Prediction of gene network models in limb muscle precursors

Abstract

The ventrolateral dermomyotome gives rise to all muscles of the limbs through the delamination and migration of cells into the limb buds. These cells proliferate and form myoblasts, withdraw from the cell cycle and become terminally differentiated. The myogenic lineage colonizes pre-patterned regions to form muscle anlagen as muscle fibers are assembled. The regulatory mechanisms that control the later steps of this myogenic program are not well understood. The homeodomain transcription factor Pitx2 is expressed in the muscle lineage from the migration of precursors to adult muscle. Ablation of Pitx2 results in distortion, rather than loss, of limb muscle anlagen, suggesting that its function becomes critical during the colonization of, and/or fiber assembly in, the anlagen. Gene expression arrays were used to identify changes in gene expression in flow-sorted migratory muscle precursors, labeled by Lbx1EGFP, which resulted from the loss of Pitx2. Target genes of Pitx2 were clustered using the “David Bioinformatics Functional Annotation Tool” to bin genes according to enrichment of gene ontology keywords. This provided a way to both narrow the target genes and identify potential gene families regulated by Pitx2. Representative target genes in the most enriched bins were analyzed for the presence and evolutionary conservation of Pitx2 consensus binding sequence, TAATCY, on the −20kb, intronic, and coding regions of the genes. Fifteen Pitx2 target genes were selected based on the above analysis and were identified as having functions involving cytoskeleton organization, tissue specification, and transcription factors. Data from these studies suggest that Pitx2 acts to regulate cell motility and expression of muscle specific genes in the muscle precursors during forelimb muscle development. This work provides a framework to develop the gene network leading to skeletal muscle development, growth and regeneration.

Gonadal Asymmetry and Sex Determination in Birds

“…For example, a direct effect on CD1 by PITX2 cannot be excluded as PITX2 has been shown to bind the CD1 promoter in other tissues [Baek et al, 2003]. In addition, a number of studies indicate that PITX2 controls genes involved in the reorganisation of the cytoskeleton, affecting cell spreading, migration and cell-cell-adhesion [Wei and Adelstein, 2002;Campbell et al, 2012].…”

Mapping the chromatin state dynamics in myoblasts

“…Pitx2 target genes in mouse Lbx1 + migratory muscle cells from E12.5 Lbx1 EGFP/+ /Pitx2 +/+ (WT) [27] and Lbx1 EGFP/+ /Pitx2 Z/Z (MT) mice (Campbell et al, 2012a) forelimbs, the dataset GSE31945 NCBI Gene Expression Omnibus and subsets of GSM791677, GSM791678, GSM791683, and GSM791684 were used (Table 1S). All data analysis was done in R using the Bioconductor package and its components (Gentleman et al, 2004).…”

“…WT data was obtained from our previous GSE49010 while MT data was newly generated and deposited in NCBI GEO under GSE71128 (Campbell et al, 2012). Fastq files along with their inputs were aligned to the mouse genome (mm10/NCBI38) reference assembly using Bowtie2 version 2.2.3, with default parameters (Langmead and Salzberg, 2012).…”

“…Pitx2 contributes to specification of the anatomical context that surrounds the muscle, including the jaw (Shih et al, 2007b) and the abdominal wall (Eng et al, 2012). Pitx2 regulates the relative amounts and types of cytoskeletal proteins that are produced as muscle cells assemble into muscles and contributes to the higher order muscle assembly (Campbell et al, 2012a). …”

Genomic Strategies for Terminal Cell Fate Specification

“…Downstream gene functions affected by pitx2 include migration, cell morphology, and cell adhesion, and these functions are mediated by both positive and negative (direct and indirect) inputs (Campbell et al, 2012;Eng et al, 2012).…”