Analysis of nuclear proteins in primary spermatocytes of Drosophila hydei: the correlation of nuclear proteins with the function of the Y chromosomal loops

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Abstract
The protein content of spermatocyte nuclei from X/Y males and mutants of D. hydei which lack different Y chromosomal loop forming sites, was compared with that of X/0 males in 14C/3H double labelling experiments. Proteins of 45,000, 52,000, 54,000, 66,000, 80,000, 84,000, and 170,000 Dalton are found to be enriched in nuclei containing two or more active Y chromosomal loop forming sites. These proteins are also present in the nuclei of X0 males. In the complete absence of the Y-chromosomal loops proteins of 35,000, 46,000, 58,000 and 110,000 Dalton become enriched in the spermatocyte nuclei. — Analysis of the nuclear RNP of spermatocytes led to the isolation of an hnRNP-containing fraction with an S-value of >900S (RNP-PP). — In the RNP-PP of XY males labelled protein material associated with hnRNA is enriched by a factor of ∼3 in respect to the X0 genotype. The nuclear RNP has a heterogenous buoyant density in CsCl of p = 1.33 to 1.43 g/cm3. RNase T1 treatment of the crude nuclear RNP from XY males prior to sucrose gradient analysis shows that the 66,000 Dalton protein which is also strongly enriched in the nuclei in the presence of active Y chromosomal loop forming sites, is the main protein associated with protected RNA-sequences of 80–120 and 200–300 nucleotides in length. Competitive nitrocellulose filter binding assays reveal that the 66,000 Dalton protein predominantly forms in 2 M NaCl stable RNA/protein complexes with the poly A +hnRNA of the RNP-PP. These RNP complexes have a buoyant density of p = 1.43 g/cm3 in CsCl. The results are discussed in relation to the nuclear structure and the function of the Y chromosomal loops during spermatogenesis in Drosophila hydei.