A Qualitative Change in the Transcriptome Occurs after the First Cell Cycle and Coincides with Lumen Establishment during MDCKII Cystogenesis

Abstract

Summary Madin-Darby canine kidney II (MDCKII) cells are widely used to study epithelial morphogenesis. To better understand this process, we performed time course RNA-seq analysis of MDCKII 3D cystogenesis, along with polarized 2D cells for comparison. Our study reveals a biphasic change in the transcriptome that occurs after the first cell cycle and coincides with lumen establishment. This change appears to be linked to translocation of β-catenin, supported by analyses with AVL9 - and DENND5A -knockdown clones, and regulation by HNF1B, supported by ATAC-seq study. These findings indicate a qualitative change model for transcriptome remodeling during epithelial morphogenesis, leading to cell proliferation decrease and cell polarity establishment. Furthermore, our study reveals that active mitochondria are retained and chromatin accessibility decreases in 3D cysts but not in 2D polarized cells. This indicates that 3D culture is a better model than 2D culture for studying epithelial morphogenesis.

Canine tumor mutational burden is correlated with TP53 mutation across tumor types and breeds

“…Mammary adenomas or carcinomas originate from epithelial cells. The establishment of epithelial cell apical-basolateral polarity decreases cell proliferation and invasiveness, acting as a potent tumor suppressor [36][37][38] . PIK3CA H1047R mutation, common in canine mammary tumors, increases cell stemness 39 and decreases epithelial cell polarity, leading to accelerated cell proliferation and tumorigenesis.…”

Shared hotspot mutations in spontaneously arising cancers position dog as an unparalleled comparative model for precision therapeutics

“…This may be due to differences in the cell of origin and the mechanisms of tumorigenesis in these cancer types. Carcinomas originate from polarized epithelial cells or their progenitors, and alterations of cell polarity genes and loss of cell polarity are likely the major drivers of accelerated cell proliferation in carcinoma development 63–65 . Sarcomas originate from mesenchymal cells, and loss of function of TP53 , which leads to defective cell cycle checkpoints and accelerated proliferation.…”

Drivers of Transcriptional Variance in Human Intestinal Epithelial Organoids

“…To further understand these gene expression patterns, we used GSEA analysis to identify pathways enriched in distal intestinal organoids compared to proximal organoids. The results for intersegmental comparisons within specific culture formats revealed novel enriched terms as well as those that confirm known functional differences between segments (LaPointe et al, 2008; Mach et al, 2014; Uhlen et al, 2010; Yalong Wang et al, 2020). It is well known that bile acids are released into the duodenum and are reabsorbed by the enterocytes in the ileum and recirculated back to the liver via enterohepatic circulation.…”

“…The format and substrate that the organoids were grown on were the two largest drivers of variance in our dataset. The reason why format is a large driver of variance remains unclear as few investigators have compared organoids grown as 3D, 2D Monolayers and/or Transwells (Guo et al, 2008; Kasendra et al, 2018; Öhlund et al, 2017; T. Wang et al, 2020; Zhang et al, 2017). The differences that we observed between 3D and 2D (monolayers and transwells) formats could be potentially due to the different cell shapes in sheet and spherical conformations, which may alter attachment to the Matrigel substrate.…”