A high-resolution morphogenetic map of the second-leg basitarsus inDrosophila melanogaster

Summary The lineages of cells on the second-leg basitarsus ofDrosophila melanogaster were analyzed by examining gynandromorphs andMinute mosaics. Bracts lie proximal to bristles on the adult basitarsus, yet bract precursor cells were found to originate lateral to bristle precursor cells. In 6 of the 8 longitudinal rows of bristles on this segment, the bract cells arise ventral to the bristle cells; in the others they arise dorsally. The lateral cell origins are interpreted as reflecting a pattern of lateral cell movements associated with evagination of the leg disc. An unusual discrepancy was observed in the relative frequencies of male vs. female bracts and bristles in gynandromorphs. The discrepancy suggests that there is a cell-autonomous sexual difference in either the time at which cells begin moving during evagination or the speed with which they move.On the basis of the results, it is reasoned that the bristle pattern of the basitarsus does not originate in its final form. Prior to evagination, the bristle cells of each row are apparently closer together than in the final pattern, and the rows are farther apart. Evidence is presented which suggests that the bristle cells of each row may originally be arranged in a jagged line which is later straightened by cell movements.The two locations where the anterior/posterior compartment boundary of the second leg passes through the basitarsus were found to vary relative to the bristle pattern. If this boundary is assumed to be a fixed line of positional values, then the extent of the observed variability — which is estimated to be ± 1 or 2 cell diameters — provides a measure of the precision of patterning around the circumference.     

Pattern formation inDrosophila melanogaster: The effects of mutations on polarity in the developing leg

Summary The effects of the mutations eyeless dominant (ey ^D) and shibire (shi) on bristle pattern in the legs ofDrosophila melanogaster were examined. Both mutations cause gaps in the intersegmental membranes which separate leg segments and often alter the position of these membranes. It was observed that pattern disturbances including reversed bristle polarity and duplication of structures such as sex combs and transverse rows were associated with defects in the intersegmental membranes. The alterations in bristle polarity and most of the duplication of structures could be accounted for by a segmentally reiterated gradient in the legs which controls bristle polarity and which requires the integrity of the intersegmental membrane. A computer simulation of this gradient model was devised which accounted for the observed results. The possible role of cell death as a cause of the gaps in the intersegmental membrane and of some of the pattern disturbances was examined.     

Updating the Trachelocercids (Ciliophora, Karyorelictea). I. A Detailed Description of the Infraciliature of Trachelolophos gigas N. G., N. Sp. and T. filum (Dragesco & Dragesco-Kernéis, 1986) N. Comb

ABSTRACT. Trachelolophos gigas n. g., n. sp. and T. filum (Dragesco & Dragesco-Kernéis, 1986) n. comb. (basionym: Tracheloraphis filum) were discovered in the mesopsammon of the French Atlantic coast at Roscoff. Their morphology and infraciliature were studied in live and protargol impregnated specimens. The new genus, Trachelolophos, belongs to the family Trachelocercidae and is unique in having a conspicuous ciliary tuft, which is very likely a highly modified brosse, in the oral cavity. The two species investigated have a very similar infraciliature, differing only in morphometric characteristics and in the nuclear configuration. The entire somatic and oral infraciliature consists of dikinetids which have both basal bodies ciliated or only the anterior or posterior ones, depending on the region of the cell. The right side is densely and uniformly ciliated. Its kineties extend onto the left side to the glabrous stripe, where an anterior and posterior secant system are formed, reducing the number of kineties in the narrowed neck and tail region. The left side bears a narrow glabrous stripe bordered by slightly irregularly arranged dikinetids having rather stiff cilia (bristles), possibly forming an uninterrupted, prolate ellipsoidal (bristle) kinety as indicated by their ciliation. The bristle kinety commences subapically at the right margin of the glabrous stripe, extends posteriorly, then anteriorly at the left, to end up at the right margin again. The dikinetids of the right posterior portion of the bristle kinety have the posterior basal bodies ciliated, whereas the anterior basal bodies are ciliated in its left and right anterior portion. The ends of the bristle kinety meet distinctly subapically at the right margin of the glabrous stripe, as indicated by the diametrically opposed ciliation of the dikinetids. The anterior region (head) of the cell bears a distinct circumoral kinety composed of very regularly arranged dikinetids, associated with nematodesmata forming an oral basket together with the nematodesmal bundles originating from the oralized somatic dikinetids at the anterior end of the somatic kineties. The systematics of trachelocercid ciliates are briefly reviewed and discussed.     

