Abstract LB-118: ChIP-seq based identification of genes regulated by a pancreatic β-cell differentiation factor, Hlxb9, that is dysregulated in β-cell tumors.

The homeobox transcription factor Hlxb9 controls pancreatic β-cell differentiation during the early and late stages of endocrine pancreas development. Recently, increased expression of Hlxb9 was observed in β-cell tumors associated with multiple endocrine neoplasia type 1 (MEN1), a tumor syndrome caused by germline mutation in the MEN1 tumor suppressor gene encoding menin. In MIN6 cells, a mouse pancreatic β-cell line, overexpression of Hlxb9 has been shown to cause apoptosis in a menin-dependent manner. Although Hlxb9 is known to act as a transcriptional repressor, its DNA-binding site at its target genes in pancreatic β-cells has not been elucidated. To identify targets of Hlxb9 and genomic binding sites in β-cells (MIN6 cells) we used chromatin immunoprecipitation and deep sequencing (ChIP-seq) followed by bioinformatics analysis of the ChIP-seq data. The Hlxb9 specific ChIP-seq libraries and corresponding input controls were sequenced on the Illumina Hi Seq 2000. The ChIP-seq reads were subjected to peak calling tools, and upon stringent filtering 2569 peaks were obtained. After assigning chromosomal positions to the peaks, 24 statistically significant (FDR-P de novo using the Genomatix Genome Analyzer suite that appeared to be common in most of the targets (Re-value: 1.88). Further analysis revealed that this motif was similar to binding sequences of the following proteins: Tcfe2a (involved in brain development) , Ascl2 (involved in nervous system) , REI1 (involved in mitotic signal network) , ZFp691 (zinc finger protein) , myf (involved in mitosis), and Bcl6b (involved in apoptosis). Gene Ontology analysis of the identified targets showed that 18.3% were associated with the Wnt signaling pathway, 27.3% with cell proliferation, 54.5% were involved in cellular response to stimulus, 13.6% in MAPK cascade and 31.8% in negative regulation of biological processes. Our genome-wide analysis of Hlxb9 target genes demonstrates the presence of a potential Hlxb9-binding site in target genes associated with important processes relevant for cellular differentiation and neoplasia. Further studies investigating these targets for menin-dependent and menin-independent actions of Hlxb9 will provide insights into the role of menin-Hlxb9 interaction in normal physiology and in the development of pancreatic β-cell tumors. Citation Format: Shruti S. Desai, Sita D. Modali, Weiping Chen, Vaishali I. Parekh, Sunita K. Agarwal. ChIP-seq based identification of genes regulated by a pancreatic β-cell differentiation factor, Hlxb9, that is dysregulated in β-cell tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-118. doi:10.1158/1538-7445.AM2013-LB-118