DNA replication in nurse cell polytene chromosomes of Drosophila melanogaster otu mutants

Abstract

Drosophila cell lines are used extensively to study replication timing, yet data about DNA replication in larval and adult tissues are extremely limited. To address this gap, we traced DNA replication in polytene chromosomes from nurse cells of Drosophila melanogaster otu mutants using bromodeoxyuridine incorporation. Importantly, nurse cells are of female germline origin, unlike the classical larval salivary glands, that are somatic. In contrast to salivary gland polytene chromosomes, where replication begins simultaneously across all puffs and interbands, replication in nurse cells is first observed at several specific chromosomal regions. For instance, in the chromosome 2L, these include the regions 31B-E and 37E and proximal parts of 34B and 35B, with the rest of the decondensed chromosomal regions joining replication process a little later. We observed that replication timing of pericentric heterochromatin in nurse cells was shifted from late S phase to early and mid stages. Curiously, chromosome 4 may represent a special domain of the genome, as it replicates on its own schedule which is uncoupled from the rest of the chromosomes. Finally, we report that SUUR protein, an established marker of late replication in salivary gland polytene chromosomes, does not always colocalize with late-replicating regions in nurse cells.

Similarity in replication timing between polytene and diploid cells is associated with the organization of the Drosophila genome

“…In recent years, the use of fluorescently labeled 5-bromo-2'-deoxyuridine and antibodies to PCNA, a protein marker of replication forks, has increased mapping resolution and revealed that there is no such thing as continuous labeling in salivary gland polytene chromosomes or in polytene chromosomes of pseudonurse cells of otu 11 mutants. When replication is seen virtually everywhere along the chromosome arms, the large black bands have not even started, but some regions of lightly packed chromatin have completed replication [ 39 , 40 ]. Thus, the propensity of thick black bands to be the last to commence replication is conserved in polytene chromosomes across different D .…”

“…melanogaster tissues. The order of replication completion by these bands is conserved too, as demonstrated by analysis of replication patterns in polytene chromosomes from different tissues in Drosophila [ 10 , 40 ] and in the mosquito Anopheles stephensi [ 41 ]. When 3 H-thymidine is used, analysis of the order of replication completion by bands becomes possible only after ≥40–45 sites on each chromosome arm have been labeled.…”