Veikko Sorsa

Electron microscopic observations on interband fibrils in Drosophila salivary chromosomes.

Acetic alcohol squash, preparations of salivary chromosomes of Drosophila melanogaster stained with uranylacetate during fixation and dehydration were sectioned and examined with the electron microscope. At high magnification the interband regions are seen to be composed of coiled fibrils in the range of 50 Å which often seem to be arranged in pairs. Submicroscopic bodies found along the interband fibrils seem to delimit successive subunits. It is suggested that transverse coiling of the fibrils within the boundaries of each subunit leads to the formation of chromomeres.

Electron Microscopic Mapping and Ultrastructure of Drosophila Polytene Chromosomes

The number of molecular studies involving cloned segments of DNA has been growing rapidly. This development holds a promise of rapid progress in the study of individual genes of Drosophila melanogaster. The exact sequences of DNA of several genes evidently will be available in the near future (cf. Spradling and Rubin, 1981). Consequently, the cytogenetic localization of cloned sequences on the chromosomes is also going to meet new challenges. More exact cytological methods have to be developed to demonstrate the normal banding pattern of polytene chromosomes and to recognize minute rearrangements in it. The quality and usability of all new methods should be tested against the revised light microscopic reference maps of Bridges (cf. Lindsley and Grell, 1968). An urgent task, however, is the reliable verification and possible revision of light microscopic maps by means of high-resolution electron microscopy. EM division maps for the salivary gland chromosome 2L of D. melanogaster are included in this review.

Ultrastructure of induced transitions in the chromatin organization of Drosophila polytene chromosomes.

The structural changes taking place in the salivary chromosomes of Drosophila melanogaster after treatment with urea-sodium hydroxide solution were studied by light and electron microscopy. An essential effect of the treatment is the gradual disappearance of the chromosomal banding pattern due to uncoiling of the chromomeric fibrils. During this process a huge amount of very thin fibrillar network is detached from the salivary chromosomes, and the longitudinal interband fibrils become aggregated to form a distinct central axis. This gives apparent likeness to a lampbrush chromosome. Even though at the light microscope level certain regions of the axial core appear to have been lost, no signs of breaks in the linear coherence of the chromosome can be observed in the electron micrographs. Because uncoiling of the chromomeres does not interrupt the continuity of the linear fibres, these observations on induced transitions lend support to the idea that the chromomeric fibrils are to some extent independent and dissimilar as compared to the interchromomeric fibres.

Interband fibrils of polytene chromosomes

Abstract Polytenization of chromosome complement is characteristic to the differentiation of salivary gland cells of Dipteran insects. Ultrastructure of polytenized chromosomes was studied in Drosophila cells by means of thin section EM. According to the high-voltage EM of whole-mounted polytene chromosomes the interband fibers are 20–30 nm wide helices of nucleosome fibril, which implies that the DNA content of interband fibres is ca. 20–30 times their axial length in chromosomes. To test this concept the ultrastructural organization of interband fibrils was studied from the salivary gland chromosomes of Drosophila melanogaster after different fixations and differential stretching of identified interbands. The results do not support the existence of nucleosome helices in the interbands. The thickness of individual interband fibrils is only about 5 nm in the thin sections of unstretched chromosomes and the length of fibrils can be increased only ca. 2 times by the stretching, which indicates that their DNA is already in relatively extended stage.

Ultrastructure of regions containing homologous loci in polytene chromosomes of Drosophila melanogaster and Drosophila subobscura

Abstract. We have used a new approach involving in situ hybridisation and electron microscopy to establish ultrastructural homologies between polytene chromosome regions of Drosophila melanogaster and Drosophila subobscura. Twelve probes were chosen to cover all the chromosomal elements: the myospheroid gene, the collagen type IV gene, the collagen-like gene, the w26 homeobox gene, the β3 tubulin gene, the kinesin heavy chain gene, the tryptophan hydrolase gene, the Hsp82, Hsp22–26 and Hsp23–28, Hsp68, Hsp70 genes and the β unit of the F0–F1 ATPase gene. Most of these loci were previously undescribed in D. subobscura and imprecisely located in D. melanogaster. We have demonstrated here, by an ultrastructural analysis of each chromosomal region, that homologous genetic loci tend to show a similar ultrastructure in the two species. With a few exceptions, the structural homology extends to the chromosomal regions surrounding the loci. In some cases, however, no structurally recognisable homology can be seen either in the locus or in its flanking regions.

