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  • Predominant Distribution of the RNAi Machinery at Apical Adherens Junctions in Colonic Epithelia Is Disrupted in Cancer.

Predominant Distribution of the RNAi Machinery at Apical Adherens Junctions in Colonic Epithelia Is Disrupted in Cancer.

International journal of molecular sciences (2020-04-11)
Joyce Nair-Menon, Amanda C Daulagala, Dean M Connor, Lauren Rutledge, Trevor Penix, Mary Catherine Bridges, Bridgette Wellslager, Demetri D Spyropoulos, Cynthia D Timmers, Ann-Marie Broome, Antonis Kourtidis
ABSTRACT

The RNA interference (RNAi) machinery is an essential component of the cell, regulating miRNA biogenesis and function. RNAi complexes were thought to localize either in the nucleus, such as the microprocessor, or in the cytoplasm, such as the RNA-induced silencing complex (RISC). We recently revealed that the core microprocessor components DROSHA and DGCR8, as well as the main components of RISC, including Ago2, also associate with the apical adherens junctions of well-differentiated cultured epithelial cells. Here, we demonstrate that the localization of the core RNAi components is specific and predominant at apical areas of cell-cell contact of human normal colon epithelial tissues and normal primary colon epithelial cells. Importantly, the apical junctional localization of RNAi proteins is disrupted or lost in human colon tumors and in poorly differentiated colon cancer cell lines, correlating with the dysregulation of the adherens junction component PLEKHA7. We show that the restoration of PLEKHA7 expression at adherens junctions of aggressively tumorigenic colon cancer cells restores the junctional localization of RNAi components and suppresses cancer cell growth in vitro and in vivo. In summary, this work identifies the apical junctional localization of the RNAi machinery as a key feature of the differentiated colonic epithelium, with a putative tumor suppressing function.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-Actin antibody produced in rabbit, affinity isolated antibody, buffered aqueous solution
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4′,6-Diamidino-2-phenylindole dihydrochloride, powder, BioReagent, suitable for cell culture, ≥98% (HPLC and TLC), suitable for fluorescence
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Anti-PLEKHA7 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution
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Anti-p120 Catenin Antibody, clone 15D2, clone 15D2, from mouse
SAFC
McCoy′s 5A Medium, Modified, with L-glutamine, without sodium bicarbonate, powder, suitable for cell culture
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PP2, ≥98% (HPLC)