PROJECT SUMMARY The research proposal aims to investigate the novel regulatory sequences and elements underlying novel phenotypes, to gain a deeper understanding of the genetic basis of morphological and cellular innovation. The evolution of morphological structures and traits is complex, and the origins and genetic mechanisms that drive the development of new cell types, tissues, and organs are not fully understood. Our long-term goal is to uncover the processes that govern morphological and functional diversity and complexity, which is a crucial step in understanding the evolution of complex life. Despite progress in this field, our current understanding is still limited. The goal of this research is to understand the origins of novel regulatory sequences and elements, how they are integrated into regulatory networks to contribute to genic and phenotypic innovation, and further impact evolution. To accomplish this, the laboratory has two major focuses: the study of how new genes and phenotypes, including expression phenotypes, are regulated, and the deciphering of the principles of sex-biased regulation from an evolutionary biological perspective. The first part of the proposed research aims to investigate the mechanisms governing the gain of expression of evolutionarily young genes in Drosophila. Specifically, the project will focus on the regulatory basis of new genes in Drosophila and using scRNA-seq and scATAC-seq to pinpoint enhancers and promoters for new genes and new expression. Additionally, the project will investigate the mechanism of pre-meiotic dosage compensation and identify putative novel players. The second part of the proposed research will focus on the genetic and epigenetic mechanisms for sex-biased novel expression. This will include using deep learning to reveal the basis of sex-biased chromatin accessibility, investigating the origin of novel enhancers from poised sequences, and studying the role of distant enhancers in gene expression novelties and sex-biased novelties. Overall, this project will use cutting-edge techniques and approaches to gain insight into the mechanisms that drive the evolution of new genes and their expression patterns in Drosophila. This study will provide important insights into the evolution of transcription regulatory networks and their contributions to novel traits, including expression phenotypes. Altogether, our integrative approach will help to elucidate the origination and evolution of novel regulatory circuits and their contributions to phenotypic innovation and evolution.