Electron Microscopic Map for the Salivary Gland Chromosome X of Drosophila Melanogaster. Divisions 1–5

Shortly after Painter’s first report on the salivary gland chromosomes of Drosophila melanogaster also C.B. Bridges started an extensive study to achieve a detailed mapping of the banding pattern in these chromosomes (Painter, 1934). A year later, in December 1934, when Painter’s map for the whole genome of Drosophila was published, Bridges also presented the complete camera lucida maps of all salivary gland chromosomes in the annual exhibition of the Carnegie Institution of Washington. The same maps demonstrating 3540 bands were published in 1935 with a reference system for identification of all bands (Bridges, 1935). Unfortunately, as pointed out by Bridges himself, the fainter bands were shown for technical reasons too conspicuous and dark in the printed map. This was clearly shown also by Lefevre (1976) in a comparison of the photographic maps and Bridges’ (1935) maps.

Structural Analysis of Polytene Chromosome Bands and Interbands

Beermann (1972) estimated that at least 95% of the DNA of polytene chromosomes of Drosophila is located in bands and only less than 5% in the interband regions. Recent studies of Laird et al. (1980) on high-voltage electron micrographs of whole-mounted polytene chromosomes have provided arguments which call for a reevaluation of the previous results. Densitometric determination of cross sectional relative dry mass in polytene chromosome bands and interbands from the EM negatives has suggested that the earlier estimates of the proportion of interband DNA are too low (Laird, 1980). According to the results obtained from the densitometric studies on the whole-mounted chromosomes at least 26

The polytene dot chromosome of Drosophila: D. melanogaster and D. subobscura.

of the total DNA is located in the interbands of polytene chromosomes. This new result, if it holds true, has several interesting and important implications. It favors the hypotheses that the interband regions are the sites of permanently active “housekeeping” genes in polytene chromosomes (e.g., Crick, 1971; Speiser, 1974). The high number of interbands, more than 5,060 in the genome of Drosophila melanogaster (Bridges, 1942), and obviously the high variability in their number in polytene chromosomes of related species, like Drosophila hydei (Berendes, 1963), contradict the view, that all interbands are active genes.

Ultrastructure of E1 + 2 + 9 + 12 inversion breakpoints in Drosophila subobscura

Abstract. The chromosome arms are assumed to be homologous within the genus Drosophila. Homology at the level of the polytene chromosome banding pattern between non-sibling species is, however, almost impossible to establish as different processes such as inversion, transposition and unequal crossing over, have disturbed it. Even though the band sequences cannot be followed, we may ask whether there is a correlation in the total number of bands between species. The polytene dot chromosome is an excellent starting point for such an approach. Here we present the detailed cytology of polytene chromosome 4 of D. melanogaster and the polytene dot chromosome of D. subobscura using electron microscopy. The results show that the number of bands is about the same, around 30, in both species. We predict that by using thin sections and electron microscopy for the longer polytene chromosome arms, both species will turn out to have approximately equal band numbers.

Electron microscopic analysis of the banding pattern in the salivary gland chromosomes of Drosophila melanogaster

The ultrastructure of the Drosophila subobscura chromosome regions around the breakpoints of the complex E1 + 2 + 9 + 12 gene arrangement was analyzed. This overlapping inversion is formed by the association of the E1, E2, E9, and E12 simple inversions. Ultrastructure of sections involving 58D/59A, 61C/D, 62D/63A, 64B/C, 67A/B, and 68B/C breakpoints on Est chromosomes were compared with the ultrastructure of sections involving 58D/68B, 62D/64C, 59A/63A, 64B/68C, 67B/61C, and 67A/61B breakpoints on E1 + 2 + 9 + 12 chromosomes. No detectable changes of structural organization on banding patterns induced by the E1 + 2 + 9 + 12 inversion were found. Ultrastructural analysis of the two E12 breakpoints has, however, facilitated the analysis of the left boundary of E12 inversion. Accordingly, we propose 61B/C as a new breakpoint instead of 61C/D.