3 expression. In contrast, cyclin D1 overexpression promoted cell proliferation, attenuated apoptosis, and enhanced invasive capacity. Furthermore, cyclin D1 overexpression was associated with increased expression of MDR1 and Bcl-2, and decreased caspase-3 expression. Results using the U251 cell line confirmed the effects of CCND1-targeted shRNA and lentivirus-mediated overexpression on proliferation and apoptosis of glioblastoma cells. Overexpression of cyclin D1 enhanced the proliferation and invasive potential of human glioblastoma cells, while reducing apoptosis. The ability to suppress the malignant phenotype by downregulating cyclin D1 expression may provide a new gene therapy approach for patients with malignant glioma.
80 citations
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TL;DR: Through its effects on MITF deubiquitination, USP13 was observed to modulate expression of MITF downstream target genes and, thereby, to be essential for melanoma growth in soft agar and in nude mice, suggesting that as a potentially drugable protease,USP13 might be a viable therapeutic target for melanomas.
Abstract: The microphthalmia-associated transcription factor (MITF) is essential for melanocyte development. Mutation-induced MAPK pathway activation is common in melanoma and induces MITF phosphorylation, ubiquitination, and proteolysis. Little is known about the enzymes involved in MITF ubiquitination/deubiquitination. Here we report the identification of a deubiquitinating enzyme, named ubiquitin-specific protease 13 (USP13) that appears to be responsible for MITF deubiquitination, utilizing a short hairpin RNA library against known deubiquitinating enzymes. Through deubiquitination, USP13 stabilizes and upregulates MITF protein levels. Conversely, suppression of USP13 (through knockdown) leads to dramatic loss of MITF protein, but not messenger RNA. Through its effects on MITF deubiquitination, USP13 was observed to modulate expression of MITF downstream target genes and, thereby, to be essential for melanoma growth in soft agar and in nude mice. These observations suggest that as a potentially drugable protease, USP13 might be a viable therapeutic target for melanoma.
80 citations
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TL;DR: The results suggest that both G3BP isoforms may act as negative regulators of p53, and members of the Ras network of proteins, Ras-GTPase activating protein-SH3-domain-binding proteins 1 and 2 (G3BP1 and 2), bind to p53 in vitro and in vivo.
Abstract: Inactivation of the p53 tumor suppressor pathway is a critical step in human tumorigenesis. In addition to mutations, p53 can be functionally silenced through its increased degradation, inhibition of its transcriptional activity and/or its inappropriate subcellular localization. Using a proteomic approach, we have found that members of the Ras network of proteins, Ras-GTPase activating protein-SH3-domain-binding proteins 1 and 2 (G3BP1 and 2), bind to p53 in vitro and in vivo. Our data show that expression of G3BPs leads to the redistribution of p53 from the nucleus to the cytoplasm. The G3BP2 isoform additionally associated with murine double minute 2 (MDM2), a negative regulator of p53. G3BP2 expression resulted in significant reduction in MDM2-mediated p53 ubiquitylation and degradation. Interestingly, MDM2 was also stabilized in G3BP2-expressing cells and its ability to ubiquitylate itself was compromised. Accordingly, short hairpin RNA (shRNA)-mediated knockdown of G3BP2 caused a reduction in MDM2 protein levels. Furthermore, expression of shRNA targeting either G3BP1 or G3BP2 in human cancer cell lines resulted in marked upregulation of p53 levels and activity. Our results suggest that both G3BP isoforms may act as negative regulators of p53.
80 citations