Abstract:Notch signaling largely determines intestinal epithelial cell fate. High Notch activity drives progenitors toward absorptive enterocytes by repressing secretory differentiation programs, whereas low Notch permits secretory cell assignment. Myeloid translocation gene-related 1 (MTGR1) is a transcriptional corepressor in the myeloid translocation gene/Eight-Twenty-One family. Given that Mtgr1(-/-) mice have a dramatic reduction of intestinal epithelial secretory cells, we hypothesized that MTGR1 is a key repress… Show more
“…We defined the ATOH1 targetome as the 658 genes in the colon and 193 genes in the ileum with significantly enriched expression in ATOH1-positive cells that also were bound by ATOH1 (Figure 5 A , Supplementary Table 9). Consistent with the concept that ATOH1 functions as a key transcription factor for differentiation of the intestinal epithelium, several ATOH1 target genes were known to be involved in intestinal secretory lineage differentiation and function, such as Notch ligands Dll1 and Dll4 ; transcription factors Spdef ,17, 18
Sox9 ,39, 40
Gfi1 ,19, 41 and Creb3l1 42 ; transcription co-repressors Cbfa2t2 and Cbfa2t3 43, 44, 45, 46; and secretory lineage-specific genes such as Best2 , Spink4 , Muc2 , Sct , EphB3 , Xbp1 , and Clca3 17, 47, 48, 49. To gain broader insight into ATOH1 target genes in the intestines, we performed GO analysis using DAVID.…”
Background & AimsThe transcription factor atonal homolog 1 (ATOH1) controls the fate of intestinal progenitors downstream of the Notch signaling pathway. Intestinal progenitors that escape Notch activation express high levels of ATOH1 and commit to a secretory lineage fate, implicating ATOH1 as a gatekeeper for differentiation of intestinal epithelial cells. Although some transcription factors downstream of ATOH1, such as SPDEF, have been identified to specify differentiation and maturation of specific cell types, the bona fide transcriptional targets of ATOH1 still largely are unknown. Here, we aimed to identify ATOH1 targets and to identify transcription factors that are likely to co-regulate gene expression with ATOH1.MethodsWe used a combination of chromatin immunoprecipitation and messenger RNA–based high-throughput sequencing (ChIP-seq and RNA-seq), together with cell sorting and transgenic mice, to identify direct targets of ATOH1, and establish the epistatic relationship between ATOH1 and SPDEF.ResultsBy using unbiased genome-wide approaches, we identified more than 700 genes as ATOH1 transcriptional targets in adult small intestine and colon. Ontology analysis indicated that ATOH1 directly regulates genes involved in specification and function of secretory cells. De novo motif analysis of ATOH1 targets identified SPDEF as a putative transcriptional co-regulator of ATOH1. Functional epistasis experiments in transgenic mice show that SPDEF amplifies ATOH1-dependent transcription but cannot independently initiate transcription of ATOH1 target genes.ConclusionsThis study unveils the direct targets of ATOH1 in the adult intestines and illuminates the transcriptional events that initiate the specification and function of intestinal secretory lineages.
“…We defined the ATOH1 targetome as the 658 genes in the colon and 193 genes in the ileum with significantly enriched expression in ATOH1-positive cells that also were bound by ATOH1 (Figure 5 A , Supplementary Table 9). Consistent with the concept that ATOH1 functions as a key transcription factor for differentiation of the intestinal epithelium, several ATOH1 target genes were known to be involved in intestinal secretory lineage differentiation and function, such as Notch ligands Dll1 and Dll4 ; transcription factors Spdef ,17, 18
Sox9 ,39, 40
Gfi1 ,19, 41 and Creb3l1 42 ; transcription co-repressors Cbfa2t2 and Cbfa2t3 43, 44, 45, 46; and secretory lineage-specific genes such as Best2 , Spink4 , Muc2 , Sct , EphB3 , Xbp1 , and Clca3 17, 47, 48, 49. To gain broader insight into ATOH1 target genes in the intestines, we performed GO analysis using DAVID.…”
Background & AimsThe transcription factor atonal homolog 1 (ATOH1) controls the fate of intestinal progenitors downstream of the Notch signaling pathway. Intestinal progenitors that escape Notch activation express high levels of ATOH1 and commit to a secretory lineage fate, implicating ATOH1 as a gatekeeper for differentiation of intestinal epithelial cells. Although some transcription factors downstream of ATOH1, such as SPDEF, have been identified to specify differentiation and maturation of specific cell types, the bona fide transcriptional targets of ATOH1 still largely are unknown. Here, we aimed to identify ATOH1 targets and to identify transcription factors that are likely to co-regulate gene expression with ATOH1.MethodsWe used a combination of chromatin immunoprecipitation and messenger RNA–based high-throughput sequencing (ChIP-seq and RNA-seq), together with cell sorting and transgenic mice, to identify direct targets of ATOH1, and establish the epistatic relationship between ATOH1 and SPDEF.ResultsBy using unbiased genome-wide approaches, we identified more than 700 genes as ATOH1 transcriptional targets in adult small intestine and colon. Ontology analysis indicated that ATOH1 directly regulates genes involved in specification and function of secretory cells. De novo motif analysis of ATOH1 targets identified SPDEF as a putative transcriptional co-regulator of ATOH1. Functional epistasis experiments in transgenic mice show that SPDEF amplifies ATOH1-dependent transcription but cannot independently initiate transcription of ATOH1 target genes.ConclusionsThis study unveils the direct targets of ATOH1 in the adult intestines and illuminates the transcriptional events that initiate the specification and function of intestinal secretory lineages.
