Abstract 5236: A qualitative change in transcriptome during MDCKII 3D epithelial morphogenesis is linked to intracellular trafficking

Abstract

Abstract The majority of human cancers are derived from epithelial tissues. Understanding epithelial morphogenesis is hence important in cancer research. Madin-Darby canine kidney II (MDCKII) cells, cultured in 3D and 2D conditions, are used extensively as a model for studying cell polarity, epithelial morphogenesis and carcinogenesis. Like other cell differentiation systems, gene expression plays a key role in MDCKII epithelial morphogenesis. To better understand the change in the transcriptome, we performed a time course RNA-seq analysis of MDCKII 3D cytogenesis, along with fully polarized 2D cells for comparison. Surprisingly, our study reveals that the change is not linear, but rather clearly biphasic. Specifically, about 3,000 genes are significantly up- or down-regulated between 24 hour and day 3 after seeding, when the lumen is forming, compared to < 120 such genes during other time intervals. Because previous studies have shown that Rab11-coordinated intracellular trafficking plays an essential role in MDCKII lumen formation, we hypothesize this qualitative change in the transcriptome is linked to intracellular trafficking. To test this hypothesis, we used knockdown clones of AVL9 and DENND5A, both interacting with Rab11 and participating in trafficking. Knockdown cells have altered cell polarity and defective trafficking, but not as significant changes in transcriptome as wild type cells, supporting our hypothesis. We then focused on β-catenin to better understand the mechanism. Our study reveals that in wild type cells, following the first cell division, β-catenin is depleted from the nucleus and enriched in cell-cell junction. This results in MYC transcriptional silencing (supported by ATAC-seq analysis), which in turn initiated down-regulation of numerous MYC target genes. These observations are however not observed in knockdown cells. Our study supports the qualitative change (“switch on-off”) model, rather than the quantitative (gradual) change model, in transcriptome remodeling during epithelial cell differentiation. Moreover, our work supports that intracellular trafficking likely initiates this quantitative change in transcriptome. Citation Format: Tianfang Wang, Shaying Zhao, Shi-Yuan Cheng. A qualitative change in transcriptome during MDCKII 3D epithelial morphogenesis is linked to intracellular trafficking [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5236.