Arrangement of bristles as a function of bristle number on a leg segment inDrosophila melanogaster

Summary The arrangement of bristles on a leg segment of the fruitflyDrosophila melanogaster was studied in various mutants that have abnormal numbers of bristles on this segment. Eighteen mutations at six different genetic loci were analyzed, plus five double or triple mutant combinations. Recessive mutations at theachaete-scute locus were found to affect distinct groups of bristles:achaete mutations remove mechanosensory bristles, whereasscute mutations remove mainly chemosensory bristles. Mechanosensory bristles remain uniformly spaced along the longitudinal axis unless their number decreases below a certain threshold, suggesting that spacing is controlled by cell interactions that cannot function when bristle cells are too far apart. Above a certain threshold, bristle spacing and alignment both become irregular, perhaps due to excessive force from these same interactions. Chemosensory bristles occupy definite positions that are virtually unaffected by removal of individual bristles from the array. Extra chemosensory bristles develop only near the six normal sites. At two of the six sites the multiple bristles tend to exhibit uniform longitudinal spacing — a property confined to mechanosensory bristles in wild-type flies. To explain the various mutant phenotypes the following scheme is proposed, with different mutations directly or indirectly affecting each step: (1) spots and stripes are demarcated within the pattern area, (2) one bristle cell normally arises within each spot, multiple bristle cells within each stripe, (3) incipient bristle cells inhibit neighboring cells from becoming bristle cells, and (4) the bristle cells within each stripe become aligned to form rows and then repel one another to generate uniform spacing.     

scabrous Modifies Epithelial Cell Adhesion and Extends the Range of Lateral Signalling during Development of the Spaced Bristle Pattern in Drosophila

The role of scabrous (sca) in the evenly spaced bristle pattern of Drosophila is explored. Loss-of-function of sca results in development of an excess of bristles. Segregation of alternately spaced bristle precursors and epidermal cells from a group of equipotential cells relies on lateral inhibition mediated by Notch and Delta (Dl). In this process, presumptive bristle precursors inhibit the neural fate of neighbouring cells, causing them to adopt the epidermal fate. We show that Dl, a membrane-bound ligand for Notch, can inhibit adjacent cells, in direct contact with the precursor, in the absence of Sca. In contrast, inhibition of cells not adjacent to the precursor requires, in addition, Sca, a secreted molecule with a fibrinogen-related domain. Over-expression of Sca in a wild-type background, leads to increased spacing between bristles, suggesting that the range of signalling has been increased. scabrous acts nonautonomously, and we present evidence that, during bristle precursor segregation, Sca is required to maintain the normal adhesive properties of epithelial cells. The possible effects of such changes on the range of signalling are discussed. We also show that the sensory organ precursors extend numerous fine cytoplasmic extensions bearing Dl molecules, and speculate on a possible role for these structures during signalling.     

The fine structure of developing bristles in wild type and mutant Drosophila melanogaster

In an attempt to understand the factors involved in morphogenesis of a complex cell like a scale or bristle, the fine structure of the normal development of bristle cells in Drosophila melanogaster (Oregon R) has been studied and compared with that of the mutants sn³ and Sb. In the development of the normal bristle rounded bundles of longitudinally oriented fibrils lie just beneath the cell surface at regularly spaced intervals. Fiber bundles constitute about 20% of the cross sectional area. The cytoplasmic surface between these bundles is active in enveloping the nerve fiber associated with the bristle and in sending out cytoplasmic processes associated with which the longitudinally oriented bristle ridges form. Singed bristles are bent and twisted and the fiber bundles are present as flattened bands constituting only about 5% of the cross-sectional area. In Sb mutants the total cross-sectional area of fiber bundle material is the same as that in Oregon R, but fiber bundles are smaller and more numerous, being distributed over the larger surface of this thicker and shorter bristle. They constitute only 7% of the cross-sectional area of the bristle. In Sn³Sb mutants characteristics of each gene are exaggerated and an extremely short, wide, and irregular bristle is formed.     

