Library

Notes: Summary The refractive indices of a number of bands and interbands of the polytene chromosomes in the living salivary gland cells of two species of Chironomus were measured with an interference microscope, and values were obtained for the total solid material present in each region. To make these measurements it was necessary to compress the cells until virtually no cytoplasm lay above or below the nucleus, but this produced no deleterious effects on the cells as a whole. The results indicated that there is not less than 25% of solid material in the band regions of these chromosomes, and not more than 15% of solids in the interbands. The nuclear sap of these cells was found to contain about 12% of solid material. Similar refractive index measurements were made on the bands and interbands of isolated chromosomes mounted in saline and in a 12% isotonic saline/protein solution of the same refractive index as the nuclear sap. In both these media the bands were again found to contain at least 10% more material than the interbands, but their refractive indices were lower in the saline than they were in the intact cells and markedly higher in the saline/protein. It is therefore concluded that the isolated chromosomes are, to some extent, permeable, and can permit the entry of protein molecules from the mounting medium and the egress of some or their own material.

Notes: Summary A cytological map of Drosophila hydei Sturtevant, one of the species of the repleta group, is presented together with some cytological observervations, including the description of three X-ray-induced rearrangements. This map of D. hydei is compared with the existing map of D. repleta Wollaston. In addition to the large standard rearrangements, a number of small differences are indicated. On the basis of rearrangements, two of the linkage groups are attributed to definite chromosomes.

Notes: Summary In Drosophila hydei abnormal puffing activities could be induced by temperature shocks and treatments with 1.2% and 1.4% KCl solutions. After temperature shocks in vivo and in vitro, a number of puffs showed a similar change in activity. Other puffs were found to show a change in activity only after a distinct treatment. Some of the puffs, specific for temperature shocks, showed a change in activity only at a distinct stage of development. In discussing the results, particular attention is paid to puffs observed in common after all treatments.

Notes: Summary The salivary glands of D. hydei larvae show differences between the cells in the distal (posterior) part and those of the proximal (anterior) part during the third instar. The first sign of these differences is an increase in cellular and nuclear volume in the distal cells of the gland, beginning at 103 hours after oviposition. After 125 hours the cytoplasm of the extreme distal cells acquires a reticulated structure, and at 130 hours these cells contain large granules or droplets of mucoprotein. From this moment up to puparium formation the number of cells containing these granules increases and the boundary of this type of cells shows a shift in the proximal direction. Just before puparium formation the granules disappear from the cells and a glue substance is secreted by the larvae. At this moment only a few cells in the extreme proximal part still lack granules. Electron-microscopical observations indicate that these cells were active in secretion, whereas all cells containing large granules are inactive in this respect during most of the third instar. During the early third instar a change in cell function occurs, i.e. from synthesis of substances presumed to be digestive enzymes which are secreted, to a synthesis of a glue substance which is stored. This change begins in the extreme distal cells of the gland. Investigation of the chromosomal puffing pattern revealed that a total number of 148 puffs were present during some period of the third instar, prepupal, and early pupal stages. The activity of 110 puffs was evaluated during a series of successive time intervals. Changes in the puffing pattern during puparium formation were compared with those observed during pupation. Proximal and distal nuclei differ in the activity level of a number of puffs, but only puff 47 B is restricted in activity to the distal cells. This puff becomes active at 119 hours and disappears 4 hours before puparium formation (156 hours). Determination of nuclear diameter and DNA in nuclei of both parts of the gland revealed a correlation between a particular DNA content and the function of the cell. Distal cells show higher nuclear diameters than proximal cells after the onset of granule production. The first differences in nuclear diameter can be seen at 103 hours. Cells in the transitional part of the gland, located between distal granulecontaining and proximal granule-negative cells, always show the same DNA content. These cells are found at different locations within the gland during the third instar. This zone of cells shows a shift in proximal direction during the third instar, identical to that of the neighbouring granule-containing cells. The possible interrelation between nuclear DNA content, the activity of puff 47 B, and the production of the glue substance were discussed.

Notes: Abstract The replication patterns of larval salivary gland chromosomes of D. hydei and D. melanogaster were studied by autoradiography with tritiated thymidine injected in mid third instar larvae. The male X chromosome showed a different replication behavior in comparison to that of the female X chromosome and autosomes. It is concluded that the male X chromosome finishes its replication earlier than the female X chromosome. Moreover, the time needed for a complete replication cycle of individual identical replication units was found to be shorter in the male than in the female X chromosome. Although the whole X chromosomes behave different there were no differences observed in the sequence of the discontinuous labeling patterns of the two types of X chromosome. One autosomal replication unit was observed which showed a different replication behavior in males and females. The possible origin of the differential behavior of the two X chromosomes is discussed in terms of their difference in degree of polyteny.

Notes: Abstract The hormone ecdysone induces a large number of changes in the puffing pattern of mid third instar larvae of Drosophila hydei. The pattern of changes occurring after experimental administration of the hormone are identical with those observed in normal development during a 6 hour period before puparium formation. After administration of the hormone a considerable number of puffs react with a change in activity within 15–20 min. During this period 3 puffs arise newly, 12 puffs show a strong increase in activity, 6 puffs show a less pronounced increase in activity and 12 puffs show a decrease in activity. At a period of 4–6 hours after administration of the hormone another 5 puffs arise newly. The effect of the hormone was identical in both in vivo and in vitro experiments. — Diameter measurements on several puffs reacting within 30 min with an increase in diameter showed that these puffs reacted simultaneously. Most of the puffs that showed a decrease in activity reacted with some delay. — A study of the effect of different hormone concentrations revealed that the kinetics of 4 puffs with respect to the relationship between concentration and puff size was identical over a range of concentrations from 33·10−5 to 33CU/μl. Three of these puffs showed a reaction with even lower concentrations. Maximum puff size is attained by all puffs at a concentration of 33·10−4CU/μl. Among the puffs studied no difference in their reaction threshold was found. — A study of the behavior of 5 puffs of the group reacting within 15–20 min and one of the group reacting after 4–6 hours in midintestine and Malpighian tubules revealed that these puffs showed the same reaction after injection of the hormone as observed in the salivary glands. — All puffs activated by administration of the hormone showed particularly strong uptake of tritiated uridine and accumulation of acidic protein. — It is concluded that the hormone ecdysone induces a pattern of changes in gene activity that is far more complex in Drosophila hydei than in Chironomus tentans.

