Abstract
The gastrointestinal (GI) epithelium is constantly renewing, depending upon the intestinal stem cells (ISC) regulated by a spectrum of transcription factors (TFs), including Myb. We noted previously in mice with a p300 mutation (plt6) within the Myb-interaction-domain phenocopied Myb hypomorphic mutant mice with regard to thrombopoiesis, and here, changes in GI homeostasis. p300 is a transcriptional coactivator for many TFs, most prominently cyclic-AMP response element-binding protein (CREB), and also Myb. Studies have highlighted the importance of CREB in proliferation and radiosensitivity, but not in the GI. This prompted us to directly investigate the p300–Myb–CREB axis in the GI. Here, the role of CREB has been defined by generating GI-specific inducible creb knockout (KO) mice. KO mice show efficient and specific deletion of CREB, with no evident compensation by CREM and ATF1. Despite complete KO, only modest effects on proliferation, radiosensitivity and differentiation in the GI under homeostatic or stress conditions were evident, even though CREB target gene pcna (proliferating cell nuclear antigen) was downregulated. creb and p300 mutant lines show increased goblet cells, whereas a reduction in enteroendocrine cells was apparent only in the p300 line, further resembling the Myb hypomorphs. When propagated in vitro, crebKO ISC were defective in organoid formation, suggesting that the GI stroma compensates for CREB loss in vivo, unlike in MybKO studies. Thus, it appears that p300 regulates GI differentiation primarily through Myb, rather than CREB. Finally, active pCREB is elevated in colorectal cancer (CRC) cells and adenomas, and is required for the expression of drug transporter, MRP2, associated with resistance to Oxaliplatin as well as several chromatin cohesion protein that are relevant to CRC therapy. These data raise the prospect that CREB may have a role in GI malignancy as it does in other cancer types, but unlike Myb, is not critical for GI homeostasis.
Highlights
Several pathways involved in the homeostasis of gut have been identified
A growing collective of transcription factor (TF) are involved in achieving intestinal crypt homeostasis and previously we have shown a role of Myb,[20] exploiting several Myb hypomorphic mutant mice,[18,19] sustained survival of which allowed the analysis of adult tissues not possible with embryonically lethal MybKO mice[22] and that complemented the inducible intestinalspecific MybKO studies.[20]
Our discovery that the p300plt6/plt[6] mutant in part phenocopied the Myb hypomorphs[17] and led to a substantial reduction in Myb expression highlights the utility of these mutants
Summary
P300plt6/plt[6] intestines phenocopy Myb hypomorphs. As part of a comprehensive ENU mutagenesis screen focused on thrombopoiesis[18] or hematopoiesis,[19] Myb emerged as a central factor in these processes.[21]. As the anticipated gene expression effects that might be expected in crebKO tissue were not observed, we isolated SI crypts for gene expression studies (these were depleted of the villi) from three creb/villinCreERT2 and three crebfl/flcontrol mice where each cohort had been subjected to 4-week Tamoxifen treatment This was important for both groups, as Tamoxifen influences transcription.[41,42] This strategy was considered to most closely parallel that used to characterize the GI morphology, to examine the GI cells most likely to express pCREB and to capture unanticipated gene expression changes. The crebKO organoids grew less efficiently than the WT organoids, beyond this baseline difference, colony
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
