The transition from euchromatin to heterochromatin is a fundamental process that particularly reshaped the epigenomic landscape of Y chromosome. Its definitive genomic underpinning and broad functional impact are still unclear, as heterochromatin (e.g., that of human Y) is usually too repetitive to study. I have previously demonstrated that, the young Y (‘neo-Y’) chromosome of Drosophila miranda has just initiated such a transition, thus is a powerful model to unveil the evolution, regulation and functional interaction of heterochromatin. I showed that this neo-Y still harbours over 1800 genes, and only 20-50% of the sequences are transposable elements (TE). Over five years, I aim to: 1) precisely resolve the structure and insertion sites of TEs as a pre-requisite for studying heterochromatin, by combining state-of-art sequencing and bioinformatic techniques. 2) I will reveal the de novo heterochromatin formation triggered by TE insertions or the heterochromatin/euchromatin boundary shifts on the neo-Y, by comparing the binding profiles of histone modification hallmarks and insulator proteins of D. miranda to its sibling species D. pseudoobscura, which lacks the neo-Y. Such epigenomic changes have likely driven the exaptation or innovation of small RNA pathways that govern the TE mobility. 3) I will then identify the responsible small RNAs and their encoding loci, which are expected to have newly emerged or differentially expressed in D. miranda relative to D. pseudoobscura. 4) Finally, I will develop CRISPR/Cas9 in D. miranda to manipulate the expression of TEs encoding such small RNAs on the neo-Y, in order to scrutinize how TE/heterochromatin evolution on the Y would impact the chromatin landscape of the entire host genome. The combined aim of this multidisciplinary project is to generate a framework for understanding the basic mechanisms of how heterochromatin evolves; and open a new avenue toward the discovery of Y chromosome function beyond male determination.