The Su(Hw) chromatin insulator protein alters double-strand break repair frequencies in the Drosophila germ line.

P element-induced double-strand breaks (DSBs) on the X chromosome of Drosophila melanogaster were repaired up to four times more frequently when functional Su(Hw) chromatin insulator protein was removed from all genomic binding sites. Simultaneous comparisons of interallelic gap repair frequencies at two target loci on the X chromosome confirmed that a Su(Hw) binding site nested within a template had no effect on DSB repair efficiency. The results suggest that the genome-wide homology search of broken ends for homologous template sequences is affected because it is the only step in the recombinational repair process with an apparent genome-wide interaction. We propose that the searching 3'-hydroxy ends gain a higher degree of freedom for the search in a su(Hw) mutant background.

Detection of poly(ADP-ribose) synthesis in Drosophila testes upon gamma-irradiation.

Poly(ADP-ribose) polymerase (PARP) activity is widespread among eukaryotes. Upon DNA damage PARP binds to DNA strand breaks and transfers ADP-ribose residues from NAD+ to acceptor proteins and to ADP-ribosyl protein adducts. This leads to branched polymers of protein-coupled poly(ADP-ribose) (pADPr). Because the germline of Drosophila has recently become important in the study of DNA double-strand break repair (DSBR) as opposed to somatic DSBR we tested whether the catalytic activity of PARP can be stimulated by gamma-irradiation during Drosophila spermatogenesis. Using antibodies against pADPr we detected a significant increase in PARP activity in male germline cells during spermatogenesis upon gamma-irradiation. Different stages of spermatogenesis revealed different subnuclear localization patterns of pADPr. In premeiotic and postmeiotic cells pADPr localized in a pattern overlapping with lamin and topoisomerase II at the nuclear rim. In primary spermatocytes pADPr is associated with three loci corresponding to the chromosomes at the nuclear periphery.

The Drosophila micropia retrotransposon encodes a testis-specific antisense RNA complementary to reverse transcriptase.

The micropia transposable element of Drosophila hydei is a long terminal repeat-containing retrotransposon present in both the autosomes and the Y chromosome. micropia expression gives rise to a complex set of sense and antisense RNAs transcribed primarily during spermatogenesis. The most abundant sense RNAs constitute an assortment of heterogeneous high-molecular-weight transcripts expressed as constituents of the Y-chromosomal lampbrush loops of primary spermatocytes. In addition, micropia encodes a full-length RNA that extends between the two long terminal repeats of the element. The major 1.0-kb antisense RNA characterized is complementary to the reverse transcriptase and RNase H coding regions of micropia. It is expressed from a testis-specific promoter during the primary spermatocyte stages and is detectable until spermatid elongation stages. Sequence comparison of this promoter with the 5' region of other testis-specific genes allows the conception of a conserved sequence that is responsible for this pattern of expression. A 284-bp fragment containing this sequence is able to drive testis-specific expression of the Escherichia coli lacZ gene in Drosophila melanogaster. This sequence is conserved in the micropia elements present in other Drosophila species that also encode an antisense RNA. The evolutionary conservation of micropia antisense RNA expression and the sequences responsible for its testis-specific transcription suggests a role for this antisense RNA in the control of germ line expression of the full-length transcript or transposon-encoded proteins.

Y chromosomal fertility genes of Drosophila: a new type of eukaryotic genes.

The Y chromosomal fertility genes of Drosophila are required for sperm differentiation. They are active only in primary spermatocytes where they form giant lampbrush loops. The molecular structure of these genes was investigated and revealed an unusual composition of DNA. Short, tandemly repeated sequence clusters are interrupted by longer and more heterogeneous sequences, which probably all represent transposable elements. No indication of the presence of protein-coding regions has been found within the fertility genes. However, the lampbrush loops bind site-specific proteins recognized by immunofluorescence techniques. This, together with other experimental data, led to the hypothesis that the Y chromosomal genes have a function in binding chromosomal proteins. The data and arguments in support of this gene model are summarized in this paper.