image: Representative images of chicken embryos used for RNA preparation and subsequent CAGE sequencing. Top panels show embryos in the first day of embryonic development (21 days in total), equivalent to the first five weeks of human gestation (first three weeks of human post-fertilization development). Bottom panels show embryos from the establishment of circulatory system to about a week before hatching, similar to the beginning of third trimester in human development. [Image modified from the original (Figure 1A) in Lizio, M. et. al. (2017). Systematic analysis of transcription start sites in avian development. PLOS Biology, 15(9), e2002887. doi:10.1371/journal.pbio.2002887 under the CC by 4.0 license.] view more
Credit: Professor Guojun Sheng
Early developmental research on human embryos is impossible due to ethical considerations, and is technically difficult to perform in other mammals, making resources for this type of research extremely rare. For example, the Functional Annotation of Mammalian Genome (FANTOM) 5 project based in Japan, established in 2000, is a worldwide collaborative project with the goal of identifying all functional elements in mammalian genomes, but it has no Cap Analysis of Gene Expression (CAGE)-based transcription start site (TSS) data for human embryos or for murine embryos representing early developmental stages.
To get around this paucity of information, an international collaboration of researchers from Japan (Kumamoto University & RIKEN), Russia (Kazan Federal University), Spain (University of Cantabria), and Australia (University of Western Australia) has put together the world's first genome-wide set of avian TSSs. These sites mark the starting point for transcription and are extremely important for cell differentiation and embryo development until birth, or, in the case of an avian species, until hatching. While birds look quite different from humans, early developmental stages are very similar, and it has been shown that about 60% of human protein-coding genes have one-to-one correspondence with chicken orthologs (genes that evolved from a common ancestor).
Using CAGE technology, a highly reliable method for finding TSSs and cis-regulatory elements (those sequence blocks regulating the position and robustness of transcription) in the genome over the full chick developmental period, the collaborators were able to map 60% of all developmental chicken TSSs to the most recent chicken genome, with the other 40% likely being as of yet uncharacterized alternative promoters or noncoding RNA genes. The updated genome was added to RIKEN's interactive, online platform ZENBU (The Japanese word "zenbu" translates to "all" in English.) under the name "Chicken-ZENBU" and is free for all to use.
The researchers also showed that their genome-wide TSS mapping could be applied to CRISPR-on technology to activate specific genes during development by successfully activating the T gene for Brachyury protein encoding at developmental stage HH10 (about 1.5 days of development after laying). This achievement was followed up by several more successful gene activations thereby showing the effectiveness of CRISPR-on when combined with TSS mapping.
"In the process of developing our CAGE-based developmental TSS profiling method, we identified new transcription factors and their regulatory modules, as well as several new housekeeping genes," said project leader Professor Guojun Sheng of Kumamoto University's IRCMS. "We expect this work and the data we added to the ZENBU online database to greatly advance developmental research in both birds and mammals."
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This research was published online in the open access journal PLOS Biology on 5 Sept. 2017.
[Resource]
Lizio, M., Deviatiiarov, R., Nagai, H., Galan, L., Arner, E., Itoh, M., ... Sheng, G. (2017). Systematic analysis of transcription start sites in avian development. PLOS Biology, 15(9), e2002887. doi:10.1371/journal.pbio.2002887
Journal
PLOS Biology