DNA clones and RNA transcripts of four lampbrush loops from the Y chromosome of Drosophila hydei

Drosophila hydei clones representing transcribed middle-repetitive sequences from four of six major lampbrush loops of the Y chromosome were isolated. Sequences homologous to each clone are clustered in a particular locus on the Y chromosome, but additional euchromatic sites were found for one of the transcribed clones. In situ hybridization to lampbrush-loops RNA permitted the identification of clones homologous with the two "nooses" loops on YS and with the "clubs" and "tubular ribbons" on the YL arm. Loop-specific nuclear RNA molecules range in size from 10S to 60S. Loop RNA is accumulated in the nucleus and remains attached to the loops during the course of primary spermatocyte growth. It disappears, however, along with the loop structures, during the first meiotic prophase. The structure and function of the Y chromosome and its lampbrush loops are briefly considered in the light of these findings.     

Y chromosomal DNA of Drosophila hydei

Six recombinant DNA clones are described, which are derived from the Y chromosome of Drosophila hydei. They reveal characteristic features of Y chromosomal DNA sequences. Three of the cloned inserts are Y-specific and are members of the same family of repeated sequences associated with the lampbrush loop-forming fertility gene "nooses" in the short arm of the Y chromosome. The other three cloned sequences are members of three different families of repeated sequences, but display a small amount of homology to one another and to the family of the nooses sequences. These three cloned sequences are found preferentially in the Y chromosome, but also in other chromosomal positions. The Y chromosomal copies are located in the short arm of the Y chromosome. The other copies are found in autosomal kinetochore-associated heterochromatin or, for one of the cloned sequences, in one band of the giant chromosome 4, in addition to the kinetochore heterochromatin.     

Characterization of the long terminal repeats of micropia elements microdissected from the Y-chromosomal lampbrush loops "threads" of Drosophila hydei

Four micropia elements from Drosophila melanogaster and D. hydei have been analysed by sequencing. Two elements, from D. hydei, micropia-DhMiF8 and -DhMiF2, were recovered by cloning microdissected Y-chromosomal lampbrush loops "threads". This method allows isolation of repetitive sequences from defined chromosomal positions, but recovery of large and overlapping inserts is difficult. In case of the Y-chromosomal micropia elements it was not possible to define the endpoints of their long terminal repeat sequences precisely. Comparison of these locus-defined micropia elements to complete micropia elements isolated from D. melanogaster allowed identification of micropia-DhMiF8 and micropia-DhMiF2 long terminal repeats (LTRs). LTR sequences from the two Drosophila species are not conserved except for a few short sequences found at comparable positions that are believed to have functional significance. In contrast, the Leu-tRNA primer binding site and plus strand primer binding site are conserved between D. melanogaster and D. hydei.     

Molecular cloning of microdissected lampbrush loop DNA sequences of Drosophila hydei

We microdissected a Y chromosomal lampbrush loop pair from primary spermatocyte nuclei of Drosophila hydei and cloned the DNA directly at the microscale. Four of the 12 recombinant DNA clones recovered display in situ hybridization to mitotic metaphase Y chromosomes, preferentially in the chromosomal region identified as the origin of the lampbrush loop pair. All clones, however, also hybridize to autosomal and X chromosomal loci in polytene chromosomes. Y chromosomal DNA sequences of D. hydei again prove to be members of different families of repeated sequences distributed throughout the genome. These microcloning experiments, which were carried out under very unfavourable experimental conditions (low DNA content of the lampbrush loops in the presence of large amounts of RNA) prove that almost any chromosomal structure detected by light microscopy is directly accessible to molecular cloning experiments by micromethods.     

Temperatursensitive Mutanten im Y-chromosom von Drosophila hydei

Mutations were induced in the Y chromosomal fertility genes of Drosophila hydei by EMS treatment of adult males. Four types of mutants were observed: 1. Sterile mutants without detectable cytological changes in Y chromosomal lampbrush loops. 2. Sterile males with morphologically changed loops. 3. Sterile males where one or several Y chromosomal loops are missing in the spermatocytes. 4. Mutants which are temperature-sensitive for sterility, development of loops or altered loop morphology. In this paper four Y mutants are described which are temperature-sensitive as regards fertility but which show unchanged lampbrush loops. They can be mapped in four different complementation groups. Two of those occur probably in regions of the Y chromosome without cytologically detectable lampbrush loops. All mutations are found in the distal half of the long arm. The temperature-sensitive period occurs during the primary spermatocyte stage and in early spermatid development while the manifestation of the effect occurs postmeiotically. The mutants are further characterized with respect to changes in the ultrastructure of the sperm at the restrictive temperature.     

Evolution of Y chromosomal lampbrush loop DNA sequences of Drosophila

The evolutionary conservation of Y chromosomal DNA sequences of Drosophila hydei in different species of the genus Drosophila was studied by in situ hybridization and on genomic DNA blots of restriction enzyme digested DNA. We demonstrated that Y specific DNA sequences, which form major parts of lampbrush loops related to the male fertility genes, are only retained in a few closely related species during evolution. Other Y chromosomal DNA sequences, also present in lampbrush loops but with homology to autosomal and X chromosomal locations, were found in distant species. We propose a model for the evolution of the Y chromosomal lampbrush loops.     

Genetic function correlated with unfolding of lampbrush loops by the Y chromosome in spermatocytes of Drosophila hydei

Summary Deficiencies of the Y chromosome of Drosophila hydei including sites which develop lampbrush loops invariably cause sterility of males. Suppression of loop unfolding in one or more sites equally results in similar morphogenetic defects of spermiogenesis. A variegated type repression of lampbrush loop unfolding observed during the spermatocyte stage results in varying morphogenetic effects on spermiogenesis. This demonstrates the existence of causal relationships between the active phase of Y chromosomal factors in spermatocytes and the differentiation processes in spermatids.In some translocated Y fragments the mode of unfolding of a particular pair of lampbrush loops may be permanently changed. As a result, lampbrush loops of a mutant phenotype are developed. Some alterations of this type are correlated with functional alterations resulting in defective spermiogenesis.Three different fragments of the Y chromosome in which lampbrush loop formation was repressed have been tested for possible reversions of loop suppression by means of X irradiations. In none of the three cases reversion has been detected among two thousand tested chromosomes.To the memory of Karl-Heinz Bier.     

The function of the lampbrush loops formed by the Y chromosome of Drosophila hydei in spermatocyte nuclei

Summary The function of pairs of translocated fragments of the Y chromosome of Drosophila hydei was tested. As the pairs of fragments together had a complete set of Y chromosomal sites, complementation of their function could be predicted according to results of earlier experiments. In contrast to the earlier experiments the development of lampbrush loops during the spermatocyte stage was blocked in one partner of each combined pair. As a consequence, no complementary effect on spermiogenesis is detectable. The results indicate that the formation of lampbrush loops by seven sites in the Y chromosome is a necessary prerequisite for the normal progress of spermiogenesis. This can be considered as further support of the view that the lampbrush loops in spermatocyte nuclei of Drosophila are phenotypic manifestations of the activity of male fertility factors.Supported by the Deutsche Forschungsgemeinschaft.