N6-methyladenosine (m6A) is the most prevalent internal modification in mRNAs. Recent studies showed that, aside from existing in mRNAs, m6A is also widespread in chromatin-associated RNAs (caRNAs) and regulates gene expression by recruiting regulatory proteins for histone PTMs (e.g., methyltransferases) to deposit histone epigenetic marks in nearby regions. Based on these observations, we hypothesize that histone epigenetic marks should be enriched at those genomic loci associated with m6A-marked caRNAs. Thus, it is possible to predict the interplay between m6A and histone epigenetic marks through bioinformatic analysis of publicly available m6A-seq data for caRNAs and ChIP-seq data for histone epigenetic marks. Furthermore, we posit that a systematic investigation of protein-protein interactions and protein-nucleic acid interactions in this crosstalk network will uncover novel epitranscriptome-epigenome crosstalks.

   In this project, we propose to employ quantitative proteomics and bioinformatic methods to reveal new crosstalks between m6A and histone PTMs in human chromatin. We will also decipher the molecular mechanisms underlying these crosstalks, and elucidate the functions of these crosstalks in regulating gene expression, genome instability and cancer development.

   The outcome of the proposed research will substantially improve our understanding about the molecular mechanisms governing m6A-mediated epigenetic regulations in both physiological and pathological contexts, and it will build a strong foundation for developing effective strategies for cancer treatment.

Recent clinic studies showed that mutations in splicing factors (e.g., U2AF1 and SRSF2) are strongly associated with myelodysplastic syndromes (MDS), AML and lung cancer. Ectopic expression of these mutant proteins in human cells led to a global elevation of R-loops and Pol II pausing at transcriptional start sites (TSSs). R-loops have gained substantial attention owing to their important roles in gene regulation, DNA damage repair, and genome stability maintenance. We have recently uncovered a group of candidate proteins that can bind to both R-loops and DNA guanine quadruplexes (G4s), and revealed the function of U2AF1-R-loop interaction in co-transcriptional splicing. In our future research, we plan to examine:

(1) If the findings made for U2AF1 could be extended to other splicing factors.

(2) How disease-associated mutations of these splicing factors affect the functions of protein-protein and protein-R-loop interactions.

(3) How environmental exposure affects the functions of protein-R-loop interactions.

    The outcome of this research will provide a biochemical basis for a better understanding of the functional roles of protein-R-loop interactions in cells, as well as offer insights into how protein mutations or environmental exposure contribute to genome instability.