Dnmt3a and Dnmt3b are de novo methyltransferases required for establishing new DNA methylation patterns during early embryonic development and gametogenesis. Dnmt3s may need other interacting partners to establish specific DNA methylation patterns. Previous work in our lab has proven that both RIM-BP3 and MPc2 have the ability to interact with Dnmt3a. In order to explore their function during embryogenesis and gametogenesis, we generated the RIM-BP3-/- and MPc2-/- mice, respectively, and investigated their biological functions from developmental point of view.Part I Functional analysis of RIM-BP3 during spermatogenesis in miceRIM-BP3 belongs to the RIM-binding protein (RIM-BP) family. The other two paralogs, RIM-BP1 and RIM-BP2, are highly expressed in the brain and proposed to function as adaptors in the process of vesicle fusion and release. Our work demonstrates that RIM-BP3 is specifically expressed in the mouse testis. It is associated with the manchette, a transient microtubular structure believed to be important for morphogenesis during spermiogenesis. Targeted deletion of the RIM-BP3 gene resulted in male infertility due to abnormal sperm heads characterized by the deformed nucleus as well as detached acrosome. However, we didn’t find any abnormal pattern of DNA methylation in genome-wide level or in any paternal imprinted genes we examined. In our effort to investigate the role of RIM-BP3 during spermiogenesis, Hook1 was identified as RIM-BP3 interacting protein by affinity purification. Hook1 is a known manchette-associated protein required for sperm head morphogenesis. Interestingly, RIM-BP3 does not interact with the truncated Hook1 protein characterized in azh (abnormal spermatozoon head) mutant mice. Moreover, RIM-BP3 and Hook1 mutant mice share several common abnormalities, in particular with regard to the ectopic positioning of the manchette within the spermatid, a presumable cause for the sperm head deformities. These observations suggest an essential role for RIM-BP3 in manchette development and function through its interaction with Hook1. Since the occurrence of deformed spermatids is one of the common abnormalities that lead to malfunctional sperm, identification of RIM-BP3 might provide a molecular cue underlying certain human infertilities.Part II Functional analysis of Polycomb protein MPc2 during embryonic development and gametogenesis in mice MPc2, one of the murine homologues of the Polycomb-group genes, has been reported to be involved in transcription repression, X chromosome inactivation and epigenetic regulations. Our previous work showed MPc2 could interact with Dnmt3a and enhanced sumoylation of Dnmt3a in vivo and in vitro. MPc2-/- mice showed neonatal lethality with atelectasis and impaired thymus development, indicating abnormal organogenesis. We are generating MPc2 conditional knock-out mice in thymocytes to investigate its mechanism during thymus development. We also conditionally disrupted MPc2 in growing oocytes to study its role in maternal imprinting during murine oogenesis, but no abnormality has been observed so far.