Patterned Arrays of Intramembranous Particles in the Bristle Cilia of Three Different Species of Euplotes1

Freeze-fractured cells of three marine species of Euplctes (E. crassus, E. raikovi, and E. rariseta) show bristle cilia with patterned arrays of intramembranous particles. Such arrays are essentially of three types, in different positions along the bristle shaft. One array is located near the bristle base and shows a plate-like shape. It appears in a close spatial correspondence with the lasiosome network, which is a structure consisting of interconnected electron-dense bodies lying in between the peripheral axonemal doublets and the bristle membrane. The second type of array, apparently typical of only E. raikovi, consists of eight to ten longitudinal rows of particles that occupy most of the intermediate portion of the bristle. The third type of array appears differently shaped in different species and occurs at the bristle apex.     

Regional differences in the developing foreleg of Drosophila melanogaster

We transplanted imaginal disks of Drosophila melanogaster from larvae of the second half of the third larval instar into prepupae. Disks from the youngest donors differentiated bristles of only the distal segments of the leg. These disks also produced unusually large areas of cuticle that had no bristles. Disks from older donors differentiated bristles of more proximal segments and the area of cuticle with no bristles was reduced. To account for the regional variation in these results, there must be regional differences among the prospective leg cells at some time during the period from the second half of the third larval instar to the end of adult bristle differentiation. We asked whether prospective distal cells were more advanced than prospective proximal cells during bristle differentiation. We estimated when bristle precursor cells undergo their final cell divisions by heavily irradiating prepupae and pupae. We assumed that cells that were insensitive to the radiation had completed their cell divisions. The distal segments were the first to have insensitive bristles. Most leg bristles became insensitive between 12 and 18 hr after pupariation. The tarsus had a larger proportion of its bristles insensitive than the femur at 15 hr after pupariation. We also investigated when bristle-forming cells begin elongating their bristle shafts. We used the length of bristle rudiments as an indicator of when elongation is initiated. At 35 hr after pupariation, bristle rudiments of distal segments were two to three times longer than bristle rudiments of proximal segments. We discuss how these intersegmental differences observed during bristle differentiation can account for the regional variation in response of discs transplanted into older hosts. However, we do not exclude the possibility that regional differences among cells of the leg tissue exist at stages earlier than the time of bristle differentiation.     

The action of mitomycin C on the bristle-forming apparatus ofPhormia regina

Summary Mitomycin C, a known inhibitor of DNA synthesis, was injected into white prepupae ofPhormia regina, Adults which developed from these prepupae showed alterations of the bristle pattern, loss of whole bristle organs, and the formation of bristles without sockets or sockets without bristle shafts. Dose-dependence was found for all modifications. For the abdominal microchaetae, the period of maximum sensitivity to the drug began at 16 h after puparium formation, that is well after all of the macrochaetae and most of the microchaetae of the thorax and the head had grown insensitive. Bristle forming trichogen and tormogen cells developed high degrees of polyteny with distinctly banded chromosomes. Photometric determination of the amount of Feulgen-DNA per nucleus led to estimations of DNA classes ranging from 256C to 2048 C. DNA contents of nuclei from Mitomycin C treated animals were significantly lower during the actual growth of the bristle apparatus, but reached approximately the same level as the controls prior to the time of emergence. Cytological investigations proved that doses of Mitomycin C which yielded bristle organs either without sockets or without shafts do not affect the differential division of the bristle mother cell. Polytene chromosomes damaged by Mitomycin C displayed a diffuse and irregular banding pattern. Possible modes of action of Mitomycin C on replicating polytene chromosomes are discussed.     

Autonomous puffing patterns in thoracic and abdominal polytene bristle cell chromosomes of the flesh fly Sarcophaga barbata

The puffing patterns of the thoracic and abdominal polytene bristle cell chromosomes were investigated in Sarcophaga barbata during a 10-day period of pupal development. The autonomous differentiation of imaginal disk descendants is visualized microscopically at the chromosomal level by the cell autonomous puff activities of the polytene bristle cell chromosomes. The sequence of chromosomal activities is strictly stage specific in both cell types. The changes in the puffing pattern are closely corelated with development. The puffing pattern changes synchronously in all bristle cells of a certain body region, e.g., the scutellum or the fifth abdominal tergit. However, there is no synchrony between the puffing pattern changes of the thoracic and abdominal bristle cells. The loci of the abdominal bristle cells are activated one day later than those of the thoracic cells. Each particular puffing pattern truly represents a particular developmental state of the bristle, regardless of body location. That is, the bristle cell chromosomes of various body segments control the timing of their puffing activities autonomously and puff formation and puff regression are not hormonally synchronized.     

