Fig. 6. Mouse KIN17 protein associates with DNA and RNA in testes. (A) In vivo UVC cross-linking of DNA to KIN17 in testicular cells. Suspensions of freshly isolated testicular cells were irradiated with UVC. Cells were then lysed and treated with DNase I. KIN17 protein was immunoprecipitated with rabbit anti-KIN17 antibody (lanes 1 and 3) or pre-immune serum (PIS) (lane 2, control) (lower panel). The immune complex was end-labelled with [-³²P]ATP and T4 polynucleotide kinase and resolved on a 10% SDS-PAGE gel. The autoradiogram is shown in the upper panel. (B) The labelled material migrating at the position of KIN17 protein was excised from the gel, digested with proteinase K, subsequently re-digested with DNaseI and resolved by 10% PAGE in 8 M urea; the autoradiogram of this gel is shown. A labelled smear is observed only in lysates from UVC cross-linked cells and mainly in the lane containing the immunoprecipitation with anti-KIN17 polyclonal antibody. The deproteinated labelled polynucleotide that had been cross-linked to KIN17 protein is sensitive to degradation by DNase 1. (C) In vivo UVC cross-linking of RNA species to KIN17 in testicular cells. The cells were treated as above excepting that RNase A was used instead of DNase 1. (D) Note that the deproteinated labelled polynucleotide that had been cross-linked to KIN17 is sensitive to degradation by RNaseA.