Recommended if funds allow
Statement of Benefit to California:
SYNOPSIS: The experiments in this proposal will utilize ChIP on Chip and other techniques to provide a comprehensive map of promoters, enhancers and insulators in hESCs. Genome-wide localization studies will map the position of transcription factors, as well as different types of chromatin modifications within the genome of hESCs. This will results in a catalog of information with which to manipulate the fate choices of these cells. SIGNIFICANCE AND INNOVATION: The innovation in this proposal lies in the application of state-of-the-art genomic technologies to hESCs. Similar approaches have already been reported in the literature using the hESC and murine ESC systems. Thus the innovation is not particularly high. However, these are exactly the kinds of approaches that need to be pursued to attain the goals of the overall proposal. The significance is quite high given the potential to generate a comprehensive catalog of chromosomal phenomena that could correlate with pluripotency. These studies are largely descriptive; however, they are necessary before embarking on detailed and comprehensive functional efforts. The secondary reviewer noted that the major goal of this proposal is to identify transcriptionally active regions, and the significance is that it will identify the location of putative pre-initiation complexes, enhancers, and insulators throughout hESCs. STRENGTHS: The major strength of this proposal is the excellent level of expertise in all of the technologies to be employed. The PI's laboratory was one of the developers of the ChIP on Chip technology. There is a high level of confidence that valuable and large amounts of information will be obtained. The 4 Aims will proceed from the identification of active promoters, transcriptional enhancers, to insulator elements in hESCs. The final Aim will begin testing some of the identified elements in functional assays. Of further merit are the computational tools that will be employed in the analysis of high volume datasets. Clearly the strength of this proposal lies with the investigator, who was one of the founders of the ChIP-chip assay and has a strong track record of productivity and expertise in ChIP-chip. There is very little doubt that the proposed aims will be accomplished successfully. The outcome will provide new knowledge on the location of pre-initiation complexes, enhancers, and insulators in the genome of hESCs. WEAKNESSES: One reviewer felt that the major weakness of this proposal is its scope. Clearly, it seems to be an effort that will take more than two years. In fact, each of the four Aims could easily be a separately funded efforts. Nevertheless, this is a superb investigator, and even if not all goals are achieved, there will certainly be a wealth of information generated. There is some concern that the PI does not propose to integrate the data to be obtained with recently published datasets also obtained in the hESC system. To the other reviewer, the major weakness is that the data that will be acquired will be a catalog of sites in hESCs, however, this is unlikely to make an immediate contribution towards the understanding of hESC regulation. Ultimately this information will be important in providing a base of knowledge. But at this stage the reviewer did not see the immediate downstream utility for understanding hESC regulation. Moreover, it is not clear that the major goal of this proposal could not be attained using tiled expression arrays. DISCUSSION: There was no further discussion beyond the reviewers' comments.