Notes: Abstract In order to separate some of the factors involved in the formation of puffs the antibiotic actinomycin D was applied at different stages of puff activity. Puffs were induced by temperature shocks or eodysone. Inhibition of RNA synthesis with actinomycin D before application of a puff inducing stimulus prevents neither the appearance of the stimulus specific puffs nor the accumulation of acidic proteins in the puff regions. The puffs attained under these conditions approximately 1/3 of the size normally produced by the stimulus. Indications were obtained that during puff formation acidic protein accumulation precedes the onset of RNA synthesis. Synthesis and storage of newly synthesized RNA within the puff region was studied on the basis of grain distribution in uridine-H3 autoradiographs after various incubation periods. RNA synthesis appears to be restricted to a particular area of the puff region. After a 3 min temperature shock following injection of uridine-H3 silver grains are located only over a particular area of the newly formed puff. The same area becomes labeled during a 1 min pulse of uridine-H3 applied at a stage of maximum puff development. Longer periods of incubation result in a random distribution of the grains over the whole puff region. Grain counts on different areas of experimentally induced puffs and on the same areas at a stage of puff regression indicate that the newly synthesized RNA becomes transferred from the area where it was synthesized and is stored for a certain period within the puff region. Complete release of newly synthesized RNA from puffs in which RNA synthesis was inhibited by actinomycin D at a stage of maximal activity is accomplished within 30 to 35 min.

Notes: Abstract The submicroscopic organization of terminal chromosome regions of Drosophila hydei polytene chromosomes is described. A compact region composed of tightly packed fibrils of 100 to 125 Å diameter embedded in an amorphous material is located at each of the chromosome ends of the 5 long chromosome arms. From this compact region, sometimes containing cavities, fibrils extend onto the nearest normal band region. The diameter of the extending fibrils is 100–125 Å, 200–250 Å or 400 Å. Pronase digestion of fixed and squashed chromosomes reduced the electron density of the amorphous matrix in the compact regions but failed to affect the diameter of the fibrils. The extending fibrils, however, showed a decrease in diameter after pronase digestion. The most frequently observed diameter values were 100–125 Å. — The volume of the terminal structures, including the compact region as well as the extending fibrils, is characteristically different for the various elements of the karyotype. Chromosome 2 displays the largest terminal structure, whereas chromosome 4 only occasionally shows the presence of compact regions. — End to end association of the long chromosome arms involves the fusion of the compact terminal structures. The non-random distribution of end to end association seems to be correlated with the volume of the terminal structures. Chromosome 2 which contains the largest compact terminal region is more frequently involved in end to end associations than any other chromosome arm. — The terminal regions show replication of DNA. They belong to the group of regions which display a discontinuous labeling pattern along the chromosomes, representing a late phase of the replication cycle. — The unique structural organization of the terminal chromosome regions, which is never observed at any other location of the genome supports the idea that they are morphological manifestations of the postulated telomeres.

Notes: Abstract Vitamin B6 induces in vivo as well as in vitro the appearance of a puff at region 2–48C in Drosophila hydei. At concentrations of 10−2 M or lower, region 2–48C is the only region responding to vitamin B6 provided that oligomycin is present in the incubation medium. Pyridoxal phosphate and pyridoxamine phosphate supplied to medium containing oligomycin induce upon incubation of salivary glands a larger puff at 2–48C. — Puff 2–48C produces large quantities of a unique RNP-product; globular 140–220 Å particles which aggregate to stable complexes of 0.1–0.2 μ in diameter. Upon continuous in vitro incubation with vitamin B6, puff 2–48C becomes loaded with these aggregates which have never been observed in any other puff of Drosophila hydei.

Notes: Abstract Puparium formation in Drosophila lebanonensis casteeli is obviously restricted to a certain phase in circadian oscillation. The question whether or not the release of molting hormone is the actual process which is controlled by the circadian oscillation could be approached by using molting hormone-specific changes in genome activity as indication for changes in hormone titer. The identification of hormone specific changes in the puffing pattern of polytene chromosomes should provide a basis for this study.—To this end, a chromosome map of the 7 polytene chromosome arms (1 acrocentric and 3 metacentric chromosomes) of the species was made. Changes in the puffing pattern associated with puparium formation are described and compared with those occurring in response to experimental administration of β-ecdysone.—89 puffs were regularly observed in midthird instar larvae. Prior to puparium formation 5 new puffs arise, one at an early stage and 4 attaining their maximum size immediately before puparium formation. Concomitantly, 5 puffs increase considerably in size. These changes in the puffing pattern can be reproduced by injection of ecdysone.—Upon injection of the hormone a clear differentiation between fast reacting loci (within 30–60 min) and slow reacting loci (after 3–4 hours) can be found. As in other Drosophila species the immediate response (within 30–60 min) comprises more than one (5) locus.