MTGR1 is required to maintain small intestinal stem cell populations - RNA-sequencing

Undifferentiated intestinal stem cells (ISCs), particularly those marked by Lgr5, are crucial for maintaining homeostasis and resolving injury. Lgr5+ cells in the crypt base constantly divide, pushing daughter cells upward along the crypt axis where they differentiate into specialized cell types. Coordinated execution of complex transcriptional programs are necessary to allow for the maintenance of undifferentiated stem cells while permitting differentiation of the wide array of intestinal cells necessary for homeostasis. Previously, members of the myeloid translocation gene (MTG) family have been identified as transcriptional co-repressors that regulate stem cell maintenance and differentiation programs in multiple organ systems, including the intestine. One MTG family member, myeloid translocation gene related 1 (MTGR1), has been recognized as a crucial regulator of secretory cell differentiation and response to injury. However, whether MTGR1 contributes to the function of ISCs has not yet been examined. Here, using Mtgr1-/- mice, we have assessed the effects of MTGR1 loss specifically in ISC biology. Interestingly, loss of MTGR1 increased the total number of cells expressing Lgr5, the canonical marker of cycling ISCs, suggesting higher overall stem cell numbers. However, expanded transcriptomic and functional analyses revealed deficiencies in MTGR1 null ISCs, including deregulated ISC-associated transcriptional programs. Ex vivo, intestinal organoids established from Mtgr1 null were found nearly completely unable to survive and expand, likely due to aberrant differentiation and loss of stem and proliferative cells. Together, these results indicate that MTGR1’s role in intestinal differentiation is likely to be stem cell intrinsic and identify a novel role MTGR1 in maintaining ISC function. Small intestine crypts were isolated from 3 WT and 3 Mtgr1-/- mice. Following crypt isolation, a portion of the samples were collected and homogenized in TRIzol reagent while the remaining crypts were plated for enteroid culture. After 24 hours, half of the plated enteroids were collected for RNA extraction, while the remaining enteroids were cultured for an additional 48 hours and harvested at 72 hours post-plating. Gene expression changes between WT and Mtgr1 null samples was assessed by bulk RNA-sequencing.