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

Abstract

Morphologically, polytene chromosomes of Drosophila melanogaster consist of compact “black” bands alternating with less compact “grey” bands and interbands. We developed a comprehensive approach that combines cytological mapping data of FlyBase-annotated genes and novel tools for predicting cytogenetic features of chromosomes on the basis of their protein composition and determined the genomic coordinates for all black bands of polytene chromosome 2R. By a PCNA immunostaining assay, we obtained the replication timetable for all the bands mapped. The results allowed us to compare replication timing between polytene chromosomes in salivary glands and chromosomes from cultured diploid cell lines and to observe a substantial similarity in the global replication patterns at the band resolution level. In both kinds of chromosomes, the intervals between black bands correspond to early replication initiation zones. Black bands are depleted of replication initiation events and are characterized by a gradient of replication timing; therefore, the time of replication completion correlates with the band length. The bands are characterized by low gene density, contain predominantly tissue-specific genes, and are represented by silent chromatin types in various tissues. The borders of black bands correspond well to the borders of topological domains as well as to the borders of the zones showing H3K27me3, SUUR, and LAMIN enrichment. In conclusion, the characteristic pattern of polytene chromosomes reflects partitioning of the Drosophila genome into two global types of domains with contrasting properties. This partitioning is conserved in different tissues and determines replication timing in Drosophila.

Replication timing in Drosophila and its peculiarities in polytene chromosomes

“…Analysis of replication in polytene chromosomes using anti-PCNA immunostaining demonstrated that INTs replicate before rb-bands (Kolesnikova et al, 2018). Completion of replication in rb-bands correlates with the size of the regions.…”

“…D. Koryakov and I. Zhimulev (2015) used BrdU to study replication in polytene chromosomes from nurse cells of D. melanogaster otu mutants. In recent years, replication detection with PCNA antibodies has been extensively used (Gibert, Karch, 2011;Kolesnikova et al, 2013Kolesnikova et al, , 2018Andreyeva et al, 2017). The high resolution of the method revealed that at the continuous labeling stage, when replication was seen almost everywhere along the chromosome arms, large black bands had not even started replication, whereas some regions of loosely packed chromatin had already completed it (Kolesnikova et al, 2013;.…”

“…Kolesnikova et al (2018) mapped borders of all 159 prominent black bands on chromosome 2R to the Drosophila genome assembly with a high accuracy. These 159 bands are characterized by the presence of silent chromatin ruby (Zhimulev et al, 2014) and hence were named rb-bands (Kolesnikova et al, 2018). The rb-bands demonstrate low gene density, low gene expression level in individual tissues, and enrichment in transcriptionally inactive types of chromatin.…”

“…Border regions between rb-bands and INTs often coincide with borders of topological domains. The borders are characterized by a dramatic difference in the presence of multiple epigenetics markers such as H3K27me3, Lamin, SUUR, and histone H1 (Kolesnikova et al, 2018). INTs and rb-bands split chromosome 2R into two types of domains with contrasting properties, which are conservative in different tissues (Kolesnikova et al, 2018).…”

Polytene chromosomes reflect functional organization of the Drosophila genome

“…Ruby chromatin (former magenta) occupies about 50 % of the genome and it lacks proteins characteristic of open chromatin (Zhimulev et al, 2014). It is localized in black transcriptionally inactive bands, mainly containing tissue-and stage-specific genes, including the largest bands of intercalary heterochromatin (Khoroshko et al, 2016;Kolesnikova et al, 2018). Lazurite (former blue) and malachite (former green) chromatin states have an intermediate degree of compaction and mainly cor respond to gray bands and transition zones between the inter bands and black bands (Zhimulev et al, 2014;Boldyreva et al, 2017;Khoroshko et al, 2018).…”