Expression of Gal4-VP16 and Gal4-DNA binding domain under the control of the T lymphocyte-specific lck proximal promoter in transgenic mice

Ryu, Chun Jeih; Whitehurst, Charles E.; Chen, Jianzhu

doi:10.5483/bmbrep.2008.41.8.575

Cited by 4 publications

(4 citation statements)

References 23 publications

(36 reference statements)

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“…Flow cytometry was performed using biotin‐conjugated 297‐D4 and/or hESC‐specific antibodies as described previously . Primary antibodies used were anti‐SSEA‐1, anti‐SSEA‐3 (R&D Systems, Minneapolis, MN, http://www.rndsystems.com), anti‐TRA‐1–60, anti‐TRA‐1–81 (Millipore, Billerica, MA, http://www.millipore.com), 297‐D4, polyclonal anti‐BAP31, ployclonal anti‐Integrin β1, polyclonal anti‐E‐cadherin, or polyclonal anti‐EpCAM antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, http://www.scbt.com).…”

Section: Methodsmentioning

confidence: 99%

B-Cell Receptor-Associated Protein 31 Regulates Human Embryonic Stem Cell Adhesion, Stemness, and Survival via Control of Epithelial Cell Adhesion Molecule

et al. 2014

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B-Cell receptor-associated protein 31 (BAP31) regulates the export of secreted membrane proteins from the endoplasmic reticulum (ER) to the downstream secretory pathway. Previously, we generated a monoclonal antibody 297-D4 against the surface molecule on undifferentiated human embryonic stem cells (hESCs). Here, we found that 297-D4 antigen was localized to pluripotent hESCs and downregulated during early differentiation of hESCs and identified that the antigen target of 297-D4 was BAP31 on the hESC-surface. To investigate the functional role of BAP31 in hESCs, BAP31 expression was knocked down by small interfering RNA. BAP31 depletion impaired hESC self-renewal and pluripotency and drove hESC differentiation into multicell lineages. BAP31 depletion hindered hESC proliferation by arresting cell cycle at G0/G1 phase and inducing caspase-independent cell death. Interestingly, BAP31 depletion reduced hESC adhesion to extracellular matrix (ECM). Analysis of cell surface molecules showed decreased expression of epithelial cell adhesion molecule (EpCAM) in BAP31-depleted hESCs, while ectopic expression of BAP31 elevated the expression of EpCAM. EpCAM depletion also reduced hESC adhesion to ECM, arrested cell cycle at G0/G1 phase and induced cell death, producing similar effects to those of BAP31 depletion. BAP31 and EpCAM were physically associated and colocalized at the ER and cell surface. Both BAP31 and EpCAM depletion decreased cyclin D1 and E expression and suppressed PI3K/Akt signaling, suggesting that BAP31 regulates hESC stemness and survival via control of EpCAM expression. These findings provide, for the first time, mechanistic insights into how BAP31 regulates hESC stemness and survival via control of EpCAM expression.

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Section: Methodsmentioning

confidence: 99%

B-Cell Receptor-Associated Protein 31 Regulates Human Embryonic Stem Cell Adhesion, Stemness, and Survival via Control of Epithelial Cell Adhesion Molecule

et al. 2014

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“…Flow cytometric analysis was performed using hESC marker‐specific antibodies and anti‐hnRNP A2/B1 antibodies as described previously []. Briefly, hESCs were detached with collagenase IV and further incubated with cell dissociation buffer (Invitrogen, Seoul, Korea, http://www.lifetechnologies.com) as described before .…”

Section: Methodsmentioning

confidence: 99%

Heterogeneous Nuclear Ribonucleoprotein A2/B1 Regulates the Self-Renewal and Pluripotency of Human Embryonic Stem Cells Via the Control of the G1/S Transition

