Non-coding RNA in stem cell biology

Yale University, New Haven CT

Investigators

Abstract

PROJECT SUMMARY Non-coding RNA is an important regulator of gene expression and genomic stability. Such processes are particularly important in pluripotent stem cells where tight regulation of maintenance, division, differentiation, and (epi)genetic stability is crucial for a favorable outcome. Currently, there are major gaps in our understanding of how pluripotent stem cells are stably maintained long-term, or make the transition to differentiation. In our research program we leverage the in vivo study of a tightly controlled population of long- lived adult pluripotent cells, the neoblasts of the planarian Schmidtea mediterranea, to reveal the regulatory mechanisms surrounding pluripotency. This proposal concerns the role of non-coding RNA in the protection of the stem cell state, and the facilitation of the transition out of pluripotency during differentiation. We focus on the role of PIWI-interacting RNAs (piRNAs), and their binding partners the PIWI proteins. These small non-coding RNAs are essential for stem cell function, in particular in early embryos and regenerating animals. While piRNAs are best known for their silencing of transposable elements (TEs), we previously found that the piRNA pathway regulates a much broader range of non-coding transcripts in stem cells, and that many such transcripts are processed into piRNAs, raising the question of how the piRNA response remains contained. In this next period we will therefore address how source transcripts for the piRNAs are generated and recognized, and how this process balances the need to maintain flexibility to respond to novel invading elements with the need to keep the response away from important genes. Further we will analyze whether and how the piRNA pathway interacts with other non-coding RNA regulatory pathways that function in the stem cells. Following up on our previous findings on the role of PIWI proteins in the chromatin regulation around cell differentiation, we will identify the proteins involved in depositing histone modifications at piRNA target sites in the genome, and determine how the local chromatin changes at piRNA target sites propagate to failure of chromatin re-arrangement during differentiation. This is a fundamental question that is difficult to address in less prolific stem cell systems. Further, we will investigate a potential direct relationship between piRNA targeting and DNA damage repair. Together, the proposed experiments will provide deep mechanistic insights in the generation, interactions, and effects of piRNAs, which are essential for long-term stem cell function. Our research program addresses big questions in stem cell biology as well as molecular biology, spanning many layers of regulation. This work will have broad impact on our understanding of the regulation of pluripotent cells, and in the long run will contribute to the safe use of pluripotent cells in therapeutic applications.