Fine structure of the dorsal bristle complex and pellicle of Euplotes

The fine structure of the dorsal bristle complex and pellicle of non-developing Euplotes eurystomus is described in detail by scanning and transmission electron microscopy. The bristle-pit unit is a highly differentiated complex of organelles. The bristle complex is composed of a pair of kinetosomes (basal bodies) joined by a connective. The anterior kinetosome bears the bristle cilium, which contains a polarized network of particles (“lasiosomes”). The posterior kinetosome bears a very short, knob-like “condylocilium,” and has an associated striated fiber. Accessory ribbons of microtubules are also associated with the kinetosome couplets. Parasomal sacs, a septum connecting the bristle cilium to the anterior wall of the pit, core granules of the kinetosomes, and large membranous ampules are described. The organization of the bristle complex bears many similarities to the somatic ciliature of other ciliates. The pellicle of Euplotes is composed of a continucus outer cell membrane subtended by membranous alveoli, which contain a “fibrous mat.” Two sheets of subpellicular microtubules (longitudinal and transverse) are located just beneath the alveoli. The “epiplasm” seen in some other ciliates is apparently absent in Euplotes. The texture of the cell surface is a pattern of folds or rugae composed of the outer cell membrane and the upper membrane of the alveolus. The pattern of rugae probably defines the “silverline-system” of light microscopy.     

Sensitive periods for abnormal patterning on a leg segment inDrosophila melanogaster

Summary The development of a leg segment of the fruitflyDrosophila melanogaster was analyzed in order to determine whether the orderliness of the segment's bristle pattern originates via waves of cellular interactions, such as those that organize the retina. Fly development was perturbed at specific times by either teratogenic agents (gamma rays, heat shock, or the drug mitomycin C) or temperature-sensitive mutations (l(1)63, l(1) Notch^ts1, orl(1) shibire ^ts1 ), and the resulting abnormalities (e.g., missing or extra structures) were mapped within the pattern area. If bristles develop in a linear sequence across the pattern, then they should show sensitivity to perturbations in the same order, and wavefronts of cuticular defects should result. Contrary to this prediction, the maps reveal no evidence for any directional waves of sensitivity. Nevertheless, other clues were uncovered as to the nature and timing of patterning events. Chemosensory bristles show earlier sensitivities than mechanosensory bristles, and longer bristles precede shorter ones. The types and sequence of cuticular abnormalities imply the following stages of bristle pattern development: (1) scattered inception of bristle mother cells, each surrounded by an inhibitory field, (2) alignment of the mother cells into rows, (3) differential mitoses, (4) assignment of cuticular fates to the mitotic progeny, (5) polytenization of the bristle cells, (6) fine-tuning adjustments in bristle spacing, and (7) signalling from bristle cells to adjacent epidermal cells, inducing them to form bracts.     

The emergence of order in the Drosophila pupal retina

During pupation, long-range order is imposed on the autonomously developing ommatidia which compose the Drosophila eye. To accomplish this, eight additional cell types arise: the primary, secondary, and tertiary pigment cells, and the four cells that form the bristle. These cells form an interweaving lattice between ommatidia. The lattice is refined when excess cells are removed to bring neighboring ommatidia into register. Recent evidence suggests that in larval development, local contacts direct cell fate. The same appears to be true during pupal development: the contacts a cell makes predict the cell type it will become. Cells which contact the anterior or posterior cone cells in an ommatidium invariably become primary pigment cells. Cells which contact primary pigment cells from different ommatidia become secondary and tertiary pigment cells. Bristle development is in several ways distinct from ommatidial development. The four cells of each bristle group appear to be immediate descendents of a single founder cell. During their early differentiation, they do not make stereotyped contacts with surrounding ommatidial cells, but do make particular contacts within the bristle group. And unlike the surrounding ommatidia, differentiation of the bristles radiates from the center of the eye to the edges. As cells are removed during two stages of programmed cell death, the bristles are brought into their final position. When all cells in the lattice have achieved their final position, a second stage of retinal development begins as structures specific to each cell type are produced. This paper follows these various stages of pupal development, and suggests how local cell-cell contacts may produce the cells needed for a functional retina.     