“…Furthermore, both Mtgr1 -/-and Mtg16 -/-mice have increased enterocyte proliferation with an expansion of transit-amplifying cell populations and skewed lineage allocation, indicative of disrupted intestinal homeostasis (12,15,16). When colitis was induced with dextran sodium sulfate (DSS), both Mtgr1 -/-and Mtg16 -/-mice were more sensitive to injury in comparison with wild-type (WT) mice, demonstrating that MTGs are required for maintaining intestinal integrity when the epithelium is challenged (10,13).…”
“…As the ChIP-seq data also suggested preferential Notch regulation by MTGR1, and because we previously determined that the secretory lineage deficiency observed in the Mtgr1 −/− intestine was rescued by Notch inhibition 24 , we next determined if Notch signaling was perturbed in Mtgr1 −/− tumors. As active Notch signaling will increase absorptive enterocyte production at the expense of secretory lineages, we stained tumors with periodic acid Schiff (PAS) to identify goblet cells.…”
Section: Resultsmentioning
confidence: 99%
“…Mtg8 −/− mice demonstrate a severe mid-gut deletion phenotype 22 ; Mtgr1 −/− mice have pan-secretory cell loss and are smaller than wild-type mice 17 ; and Mtg16 −/− mice have decreased goblet cell indices 23 . The mechanisms underlying these various phenotypes remain unclear, although there are reports that MTGR1 directly binds to TCF4, competing for β-catenin occupancy and that it interacts with CSL in repressing Notch signaling 24,35 . However, evidence that MTGs occupy Wnt/Notch targets in the gut has been lacking.…”
Section: Discussionmentioning
confidence: 99%
“…qPCR was then performed using SYBR Green (Biorad) with primers for Gapdh (Realtimeprimers). Primers for CgA and Muc2 were previously described 24 . Primers for Ascl2 , Axin2 , c-Myc , and Klf4 were purchased from RealtimePrimers.…”
Myeloid translocation genes (MTGs), originally identified as chromosomal translocations in acute myelogenous leukemia, are transcriptional corepressors that regulate hematopoietic stem cell programs. Analysis of The Cancer Genome Atlas (TCGA) database revealed that MTGs were mutated in epithelial malignancy and suggested that loss of function might promote tumorigenesis. Genetic deletion of MTGR1 and MTG16 in the mouse has revealed unexpected and unique roles within the intestinal epithelium. Mtgr1−/− mice have progressive depletion of all intestinal secretory cells, and Mtg16−/− mice have a decrease in goblet cells. Furthermore, both Mtgr1−/− and Mtg16−/− mice have increased intestinal epithelial cell proliferation. We thus hypothesized that loss of MTGR1 or MTG16 would modify Apc1638/+-dependent intestinal tumorigenesis. Mtgr1−/− mice, but not Mtg16−/− mice, had a 10-fold increase in tumor multiplicity. This was associated with more advanced dysplasia, including progression to invasive adenocarcinoma, and augmented intratumoral proliferation. Analysis of ChIP-seq datasets for MTGR1 and MTG16 targets indicated that MTGR1 can regulate Wnt and Notch signaling. In support of this, immunohistochemistry and gene expression analysis revealed that both Wnt and Notch signaling pathways were hyperactive in Mtgr1−/− tumors. Furthermore, in human colorectal cancer (CRC) samples MTGR1 was downregulated at both the transcript and protein level. Overall our data indicates that MTGR1 has a context dependent effect on intestinal tumorigenesis.
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