Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptome Analysis of Developing Intestine [RNA-Seq]

ABSTRACT: Background: The muscularis externa (ME) of the adult intestine consists of two layers of visceral smooth muscle (VISM), the inner circular muscle (ICM) and outer longitudinal muscle (OLM), that form sequentially beginning at embryonic day (E) 13 and E15 in the developing mouse. Coordinated contraction of these two layers facilitates the movement of food down the digestive tract. Though abnormal ME function or development has been linked to pseudoobstruction and irritable bowel syndrome, little is known about the molecular character of the smooth muscle that comprises this tissue. We performed transcriptome analysis to identify genes that are enriched in intestinal mesenchyme tissue at E14.5, when the inner circular muscle (ICM) is well established. Results: Expression patterns of enriched mesenchyme genes were examined in publically available in situ databases, revealing over one hundred genes that are expressed in the ICM. Examination of the promoter regions for these genes revealed enrichment for cJUN transcription factor binding sites and cJUN itself was also enriched in ICM. A cJUN ChIP-seq at E14.5 showed that cJUN regulatory regions contained characteristics of muscle enhancers. E14.5 mouse intestines were harvested and grown for 24 hours in a transwell culture with or without Cyclopamine treatment. Separated epithelial and mesenchyme tissue populations or whole intestines were submitted for sequencing. Three replicates for each condition were collected.

ORGANISM(S): Mus musculus

SUBMITTER: Katherine Gurdziel

PROVIDER: E-GEOD-74920 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Diverse functions of the homeodomain transcription factor BARX1 include Wnt-dependent, non-cell autonomous specification of the stomach epithelium, tracheo-bronchial septation, and Wnt-independent expansion of the spleen primordium. Tight spatio-temporal regulation of Barx1 levels in the mesentery and stomach mesenchyme suggests additional roles. To determine these functions, we forced constitutive BARX1 expression in the Bapx1 expression domain, which includes the mesentery and intestinal mesenchyme, and also examined Barx1-/- embryos in further detail. Transgenic embryos invariably showed intestinal truncation and malrotation, in part reflecting abnormal left-right patterning. Ectopic BARX1 expression did not affect intestinal epithelium, but intestinal smooth muscle developed with features typical of the stomach wall. BARX1, which is normally restricted to the developing stomach, drives robust smooth muscle expansion in this organ by promoting proliferation of myogenic progenitors at the expense of other sub-epithelial cells. Undifferentiated embryonic stomach and intestinal mesenchyme showed modest differences in mRNA expression and BARX1 was sufficient to induce much of the stomach profile in intestinal cells. However, limited binding at cis-regulatory sites implies that BARX1 may act principally through other transcription factors. Genes expressed ectopically in BARX1+ intestinal mesenchyme and reduced in Barx1-/- stomach mesenchyme include Isl1, Pitx1, Six2 and Pitx2, transcription factors known to control left-right patterning and influence smooth muscle development. The sum of evidence suggests that potent BARX1 functions in intestinal rotation and stomach myogenesis occur through this small group of intermediary transcription factors. To investigate how Barx1 regulates gut smooth muscle development in a cell-autonomous manner, we used Affymetrix arrays to profile genes enriched in wild-type stomach and BARX1-overexpressing intestinal mesenchyme, compared to wild-type intestinal mesenchyme.

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Transcriptome Analysis of Developing Intestine [RNA-Seq]

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