Fig. 6: BbYY1 interacts with bbRAG1L/2L.

a The co-immunoprecipitation (co-IP) assay indicated that bbYY1 interacted with bbRAG1L/2L in a dose-dependent manner. b The co-IP assay indicated that bbYY1 did not interact with bbRAG1L by transfecting MBP-tagged bbRAG1L and Flag-tagged bbYY1 expression vectors into 293T cells. c The co-IP assay revealed the interaction between bbYY1 and bbRAG2L by transfection of MBP-tagged bbRAG2L and Flag-tagged bbYY1 expression vectors into 293T cells. d The GST-pulldown assay indicated the direct interaction between bbYY1 and bbRAG2L (or bbRAG1L/2L complex). e The zinc-finger domain (ZNF) of bbYY1 is indispensable for the bbYY1-bbRAG1L/2L interaction. SDS-PAGE and western blotting of the indicated GST-pulldown assays. See also Supplementary Fig. 6E, G. The co-IP and GST-pulldown assays in a–e represent three independent duplications. f A summary diagram illustrates multiple roles of bbYY1 in regulating bbRAG1L and bbRAG2L expression and ProtoRAG-mediated DNA recombination and transposition. First, through recognition of the TR5 element on ProtoRAG, bbYY1 may recruit co-repressors or compete with activators to suppress ProtoRAG transposon expression, contributing to lower transposition activity and a reduced transposition footprint in the lancelet genome. Second, by interacting with bbRAG1L/2L, bbYY1 can mediate bbRAG1L/2L precise targeting to the correct TIR signal sequences, which would lead to precise cleavages and HDJs. Hence, bbYY1 probably protects the intact transposon structure of ProtoRAG against decay. Third, bbYY1 can attenuate TIR-dependent transposition but improves TTJ formation, leading to suppressed transposition activity of ProtoRAG. For a–e, source data are provided as Source Data file.