Abstract LB-143: The role of Thoc1 in cell differentiation of the small intestine

Abstract

Abstract Thoc1 encodes an RNA binding protein (pThoc1) that was identified due to its interaction with the retinoblastoma tumor suppressor protein (Rb). pThoc1 regulates transcriptional elongation and mRNA export as part of the TREX/THO (Transcription/Export) complex. In vivo, mice lacking Thoc1 fail to develop past the late blastocyst stage and mice hypomorphic for Thoc1 have compromised gametogenesis and are sterile. Elevated levels of pThoc1 occur in multiple cancers (breast, lung, colorectal), in cancer cell lines, and in transformed fibroblasts. In vitro, neoplastic cells appear to be more sensitive to pThoc1 loss than normal cells. These observations lead to the hypothesis that cells with extended replicative potential (i.e. stem, progenitor, & cancer cells) require Thoc1. To test this hypothesis, we are examining the role of Thoc1 in vivo utilizing the crypt-villus axis of the murine small intestine as a model system. We have deleted Thoc1 in the murine small intestine using the Rosa26CreER transgene and the Tamoxifen induced CreER mediated recombination system. Thoc1F/F:Rosa26CreER (test) and Thoc1+/+:Rosa26CreER (control) 22 week old mice were treated with intraperitoneal injections of 1mg/day Tamoxifen for 5 days. Small intestinal tissue was collected 24 hours after the last injection. Histological analysis shows Thoc1 deletion disrupts small intestinal crypt architecture, but does not affect villus architecture. Immunohistochemistry illustrates Thoc1 deletion correlates with an increase in cleaved Caspase 3 and decrease in Ki67 in the crypts. No cleaved Caspase 3 is observed in the differentiated villi. These findings suggest intestinal stem cells (ISCs) and progenitor transit amplifying cells of the highly proliferative intestinal crypts require Thoc1 to a greater extent than the differentiated cells of the villi. Since the presence of Thoc1 appears to be required in the crypts, we investigated how Tamoxifen treatment and Thoc1 deletion would affect the crypt-villus axis over time. Thoc1F/F:Rosa26CreER and Thoc1+/+:Rosa26CreER mice were treated as previously described, however small intestinal tissue was collected 3 weeks after the last Tamoxifen injection. Interestingly, no change in body weight, survival, or crypt-villus architecture was observed. These findings lead us to believe ISCs lacking Thoc1 are at a competitive disadvantage and will eventually be replaced by Thoc1 positive ISCs. Consistent with this, while Thoc1 deletion and pThoc1 loss was readily detected 24 hours after Tamoxifen treatment, it was not three weeks after treatment. Future work involves deleting Thoc1 specifically in ISCs using the Lgr5CreER transgene to allow cell differentiation lineage tracing and cell fate studies of ISCs in which Thoc1 has been deleted. Additionally, since Thoc1 is elevated in multiple tumor types, we will delete Thoc1 in transformed ISCs using the Lgr5CreER transgene in ApcMin/+ mice to study Thoc1 in cancer stem cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-143. doi:10.1158/1538-7445.AM2011-LB-143