In vivo genome-wide analysis of multiple tissues identifies gene regulatory networks, novel functions and downstream regulatory genes for Bapx1 and its co-regulation with Sox9 in the mammalian vertebral column

Abstract

BackgroundVertebrate organogenesis is a highly complex process involving sequential cascades of transcription factor activation or repression. Interestingly a single developmental control gene can occasionally be essential for the morphogenesis and differentiation of tissues and organs arising from vastly disparate embryological lineages.ResultsHere we elucidated the role of the mammalian homeobox gene Bapx1 during the embryogenesis of five distinct organs at E12.5 - vertebral column, spleen, gut, forelimb and hindlimb - using expression profiling of sorted wildtype and mutant cells combined with genome wide binding site analysis. Furthermore we analyzed the development of the vertebral column at the molecular level by combining transcriptional profiling and genome wide binding data for Bapx1 with similarly generated data sets for Sox9 to assemble a detailed gene regulatory network revealing genes previously not reported to be controlled by either of these two transcription factors.ConclusionsThe gene regulatory network appears to control cell fate decisions and morphogenesis in the vertebral column along with the prevention of premature chondrocyte differentiation thus providing a detailed molecular view of vertebral column development.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-1072) contains supplementary material, which is available to authorized users.

Genome wide binding (ChIP-Seq) of murine Bapx1 and Sox9 proteins in vivo and in vitro

“…The information presented here has been previously described in Chatterjee et al (2014) “In vivo genome-wide analysis of multiple tissues identifies gene regulatory networks, novel functions and downstream regulatory genes for Bapx1 and its co-regulation with Sox9 in the mammalian vertebral column.” [1] . An overview of the experimental approach is shown in Fig.…”

“…Chromatin (~ 2 mg) was used for chromatin immunoprecipitation as previously described for Bapx1/Nkx3.2 and Sox9 [10] . With 15 ng of purified ChIP DNA from each sample, the sequencing library was generated according to the manufacturer’s instructions (Illumina) [1] , [11] . Antibodies used for Bapx1/Nkx3.2 and Sox were anti S-Peptide antibody (Bethyl laboratories, A190-134A) and anti-Sox9 antibody (R&D Systems, AF3075) respectively.…”

Gene expression profiles of Bapx1 expressing FACS sorted cells from wildtype and Bapx1-EGFP null mouse embryos

“…The data presented here pertains to Chatterjee et al (2015) “In vivo genome-wide analysis of multiple tissues identifies gene regulatory networks, novel functions and downstream regulatory genes for Bapx1 and its co-regulation with Sox9 in the mammalian vertebral column.” [1] . We have used stable transgenic mouse lines expressing EGFP under the endogenous promoter of various transcription factors expressed in various organs in the developing embryo to sort out cells to do further genomic experiments like transcriptional profiling and genome wide binding assays ( Fig.…”

“…Peaks called by MACS were ranked as described [1] . The top 200 peaks were used for motif analysis and the repeat masked genome sequence +/− 50 bp from the summit of these 200 peaks was downloaded from the UCSC genome browser ( http://genome.ucsc.edu /).…”

Revisiting the role of the Gcm transcription factor, from master regulator to Swiss army knife

“…[1][2][3] However, pleiotropic genes should be studied in a tissue/stage specific fashion to understand the molecular mechanisms at play in each system. 71 Further characterization of Gcm using conditional knock down and MARCM clones will allow a complete understanding of Gcm's role in Drosophila development and will shed light on the real impact of this fate determinant.…”