Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Expression Profile of Chicken Primitive Streak (HH4) Tissues

ABSTRACT: In birds and mammals, all mesoderm cells are generated from the primitive streak. Nascent mesoderm cells contain unique dorso-ventral (D/V) identities depending on their relative ingression position along the streak. Molecular mechanisms controlling this initial phase of mesoderm diversification are not well-understood. Using chick model, we generated high-quality transcriptomic datasets of different streak regions and analyzed their molecular heterogeneity. Streak tissues were dissected from stage-matched HH4 chick embryos and were further divided into four equal pieces along its anterio-posterior length (termed “A”, “B”, “C” and “D”, with “A” representing most anterior and therefore most dorsal and “D” most posterior and therefore most ventral). Each of the four regions was represented by three independent samples (n=3), with each sample containing 5-7 μg total RNA derived from 85-93 pooled pieces. 5 μg of RNA from each sample were used to screen Affymetrix Chicken Genome Array without an amplification step.

ORGANISM(S): Gallus gallus

SUBMITTER: Guojun Sheng

PROVIDER: E-GEOD-22230 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Transcriptomic landscape of the primitive streak.

Alev Cantas C Wu Yuping Y Kasukawa Takeya T Jakt Lars M LM Ueda Hiroki R HR Sheng Guojun G

Development (Cambridge, England) 20100728 17

In birds and mammals, all mesoderm cells are generated from the primitive streak. Nascent mesoderm cells contain unique dorsoventral (D/V) identities according to their relative ingression position along the streak. Molecular mechanisms controlling this initial phase of mesoderm diversification are not well understood. Using the chick model, we generated high-quality transcriptomic datasets of different streak regions and analyzed their molecular heterogeneity. Fifteen percent of expressed genes ...[more]

PMID: 20667916

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Project description:Genetic evidence has implicated several genes as being critical for heart development. However, the inducers of these genes as well as their other targets and the pathways they constitute, remain largely unknown. In the avian embryo, Cerberus (cCer) transcripts are detected in the anterior mesendoderm including the heart precursor cells and in the left lateral plate mesoderm. We have identified a promoter element of chick cCer able to drive EGFP expression into the heart and the hemangioblast precursor cells, allowing us to identify a population of cells that consistently exit from the anterior primitive streak region from as early as stage HH3+ and that later will populate the heart. In order to identify and study novel genes expressed and involved in the correct development and differentiation of the vertebrate H/HPC (Heart/ Hemangioblast Precursor Cells) lineages, a differential screening using Affymetrix GeneChip system technologies was performed. Remarkably, this screening led to the identification of more than 800 transcripts potentially expressed in these precursor lineages. We have identified unknown genes that are differentially expressed in the H/HPC precursor lineages. By developing a procedure to isolate the heart precursor cells using the cer2.5-EGFP construct, we were able to specifically isolate a population of H/HPC expressing already known cardiac markers, and a long list of still uncharacterized genes. From those, we defined by WISH that at least some of these in silico identified transcripts are in fact expressed in these cell populations and more importantly, functionally required for heart formation.More importantly, our study unveiled several uncharacterized genes that can now be used for further studies. Portions of chick embryos were selected and excised at early stages of development for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain homogeneous populations of embryos at early gastrula stage in order to increase the temporal resolution of expression profiles. To that end, we hand-selected and excised portions of the embryos according to morphological criteria and EGFP expression. Four GeneChip chicken arrays used for another study were added to the two arrays in this study to ensure a robust model for expression value computation (Note: The 4 samples chick_T2-1, chick_T2-2, chick_T2-3, chick_T2-4, were performed by another group and for a different array, and nothing has to do with our data. They were only run at the same time and used for normalization. Raw data unavailable.). The arrays were normalized to a baseline array with median CEL intensity by applying an Invariant Set Normalization Method. Normalized CEL intensities of the six arrays were used to obtain model-based gene expression indices (MBEI) based on a PM (Perfect Match)-only model. Processed data for all 6 samples can be found in supplementary file linked below.

Project description:Within a given vertebrate species, the total number of vertebrae in each anatomical domain is precisely defined and shows little variation among individuals. In contrast, this number can vary tremendously between different species, ranging from as few as six vertebrae in frogs to as many as several hundred in some snakes and fish. Segmental precursors of the vertebrae, called somites are produced sequentially in the embryo from the presomitic mesoderm (PSM), until a final number characteristic of the species, is reached. Here, we show in the chicken embryo that, by controlling the rate of axis elongation, Hox genes control the total number of somites generated by the embryo. We observed that activation of the most posterior Hox genes in somite precursors of the tail bud correlates with an abrupt slowing-down of the speed of axis elongation. We show that progressively more posterior Hox genes, which are collinearly activated in somitic precursors of the epiblast, repress Wnt activity with increasing strength. This leads to a graded repression of the Brachyury/T transcription factor, reducing mesoderm ingression and slowing down the elongation process. Due to the continuation of somite formation, the PSM, which is not fed with sufficient supply of new cells posteriorly, becomes progressively exhausted, ultimately leading to an arrest of segment formation. Our data provide a conceptual framework to explain how the cross-talk between the segmentation clock and the Hox clock accounts for the diversity of vertebral formulae across animal species. Primitive streak aera corresponding to PSM precursors were dissected in Stage 9 somites chicken embryo overexpressing HoxA13 or a control H2B-Venus. The experiment was designed to have biological duplicate in each conditions. The gain-of-function was obtained by electroporating the embryo at Stage 5HH with a vector containing HoxA13 under the CAGGS promoter with an H2B venus reporter. In order to have enough material for the microarray, 7 embryos were pooled in each sample before the hybridization.

Dataset Information

Expression Profile of Chicken Primitive Streak (HH4) Tissues

Publications

Transcriptomic landscape of the primitive streak.

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