Periodic Report Summary 2 - HINLOD (Roles of homeoproteins in lymphoid organ development)
Developmental and cancer biology can be seen as two complementary disciplines. Whereas developmental biology is concerned with mechanisms underlying the acquisition and maintenance of normal cellular identity and function, cancer biology focuses on the disruption of such mechanisms and thus on abnormal cell function. Understanding what goes wrong when a cell becomes cancerous requires knowledge of the processes that ensure correct function of the same cell type during normal development. It is now clear that many oncogenes and tumour suppressors encode for transcription factors that are crucial in orchestrating developmental processes including organogenesis. To this end we set to identify and characterise novel molecular networks regulated by the oncogenic transcription factors Pbx1 and Hox11 during organogenesis through the following specific objectives:
1) identification of the growth-related regulatory networks and target genes controlled by homeoproteins during organogenesis;
2) validation and functional characterisation of candidate targets during organ development.
Over the past two years, we performed a comprehensive DNA microarray analysis and used Gene set enrichment analysis (GSEA) for interpreting gene expression data, a computational method that allows focusing on set of genes (instead of single gene) that share common biological functions. By employing this approach, we found statistically significant differences in the expression profiles connected with several developmental pathways. In particular, we found that loss of Hox11 is associated with up-regulation of several pathways including retinoc acid, Wnt and BMP. In addition, to determine whether Hox11 and Pbx1 directly controls these genes / pathways, we performed chromatin immunoprecipitation (ChIP) analysis coupled to deep sequencing using and identified several genomic regions bound by these transcription factors. We selected a few genes belonging to the retinoic acid pathways and confirmed, by ChIP and qPCR analyses, they are direct target genes of Hox11 and Pbx1. By using an in vitro organ culture system, we functionally validated a few targets belonging to the RA pathways and are currently extending our analysis to other target genes / pathways.
1) identification of the growth-related regulatory networks and target genes controlled by homeoproteins during organogenesis;
2) validation and functional characterisation of candidate targets during organ development.
Over the past two years, we performed a comprehensive DNA microarray analysis and used Gene set enrichment analysis (GSEA) for interpreting gene expression data, a computational method that allows focusing on set of genes (instead of single gene) that share common biological functions. By employing this approach, we found statistically significant differences in the expression profiles connected with several developmental pathways. In particular, we found that loss of Hox11 is associated with up-regulation of several pathways including retinoc acid, Wnt and BMP. In addition, to determine whether Hox11 and Pbx1 directly controls these genes / pathways, we performed chromatin immunoprecipitation (ChIP) analysis coupled to deep sequencing using and identified several genomic regions bound by these transcription factors. We selected a few genes belonging to the retinoic acid pathways and confirmed, by ChIP and qPCR analyses, they are direct target genes of Hox11 and Pbx1. By using an in vitro organ culture system, we functionally validated a few targets belonging to the RA pathways and are currently extending our analysis to other target genes / pathways.