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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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Journal ArticleDOI
15 May 2009-PLOS ONE
TL;DR: It is found that inhibition of Erk5 decreased cancer cell proliferation and also sensitized these cells to the action of anti-HER2 therapies, and may represent a future therapeutic target.
Abstract: Background Breast cancer is the most common neoplasia in women. Even though advances in its treatment have improved disease outcome, some patients relapse. Therefore, attempts to better define the molecular determinants that drive breast cancer cell proliferation may help in defining potential therapeutic targets. Mitogen-activated protein kinases (MAPK) play important roles in tumorigenesis. One of them, Erk5, has been linked to the proliferation of breast cancer cells in vitro. Here we have investigated the expression and prognostic value of Erk5 in human breast cancer. Methodology/Principal Findings Animal and cellular models were used to study Erk5 expression and function in breast cancer. In 84 human breast tumours the expression of Erk5 was analyzed by immunohistochemistry. Active Erk5 (pErk5) was studied by Western blotting. Correlation of Erk5 with clinicopathological parameters and with disease-free survival in early stage breast cancer patients was analyzed. Expression of Erk5 was detected in most patients, and overexpression was found in 20%. Active Erk5 was present in a substantial number of samples, as well as in tumours from an animal breast cancer model. Overexpression of Erk5 was associated with a decrease in disease-free survival time, which was independent of other clinicopathological parameters of prognosis. Transient transfection of a short hairpin RNA (shRNA) targeting Erk5, and a stable cell line expressing a dominant negative form of Erk5 (Erk5AEF), were used to investigate the influence of Erk5 on drugs used in the clinic to treat breast tumours. We found that inhibition of Erk5 decreased cancer cell proliferation and also sensitized these cells to the action of anti-HER2 therapies. Conclusions/Significance Overexpression of Erk5 is an independent predictor of disease-free survival in breast cancer, and may represent a future therapeutic target.

106 citations

Journal ArticleDOI
TL;DR: Recurrent amplification of chromosome 19q13.1-2 has been reported in pancreatic cancer, but the exact target gene has not been identified and actinin-4 contributes to the invasive growth of pancreatic ductal carcinoma, and ACTN4 is one of the candidate oncogenes in this chromosome locus.
Abstract: Purpose: An invasive growth pattern is one of the hallmarks of pancreatic ductal carcinoma. Actinin-4 is an actin-binding protein associated with enhanced cell motility, invasive growth, and lymph node metastasis. Actinin-4 might play an important role in the development and progression of pancreatic cancer. Experimental Design: The expression of actinin-4 was examined immunohistochemically in 173 cases of invasive pancreatic ductal carcinoma. The copy number of the actinin-4 ( ACTN4 ) gene was calculated by fluorescence in situ hybridization. The expression of actinin-4 was stably knocked down by short hairpin RNA, and tumorigenicity was evaluated by orthotopic implantation into mice with severe combined immunodeficiency. Results: The expression level of actinin-4 was increased in 109 (63.0%) of 173 cases of pancreatic cancer. Kaplan-Meier survival curves revealed that patients with increased expression of actinin-4 had a significantly poorer outcome ( P = 0.00001, log-rank test). Multivariate analysis by the Cox proportional hazard model showed that high expression of actinin-4 was the most significant independent negative predictor of survival (hazard ratio, 2.33; P = 0.000009). Amplification (defined as more than four copies per interphase nucleus) of the ACTN4 gene was detected in 11 (37.9%) of 29 cases showing increased expression of actinin-4. Knockdown of actinin-4 expression inhibited the destructive growth of cancer cells in the pancreatic parenchyma. Conclusion: Recurrent amplification of chromosome 19q13.1-2 has been reported in pancreatic cancer, but the exact target gene has not been identified. Actinin-4 contributes to the invasive growth of pancreatic ductal carcinoma, and ACTN4 is one of the candidate oncogenes in this chromosome locus.

