Mechanisms of Gastrointestinal Growth and Transformation

University Of Michigan At Ann Arbor, Ann Arbor MI

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Abstract

DESCRIPTION (provided by applicant): The intestinal enterochromaffin cell (EC) compartment expands in response to acute injury, microbial infection and colitis to produce serotonin. Elevated levels of plasma serotonin stimulate fluid secretion and gut motility to expel the infectious or toxic irritants. Although an essential component of the innate immune response, these gut functions contribute to patient discomfort and if sustained become a source of morbidity and mortality. Despite their central role, it is not understood what modulates EC cell numbers. During the prior funding cycle, we found that the zinc finger transcription factor ZBP-89 interacts directly with the tumor suppressor protein ataxia telangiectasia mutated (ATM) in response to histone deacetylase inhibition (HDACi), e.g., butyrate or trichostatin A (TSA). Butyrate induces ZBP-89 expression and binding to a GC-rich DNA element within the promoter of the cyclin-dependent kinase inhibitor (CDKI) p21. Butyrate triggers auto- phosphorylation of ATM at serine 1981 (pATMS1981) that subsequently complexes with ZBP-89 to mediate the components of the DNA Damage/Repair (DDR) response. In the current proposal, pATMS1981 positive expression in the gut was found to occur exclusively in EC cells of the intestine and colon demonstrating that the ZBP-89/pATMS1981 complex, which activates p21-mediated growth arrest and apoptosis, is essential to lineage specification of these cells. Mice conditionally null for ZBP-89 in the colon exhibited reduced numbers of EC cells. In addition, pATMS1981-expressing cells in APCmin polyps were absent suggesting that excess Wnt signaling prevents EC cell differentiation. In the current application, three aims are proposed to test the overarching hypothesis that ZBP-89 modulates EC cell specification that subsequently is perturbed during transformation. In Aim 1, we will determine how a component of the Wnt signaling pathway antagonizes ZBP- 89 function through a protein-protein interaction. In Aim 2, we will determine how suppression of Notch signaling in the intestine affects ZBP-89, its interaction with pATMS1981 and subsequently EC cell differentiation. In Aim 3, the numbers of EC cells in the colons of germ-free mice will be compared to mice maintained under conventional housing or treated with butyrate. Since Wnt signaling reduced epithelial ATMS1981 expression in vivo, we hypothesized that inhibiting the formation of the ATM/ZBP-89 complex and expected EC cell differentiation is an early step towards colonic transformation. Dextran sulfate (DSS) and azoxymethane (AOM) will be used to activate Wnt signaling in mice conditionally null for ZBP-89 in the colon. Anticipating reduced EC cells in the colons of mice conditionally null for ZBP-89, we predict that these mice placed on a genetic background with mutated or deleted APC alleles will be more susceptible to neoplastic transformation. Collectively, we will establish a link between the ZBP-89-pATMS1981 complex, which mediates the DDR signaling network and EC cell differentiation.