understanding-early-organogenesis-using-simplified-situ-hybridization

Understanding Early Organogenesis Using a Simplified <em>In Situ</em> Hybridization Protocol in <em>Xenopus</em>

Organogenesis is the study of how organs are specified and then acquire their specific shape and functions during development.  The Xenopuslaevis embryo ...

University of Western Ontario

The lung and thyroid are derived from the anterior endoderm. Retinoic acid and Fgf signalling are known to be essential for development of the lung in mouse but little is known on how the lung and thyroid are specified in Xenopus.

Retinoic acid is a key regulatory switch determining the difference between lung and thyroid fates in Xenopus laevis.

RanBPM is a ubiquitous protein that has been reported to regulate several cellular processes through interactions with various proteins. However, it is not known whether RanBPM may regulate gene expression patterns. As it has been shown that RanBPM interacts with a number of transcription factors, we hypothesized that it may have wide ranging effects on gene expression that may explain its function. To test this hypothesis, we generated stable RanBPM shRNA cell lines to analyze the effect of RanBPM on global gene expression. Microarray analyses were conducted comparing the gene expression profile of Hela and HCT116 RanBPM shRNA cells versus control shRNA cells. We identified 167 annotated genes significantly up- or down-regulated in the two cell lines. Analysis of the gene set revealed that down-regulation of RanBPM led to gene expression changes that affect regulation of cell, tissue, and organ development and morphology, as well as biological processes implicated in tumorigenesis. Analysis of Transcription Factor Binding Sites (TFBS) present in the gene set identified several significantly over-represented transcription factors of the Forkhead, HMG, and Homeodomain families of transcription factors, which have previously been demonstrated as having important roles in development and tumorigenesis. In addition, the combined results of these analyses suggested that several signaling pathways were affected by RanBPM down-regulation, including ERK1/2, Wnt, Notch, and PI3K/Akt pathways. Lastly, analysis of selected target genes by quantitative RT-qPCR confirmed the changes revealed by microarray. Several of the genes up-regulated in RanBPM shRNA cells encode proteins with known oncogenic functions, such as the RON tyrosine kinase, the adhesion molecule L1CAM, and transcription factor ELF3/ESE-1, suggesting that RanBPM functions as a tumor suppressor to prevent deregulated expression of these genes. Altogether, these results suggest that RanBPM does indeed function to regulate many genomic events that regulate embryonic, tissue, and cellular development as well as those involved in cancer development and progression.

Jean H. Wang

Children's Health Research Institute,
Department of Paediatrics

University of Western Ontario

Understanding Early Organogenesis Using a Simplified In Situ Hybridization Protocol in Xenopus

Jean H. Wang

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Children's Health Research Institute,Department of Paediatrics

University of Western Ontario

Understanding Early Organogenesis Using a Simplified In Situ Hybridization Protocol in Xenopus

Children's Health Research Institute,
Department of Paediatrics