106 citations

Journal ArticleDOI
TL;DR: Functional evidence is provided that OPN contributes to breast tumor growth as well, in addition to the widely reported roles of OPN in late stages of tumor progression.
Abstract: Elevated expression of osteopontin (OPN), a secreted phosphoglycoprotein, is frequently associated with many transformed cell lines. Various studies suggest that OPN may contribute to tumor progression as well as metastasis in multiple tumor types. High levels of OPN have been reported in patients with metastatic cancers, including breast. We found that the expression of OPN corroborates with the aggressive phenotype of the breast cancer cells i.e. the expression of OPN is acquired as the breast cancer cells become more aggressive. To assess the role(s) of OPN in breast carcinoma, expression of endogenous OPN was knocked down in metastatic MDA-MB-435 human breast carcinoma cells using RNA interference. We targeted multiple regions of the OPN transcript for RNA interference, along with ‘scrambled’ and ‘non-targeting siRNA pool’ controls to distinguish between target-specific and potential off-target effects including interferon-response gene (PeIF2-α) induction. The OPN knockdown by shRNA suppressed tumor take in immunocompromised mice. The ‘silenced’ cells also showed significantly lower invasion and migration in modified Boyden chamber assays and reduced ability to grow in soft agar. Thus, in addition to the widely reported roles of OPN in late stages of tumor progression, these results provide functional evidence that OPN contributes to breast tumor growth as well.

106 citations

Journal ArticleDOI
TL;DR: It is demonstrated that coexpression of Gli2ΔC and Sox2 can rescue the expression of Hes5 and prevent premature neuronal differentiation in NE cells but cannot rescue its proliferation, revealing a novel transcriptional cascade, involving Gli 2 → Sox2 → Hes5, which maintains the undifferentiated state of telencephalic NE cells.
Abstract: Multipotential neural stem cells (NSCs) in the central nervous system (CNS) proliferate indefinitely and give rise to neurons, astrocytes, and oligodendrocytes. As NSCs hold promise for CNS regeneration, it is important to understand how their proliferation and differentiation are controlled. We show here that the expression of sox2 gene, which is essential for the maintenance of NSCs, is regulated by the Gli2 transcription factor, a downstream mediator of sonic hedgehog (Shh) signaling: Gli2 binds to an enhancer that is vital for sox2 expression in telencephalic neuroepithelial (NE) cells, which consist of NSCs and neural precursor cells. Overexpression of a truncated form of Gli2 (Gli2DeltaC) or Gli2-specific short hairpin RNA (Gli2 shRNA) in NE cells in vivo and in vitro inhibits cell proliferation and the expression of Sox2 and other NSC markers, including Hes1, Hes5, Notch1, CD133, and Bmi1. It also induces premature neuronal differentiation in the developing NE cells. In addition, we show evidence that Sox2 expression decreases significantly in the developing neuroepithelium of Gli2-deficient mice. Finally, we demonstrate that coexpression of Gli2DeltaC and Sox2 can rescue the expression of Hes5 and prevent premature neuronal differentiation in NE cells but cannot rescue its proliferation. Thus these data reveal a novel transcriptional cascade, involving Gli2 --> Sox2 --> Hes5, which maintains the undifferentiated state of telencephalic NE cells.

106 citations

Journal ArticleDOI
TL;DR: This work applies retrovirus-mediated RNAi to epithelial Madin-Darby canine kidney cells, whose properties limit the applicability of other RNAi methods, and shows that this system can be used to simultaneously target two genes, giving rise to double knockdowns.
Abstract: RNA interference (RNAi) is a ubiquitous mechanism of eukaryotic gene regulation that can be exploited for specific gene silencing. Retroviruses have been successfully used for stable expression of short hairpin RNAs in mammalian cells, leading to persistent inhibition of gene expression by RNAi. Here, we apply retrovirus-mediated RNAi to epithelial Madin-Darby canine kidney cells, whose properties limit the applicability of other RNAi methods. We demonstrate efficient suppression of a set of 13 target genes by retroviral coexpression of short hairpin RNAs and a selectable marker. We characterize the resulting knockdown cell populations with regard to composition and stability, and examine the usefulness of proposed guidelines for choosing RNAi target sequences. Finally, we show that this system can be used to simultaneously target two genes, giving rise to double knockdowns. Thus, retrovirus-mediated RNAi is a convenient method for gene silencing in Madin-Darby canine kidney cells, and is likely to be applicable to virtually any mammalian cell.

106 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
2023804
2022477
2021384
2020454
2019541
2018518