Identification of a 41 kDa Protein Embedded in the Biosilica of Scales and Bristles Isolatedfrom Mallomonas splendens (Synurophyceae, Ochrophyta)

Cells of the photosynthetic protist Mallomonas splendens (Synurophyceae, Ochrophyta) are encased within a highly patterned wall or scale case that consists of silicified scales and bristles. In an effort to understand the mechanisms that unicellular protists utilize to produce elaborate, mineralized structures of great complexity and hierarchical structure, we identified and characterized a 41 kDa protein from purified scales/bristles isolated from M. splendens (SP41 for Scale Protein of 41 kDa). A cDNA encoding this protein was isolated and sequence analysis indicated that it is a novel protein. Polyclonal antibodies were generated against bacterially expressed SP41 and used to localize the protein throughout scale and bristle morphogenesis. Immunoelectron microscopy confirmed the biochemical data that SP41 is a component of mature scales and bristles, the protein localizing to silicified components of the purified extracellular matrix. During scale and bristle biogenesis within the cell, SP41 is deposited into a specialized Silica Deposition Vesicle (SDV) concomitant with silica deposition, a highly regulated event during scale and bristle formation. These results argue for SP41 playing a role in morphogenesis and/or silicification within the SDV during scale and bristle biogenesis.     

Post-mating effects in the grasshopper, Gomphocerus rufus L. mediated by the spermatheca

In the female grasshopper Gomphocerus rufus mating replaces copulatory readiness with immediate and long-lasting `secondary defense', during which further mating attempts are efficiently repelled. The behavioral change is caused by secretions from the male accessory glands' white secretory tubule 1 which is injected with the spermatophore material into the female's spermathecal duct. A bristle field of contact chemoreceptors at the entry of the spermathecal duct into the endbulb is assumed to be stimulated by the secretion. Ablation of the bristle field, interruption of the nervous pathway between the spermatheca and the ventral nervecord, or severance of the latter sustains sexual receptivity after mating. Both the secretion from white secretary tubule 1 and the spermatophore contained in the spermatheca of a mated female are digested by proteolytic enzymes from spermathecal gland cells. Dissolved material is resorbed by similar glandular-like cells. The intersexual conflicts of interest and their evolutionary consequences are discussed. Accepted: 4 December 1998     

Ultrastructure of the phallic glands of the firebrat,Thermobia domestica (packard) (Thysanura : Lepismatidae)

The exocrine glands located in the penis of Thermobia domestica (Thysanura : Lepismatidae) are composed of about 100 distinct units, each containing several cell types: one large secretory cell with an apical reservoir; 2 groups of envelope cells, an inner group of 2 superimposed cells, and an outer group of 4 cells arranged in a ring, and also 2 basal cells, called ciliary cells owing to their elongated processes, which look like the dendrite of a sensory cell. Each functional unit includes cuticular differentiations: a tubular bristle, fixed on a small tubercle; and a long “internal” ductule communicating basally with the reservoir of the glandular cell and opening distally at the tip of the bristle. A study of the modifications affecting the phallic glands during moulting, shows that the inner envelope cells deposit the cuticle that forms the ductule, the outer envelope cells elaborate the cuticule of the tubercle, while a temporary distal projection of only one of these cells ensures the formation of the bristle. In addition, a lengthening of the outer dendritic segment of the 2 ciliary cells takes place before ductule formation, but this segment partially degenerates after ecdysis. These findings are compared with data already obtained on the morphogenesis of other insect integumental glands. In T. domestica, the secretion of the phallic glands is presumed to be used, during the mating sequences, for spinning fine threads before spermatophore deposition.     

