Deciphering piRNA-based immunity in germ cells across space and time

Summary My laboratory investigates how germ cells distinguish self from non-self RNAs to safeguard genome integrity across generations. Central to this work are small RNA pathways and the specialized condensates known as germ granules, which coordinate RNA regulation in the C. elegans germline. These mechanisms are crucial for protecting the genome from transposons and inappropriate gene silencing, ensuring fertility and proper development. Over the past decade, we have defined key rules that govern piRNA targeting and established how gene licensing pathways, such as CSR-1–associated small RNAs, actively protect self RNAs from piRNA-induced silencing. Our research has shown that licensing and silencing are not simply antagonistic but rely on highly orchestrated sorting decisions made within distinct sub-compartments of germ granules. We have also developed innovative tools, including synthetic piRNA systems and fluorescent reporters, that allow us to dissect small RNA activity and track regulatory outcomes in vivo. Building on these advances, our research over the next five years will address how intrinsic RNA features and associated proteins direct RNAs through specific regulatory fates—either protected or silenced. A major focus will be to understand how the spatial organization of germ granules, including functionally distinct sub-compartments, contributes to these decisions. We aim to elucidate the molecular mechanisms by which licensing pathways restrict RNAs from entering piRNA silencing domains, and how condensate interfaces regulate RNA flow and protein sorting. To achieve these goals, we are leveraging a suite of cutting-edge tools, including live- cell imaging, proximity labeling, CRISPR-based reporters, and inducible perturbation systems. These approaches will allow us to visualize dynamic RNA-protein interactions in space and time and to manipulate regulatory pathways with precision. Ultimately, our work seeks to uncover fundamental principles of small RNA-based genome defense, condensate organization, and epigenetic inheritance. These insights have broader implications for RNA regulation in other systems, and may inform the development of gene regulatory and therapeutic strategies that harness small RNAs for targeted gene regulation. Project Number: 1R35GM164060-01 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Heng-Chi Lee | Institution: UNIVERSITY OF CHICAGO, CHICAGO, IL | Award Amount: $444,788 | Activity Code: R35 | Study Section: Special Emphasis Panel[ZRG1 MGG-D (55)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11331706