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All-optical regulation of gene expression in targeted cells.

ABSTRACT: Controllable gene expression is always a challenge and of great significance to biomedical research and clinical applications. Recently, various approaches based on extra-engineered light-sensitive proteins have been developed to provide optogenetic actuators for gene expression. Complicated biomedical techniques including exogenous genes engineering, transfection, and material delivery are needed. Here we present an all-optical method to regulate gene expression in targeted cells. Intrinsic or exogenous genes can be activated by a Ca(2+)-sensitive transcription factor nuclear factor of activated T cells (NFAT) driven by a short flash of femtosecond-laser irradiation. When applied to mesenchymal stem cells, expression of a differentiation regulator Osterix can be activated by this method to potentially induce differentiation of them. A laser-induced "Ca(2+)-comb" (LiCCo) by multi-time laser exposure is further developed to enhance gene expression efficiency. This noninvasive method hence provides an encouraging advance of gene expression regulation, with promising potential of applying in cell biology and stem-cell science.

SUBMITTER: Wang Y

PROVIDER: S-EPMC4061554 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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All-optical regulation of gene expression in targeted cells.

Wang Yisen Y He Hao H Li Shiyang S Liu Dayong D Lan Bei B Hu Minglie M Cao Youjia Y Wang Chingyue C

Scientific reports 20140618

Controllable gene expression is always a challenge and of great significance to biomedical research and clinical applications. Recently, various approaches based on extra-engineered light-sensitive proteins have been developed to provide optogenetic actuators for gene expression. Complicated biomedical techniques including exogenous genes engineering, transfection, and material delivery are needed. Here we present an all-optical method to regulate gene expression in targeted cells. Intrinsic or ...[more]

PMID: 24939233

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Project description:ObjectiveTo investigate the characteristics of Tiam1 gene expression in human cholangiocarcinoma tissues and benign bile duct tissues, and to analyze the correlations between Tiam1 gene expression and the degree of tumor differentiation, invasive and metastatic abilities. To explore the effect of targeted inhibiting Tiam1 gene expression on proliferation and migration activity of human cholangiocarcinoma cells.MethodsExpression of Tiam1 in 83 cases of cholangiocarcinoma tissues and 25 cases of benign bile tissues was detected using immunohistochemistry. The clinical data of patients with cholangiocarcinoma were collected. The correlations between Tiam1 gene expression and the clinicopathologic features in patients with cholangiocarcinoma were analyzed. The human cholangiocarcinoma RBE cells were divided into 3 groups. Cells in experimental group and control group were respectively transfected with Tiam1 shRNA lentiviral vectors and negative shRNA lentiviral control vectors. Cells in blank group received no treatment. Real-time PCR endogenesis was used to verify Tiam1 gene expression. Cell cycle experiments and MTT assay were used to measure cell proliferation activity. Transwell test was used to detect cell migration activity.ResultsThe negative rate Tiam1 protein expression in cholangiocarcinoma tissues was significantly higher than that in benign bile tissues (P<0.001). Tiam1 protein expression in cholangiocarcinoma tissues had correlations with cholangiocarcinoma differentiation degree, TNM stage and lymph node metastasis (P<0.05), and had no significant correlations with gender, age and distant metastasis (P>0.05). Real-time PCR detection indicated that Tiam1 expression of experimental group was significantly lower than that in control group and blank group (P<0.05), demonstrating that Tiam1 shRNA was effective on Tiam1 gene silencing in RBE cells. Cell cycle experiment showed that the percentage of S phase in cell cycle in experimental group was lower than that in control group and blank group (P<0.05), demonstrating that after the down-regulation of Tiam1 gene expression, the speed of cell proliferation was inhibited. MTT assay results showed that the total growth speed in experimental group was significantly lower than that in control group and blank group (P<0.05), indicating that the proliferation activity of cholangiocarcinoma cells was inhibited after targeted inhibition of Tiam1 gene expression. Transwell detection results showed that the metastasis rate in experimental group was significantly lower than that in control group and blank group (P<0.05), demonstrating that targeted inhibition of Tiam1 gene expression could significantly inhibit migration ability of RBE cells.ConclusionTiam1 expression significantly increased in cholangiocarcinoma tissues, and increased along with the degree of malignancy of cholangiocarcinoma. Targeted silencing Tiam1 expression could inhibit proliferation and migration activity of cholangiocarcinoma cells.

Project description:BCL2-family proteins have a central role in the mitochondrial apoptosis machinery and their expression is known to be deregulated in many cancer types. Effort in the development of small molecules that selectively target anti-apoptotic members of this family i.e., Bcl-2, Bcl-xL, Mcl-1 recently opened novel therapeutic opportunities. Among these apoptosis-inducing agents, BH3-mimetics (i.e., venetoclax) led to promising preclinical and clinical activity in B cell malignancies. However, several mechanisms of intrinsic or acquired resistance have been described ex vivo therefore predictive markers of response as well as mechanism-based combinations have to be designed. In the present study, we analyzed the expression of the BCL2-family genes across 10 mature B cell malignancies through computational normalization of 21 publicly available Affimetrix datasets gathering 1,219 patient samples. To better understand the deregulation of anti- and pro-apoptotic members of the BCL2-family in hematological disorders, we first compared gene expression profiles of malignant B cells to their relative normal control (naïve B cell to plasma cells, n = 37). We further assessed BCL2-family expression according to tissue localization i.e., peripheral blood, bone marrow, and lymph node, molecular subgroups or disease status i.e., indolent to aggressive. Across all cancer types, we showed that anti-apoptotic genes are upregulated while pro-apoptotic genes are downregulated when compared to normal counterpart cells. Of interest, our analysis highlighted that, independently of the nature of malignant B cells, the pro-apoptotic BH3-only BCL2L11 and PMAIP1 are deeply repressed in tumor niches, suggesting a central role of the microenvironment in their regulation. In addition, we showed selective modulations across molecular subgroups and showed that the BCL2-family expression profile was related to tumor aggressiveness. Finally, by integrating recent data on venetoclax-monotherapy clinical activity with the expression of BCL2-family members involved in the venetoclax response, we determined that the ratio (BCL2+BCL2L11+BAX)/BCL2L1 was the strongest predictor of venetoclax response for mature B cell malignancies in vivo.

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All-optical regulation of gene expression in targeted cells.

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All-optical regulation of gene expression in targeted cells.

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