The Behavior and Fine Structure of the Dorsal Bristles of Euplotes minuta,E. aediculatus,and Stylonychia mytilus (Ciliata,Hypotrichida)1

ABSTRACT. Studies of the bristle (dorsal) cilia of Euplotes minuta. E. aediculatus, and Stylonychia mytilus by light and electron microscopy indicate that these cilia do not beat metachronously in any of the species. The bristle cilia in Stylonychia may beat actively, but those in Euplotes stand erect or are bent in different directions with the flow of water. The duration and degree of bending appear correlated with the duration and velocity of the water current. The fine structure of the bristle complex is similar in both Euplotes species and like other reports of Euplotes in the literature. The complex consists of paired kinetosomes, the anterior bearing a short cilium containing four to six rows of fibrous balls (lasiosomes) oriented along the anterior surface of the axoneme, the posterior lacking a cilium but with a small cap. Microtubular ribbons are associated with the paired kinetosomes, and a collar with a pronounced alveolar ring underneath the pellicular membrane tightly surrounds the cilium at the opening of the bristle pit. The bristle complex in S. mytilus differs from that of Euplotes and other hypotrichs in that it has a single kinetosome in interphase cells and, attached to the kinetosome, a prominent fibrous structure (parakinetosomal body). Microtubules are attached to the parakinetosomal body. As in Euplotes, the bristle unit is surrounded by mucocyst-like organelles (ampules). Observations of behavior and fine structure suggest that the dorsal bristles may be sensory, perhaps responding to stimuli from water currents, although other functions are possible, too.     

A balance of capping protein and profilin functions is required to regulate actin polymerization in Drosophila bristle

Profilin is a well-characterized protein known to be important for regulating actin filament assembly. Relatively few studies have addressed how profilin interacts with other actin-binding proteins in vivo to regulate assembly of complex actin structures. To investigate the function of profilin in the context of a differentiating cell, we have studied an instructive genetic interaction between mutations in profilin (chickadee) and capping protein (cpb). Capping protein is the principal protein in cells that caps actin filament barbed ends. When its function is reduced in the Drosophila bristle, F-actin levels increase and the actin cytoskeleton becomes disorganized, causing abnormal bristle morphology. chickadee mutations suppress the abnormal bristle phenotype and associated abnormalities of the actin cytoskeleton seen in cpb mutants. Furthermore, overexpression of profilin in the bristle mimics many features of the cpb loss-of-function phenotype. The interaction between cpb and chickadee suggests that profilin promotes actin assembly in the bristle and that a balance between capping protein and profilin activities is important for the proper regulation of F-actin levels. Furthermore, this balance of activities affects the association of actin structures with the membrane, suggesting a link between actin filament dynamics and localization of actin structures within the cell.     

SECRETION AND DEPLOYMENT OF BRISTLES IN MALLOMONAS SPLENDENS (SYNUROPHYCEAE)1

Mallomonas splendens (G. S. West) Playfair has a cell covering of siliceous scales and bristles. Interphase cells bear four anterior and four posterior bristles that each articulate, at their flexed basal ends via a complex of labile fibers (the fibrillar complex), on a specialized body scale (a base-plate scale). Body scales, base-plate scales and bristles are formed independently of each other and at different times in silica deposition vesicles (SDVs) that are associated with one of the two chloroplasts. The fine structure of scale and bristle morphogenesis in M. splendens agrees with that previously described for Synura and Mallomonas. Four new posterior bristles are formed at late interphase with their basal ends towards the cell posterior. The fibrillar complex is formed in situ on the bristle in the SDV. Mature bristles are secreted one by one onto the surface of the protoplast, beneath the layer of body scales, where the basal ends of the bristles adhere to the plasma membrane via the fibrillar complex. The extrusion of posterior bristles and their deployment onto the cell surface was monitored with video. A fine cellular protuberance accompanies the bristles as they are extruded from beneath the scale layer with their basal ends leading. When distant from the cell, the basal ends of the bristles appear attached to the protuberance, possibly by way of their fibrillar complexes. Once bristles are fully extruded, and their tips free in the surrounding environment, the bristle bases are drawn back to the posterior apex of the cell, apparently by the now shortening protuberance. Thus a 180° reorientation of the posterior bristles has been effected outside the cell. Thin-sections of cells that are extruding bristles show a threadlike, cytoplasmic extension of the cell posterior which may be analogous to the protuberance seen in live cells. Four new posterior base-plate scales are secreted after the bristles have reoriented. Scanning electron microscopy indicates that the fibrillar complex is involved in positioning the bristles onto their respective base-plate scales. Anterior bristles are formed in new daughter cells in the same orientation as the posterior bristles; thus they are extruded tip first and no reorientation is required.