| ○要約 (任意): | □ | (英) Epithelial cells have apicobasal polarity, which shows the asymmetric distributions of proteins between apical and basolateral surface within single cell that produce asymmetric cellular function to maintain homeostasis in the body. Therefore, apicobasal polarity is essential for multicellular organisms and its disturbance causes various diseases including cancer. Some apicobasal polarity factors have been identified in model animals through genetic studies. However, the comprehensive molecular system remains to be clarified. RNAi screening for apicobasal polarity using culturable epithelial cells has not been performed because of the robustness of epithelial sheets that protects cells from disturbance of the polarity factors. Here, we develop a screening system for apicobasal polarity factors using α-catenin-deficient R2/7 cells that are not capable of forming cell-cell adhesion. Intriguingly, R2/7 cells form the apicobasal polarity: aPKC, SCRIB and ZO-1 were localized to apical, basolateral and the boundary region, respectively. In particular, ZO-1 was observed as ring just below plasma membrane. Knockdown of known apical factors shrank the ZO-1 ring, whereas knockdown of known basolateral factors enlarged the ring. Therefore, the ZO-1 ring in R2/7 would be a good marker for sensitive detection of the change in apicobasal polarity. We established an algorithm for the detection and estimation of ZO-1 area that was ensured as acceptable assay (Z´-factor>0.5). Then we performed the screening using siRNA library (18,152 genes) and high-throughput image analysis. Using bioinformatic analyses, we could extract 1,263 and 422 candidate genes showing shrinking and enlarging type, respectively. Now, we are executing biochemical analyses for novel molecules to clarify comprehensive molecular system of the apicobasal polarity. (日)
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