et al. 2013

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Self-renewal and pluripotency of human embryonic stem cells (hESCs) are a complex biological process for maintaining hESC stemness. However, the molecular mechanisms underlying these special properties of hESCs are not fully understood. Heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) is a multifunctional RNAbinding protein whose expression is related to cell proliferation and carcinogenesis. In this study, we found that hnRNP A2/B1 expression was localized to undifferentiated hESCs and decreased upon differentiation of hESCs. hnRNP A2/B1 knockdown reduced the number of alkaline phosphatase-positive colonies in hESCs and led to a decrease in the expression of pluripotency-associated transcription factors OCT4, NANOG, and SOX2, indicating that hnRNP A2/B1 is essential for hESC self-renewal and pluripotency. hnRNP A2/B1 knockdown increased the expression of gene markers associated with the early development of three germ layers, and promoted the process of epithelial-mesenchymal transition, suggesting that hnRNP A2/B1 is required for maintaining the undifferentiated and epithelial phenotypes of hESCs. hnRNP A2/B1 knockdown inhibited hESC proliferation and induced cell cycle arrest in the G0/G1 phase before differentiation via degradation of cyclin D1, cyclin E, and Cdc25A. hnRNP A2/B1 knockdown increased p27 expression and induced phosphorylation of p53 and Chk1, suggesting that hnRNP A2/B1 also regulates the G1/S transition of hESC cell cycle through the control of p27 expression and p53 and Chk1 activity. Analysis of signaling molecules further revealed that hnRNP A2/B1 regulated hESC proliferation in a PI3K/Akt-dependent manner. These findings provide for the first time mechanistic insights into how hnRNP A2/B1 regulates hESC self-renewal and pluripotency. STEM CELLS

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“…Low tissue specificity and efficiency of exogenous gene expression are the two major obstacles in tumor-targeted gene therapy. It is possible to achieve specific and efficient expression of a target gene in tumor cells by the regulation of tumor-specific promoters (TSP) and two-step transcriptional amplification (TSTA) (1,2). Multiple promoters have been used in targeted gene therapy for ovarian cancer, including secretory leukocyte protease inhibitor (SLPI), the ovary-specific promoter OSP1 and the human epithelial tissue-specific promoter, transcription factor HES1.…”

Section: Introductionmentioning

confidence: 99%

Gene therapy for human ovarian cancer cells using efficient expression of Fas gene combined with γδT cells

Lin

Zeng

et al. 2017

Molecular Medicine Reports

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Low tissue specificity and efficiency of exogenous gene expression are the two major obstacles in tumor‑targeted gene therapy. The Fas cell surface death receptor (Fas)/Fas ligand pathway is one of the primary pathways responsible for the regulation of cell apoptosis. The aim of the present study was to explore whether the regulation of tumor specific promoters and a two‑step transcriptional amplification system (TSTA) assured efficient, targeted expression of their downstream Fas gene in human ovarian cancer cells, and to assess the killing effect of γδT cells on these cells with high Fas expression. Three shuttle plasmids containing different control elements of the human telomerase reverse transcriptase (hTERT) promoter and/or TSTA were constructed and packaged into adenovirus 5 (Ad5) vectors for the expression of exogenous Fas gene. The human ovarian cancer cell line SKOV3 and a control human embryonic lung fibroblast cell line were transfected with Ad5‑hTERT‑Fas or Ad5‑hTERT‑TSTA‑Fas. Fas mRNA and protein expression were examined by reverse transcription‑quantitative polymerase chain reaction and western blot analysis. γδT lymphocytes were isolated, cultured and mixed at different ratios with SKOV3 cells with Fas expression in order to assess the killing effect of γδT cells. hTERT promoter induced the specific expression of FAS gene in SKOV3 cells, and the TSTA strategy increased FAS expression by 14.2‑fold. The killing effect of γδT cells increased with the expression level of Fas and the effector‑target cell ratio. The killing rate for SKOV3 cells with high FAS expression was 72.5% at an effector‑target cell ratio of 40:1. The regulators of hTERT promoter and TSTA assure the efficient and targeted expression of their downstream Fas gene in SKOV3 cells. The killing effect of γδT cells for ovarian cancer cells with relatively high Fas expression was improved.

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Expression of Gal4-VP16 and Gal4-DNA binding domain under the control of the T lymphocyte-specific lck proximal promoter in transgenic mice

Cited by 4 publications

References 23 publications

B-Cell Receptor-Associated Protein 31 Regulates Human Embryonic Stem Cell Adhesion, Stemness, and Survival via Control of Epithelial Cell Adhesion Molecule

B-Cell Receptor-Associated Protein 31 Regulates Human Embryonic Stem Cell Adhesion, Stemness, and Survival via Control of Epithelial Cell Adhesion Molecule

Heterogeneous Nuclear Ribonucleoprotein A2/B1 Regulates the Self-Renewal and Pluripotency of Human Embryonic Stem Cells Via the Control of the G1/S Transition

Gene therapy for human ovarian cancer cells using efficient expression of Fas gene combined with γδT cells

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