Tudor Domain Proteins in Germline Genome Defense

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 DESCRIPTION (provided by applicant): Transposable elements (TEs) are "genomic parasites" that can replicate and re-integrate into the host cell genome. Uncontrolled TE activity in germ cells leads to DNA damage, disruption of gametogenesis, and infertility. In mammalian male germ cells, the Piwi- piRNA pathway uses small RNAs as a guide to silence mobile TEs to protect genome integrity and sustain fertility. A unique family of proteins, the Tudor domain containing proteins (TDRDs), directly bind to Piwi proteins and facilitate their roles in TE silencing and spermatogenesis. The mechanistic involvement of TDRD proteins in conjunction with Piwi proteins in meiotic TE silencing is currently unknown. Mutation of TDRD proteins TDRD5 or TDRD7 alone leads to L1 (the most abundant TE in mammals) activation and male infertility. However, the mechanism by which they are integrated into the Piwi-piRNA pathway to suppress TE expression remains elusive. In addition to the Tudor domain, TDRD5 and TDRD7 also contain the Lotus domain, a novel putative RNA binding domain of unknown function. The overall objective of this proposal is to define the molecular mechanism whereby TDRD5 and TDRD7 post transcriptionally suppress TE activation during meiosis. Our preliminary data indicate that the Lotus domain is an RNA binding domain. We hypothesize that TDRD5 and TDRD7 act as "Tudor adaptors" to directly recruit TE mRNAs to the Piwi protein complex for degradation or sequestration. This is likely achieved by virtue of their Tudor and Lotus domains binding to Piwi proteins and TE mRNAs, respectively. To test this hypothesis, we will use biochemical approaches and mouse models to: 1) Determine the Lotus domain RNA binding properties; 2) Determine the differential binding mechanisms of TDRD5 and TDRD7 with Piwi proteins Miwi and Mili; 3) Define the precise role of TDRD5 in meiotic TE silencing in mice. These studies will provide valuable new insight into the mechanisms that safeguard germline genome integrity and sustain male fertility.