Behavior of Somatic Cells Homozygous for Zygotic Lethals in DROSOPHILA MELANOGASTER

The behavior in genetic mosaics of 86 EMS-induced sex-linked lethals has been studied. Seventy-five percent of them are autonomous in gynandromorphs. Forty-three lethals nonviable in sex mosaics have been analyzed in X-ray-induced spots in the abdominal tergites and the imaginal wing derivatives. Of the lethals, 90.7% are homozygous viable in mosaic spots, and only 9.3% have been classified as epidermal cell lethal. Thus, the fraction of the Drosophila genome essential for cell viability has been estimated to be about 420 genes. The phenotypes at the cellular level of some cell-viable mutations altering cell parameters (mitotic orientation, differentiation, etc.) are described.     

Pattern formation in the imaginal discs of a temperature-sensitive cell-lethal mutant of Drosophila melanogaster

To explore the effects of cell death on pattern formation in the developing imaginal discs of Drosophila melanogaster, I have isolated a number of cell-autonomous temperature-sensitive lethal mutants. Sex-linked temperature-sensitive lethals were screened for cell-autonomy by scoring the survival of lethal-bearing clones in genetic mosaics. The mutant with the strongest effect on clone viability gave rise to a high frequency of structural deficiencies and duplications in the derivatives of the eye-antennal discs, when subjected to pulse-treatments at the nonpermissive temperature during the late second and third instars. The patterns produced were nonrandom, with some structures showing a tendency to become deficient, and others a tendency to duplicate. Duplicated structures were only found in heads in which other structures were missing. Genetic tests identified the lethal as a point mutation at the suppressor-of-forked locus. Recombination, and complementation tests with a small duplication of this region showed that a second mutational lesion is in all probability not involved in the generation of abnormal patterns in the imaginal discs. It is therefore proposed that the cell-lethal action of the mutant is sufficient to account for phenotypic effects described. According to this hypothesis, cell death primarily causes deficiencies, and duplications occur as a response of the discs to injury. In agreement with this, it was found that in gynandromorphs, pattern duplications can be found in wild-type tissue in the presence of lethal tissue in the same disc. Thus, a cell-autonomous lethal may affect the process of pattern formation in a nonautonomous way.     

Competition between cleavage nuclei in Drosophila

Distribution of tissues of XX and XO genotype in gynandromorphs resulting from elimination of unstable ring X-chromosome during initial cleavage divisions was studied. Predominantly XX-nuclei proved to give rise to cell nuclei in tissues of cranial and caudal regions of mosaics. XX nuclei are proposed to be more migrationally active than XO nuclei.     

Polymorphism of M factors in populations of the housefly, Musca domestica L., in Turkey

M factors, which determine maleness in Musca domestica, were found on the second, third, fourth and fifth linkage groups in housefly populations of Turkey. As in European populations, the male-determining factor was more frequently located on linkage group III (MIII). Some males homozygous or double heterozygous for M factors were identified. Deviations from a 1:1 sex ratio in favour of males, as well as mosaics for somatic marker mutations and sexual mosaics (gynandromorphs), were also observed. The results reveal an extensive polymorphism in the sex-determining system.     

On the Interpretation of Mutagenically Induced Mosaicism in Drosophila

This paper draws attention to the formal parallelism that exists between chromosomal-loss mosaicism and mutagenically induced mosaicism in Drosophila and suggests that, although the underlying processes by which these two types of genetic mosaics are generated are very different, the more refined methodology employed in developmental analyses of genetically induced chromosomal-loss mosaics may be profitably extrapolated to mutagenesis studies. Results obtained from various studies of genetically induced mosaics and from a previous EMS mutation induction experiment at the yellow locus are utilized to illustrate this methodology and to estimate the total mutagenicity rates of EMS.-The following are some of the tentative conclusions that have been drawn in this report regarding an EMS concentration that produced 31% F(2) lethals in the standard X-linked recessive lethal test: (1) The frequency of cuticular mosaics is at least 5 times that of F(3) lethals. (2) At least 60% of all cuticular mosaics go undetected in the standard X -linked recessive lethal test since their mutant tissue does not extend into the germ line. (3) The frequency of EMS-induced cryptic mosaics is probably less than 10% the frequency of cuticular mosaics. (4) Some EMS-induced mutations are probably bona fide completes; if confirmed, this inference must be taken into consideration in estimating the total mutagenicity rates of this agent and in molecular interpretations of its mechanism of action. (5) The fact that the proportion of mutant tissue in EMS-induced mosaics is greater than 25% is consistent with the suggestion that the action of EMS is occasionally delayed until after the first cleavage division of the embryo. (6) Such an EMS concentration causes on the average more than 5 independent genetic alterations in the entire haploid genome of an X-bearing sperm.-This report clarifies the experimental evidence that must be generated, and the methodology that can be used to analyze this evidence, if it is of interest to render these and related conclusions regarding the effect of EMS on D. melanogaster more accurate, or if it is of interest to conduct a similar analysis for other mutagens that cause a significant degree of mosaicism.     

Identification of Brain Sites Controlling Female Receptivity in Mosaics of DROSOPHILA MELANOGASTER

We have identified cells in the brain of Drosophila melanogaster that are required to be of female genotype for receptivity to copulation with males. To do this, we determined experimental conditions in which female flies virtually always copulate, then measured the minimum amount of male courtship that is required to stimulate females to indicate their receptivity to copulation. We then observed gynandromorphs with female genitalia to determine whether the sex mosaics elicited at least the minimum amount of courtship and, if so, whether they copulated. By analyzing these gynandromorphs, in which the genotype of external and internal tissues could be ascertained, we were able to identify a group of cells in the dorsal anterior brain that, when bilaterally female, is necessary and sufficient for receptivity to copulation. This group of cells is anatomically distinct from those that are required to be of male genotype for the performance of courtship behaviors.     

Temperature-sensitive autosomal dominant lethal mutations in Drosophila melanogaster induced by ethylmethane sulfonate

Cold- and heat-sensitive dominant autosome and recessive sex-linked lethals were scored using C(1)RM, y; vg bw; e ss tester stock. The frequencies of heat-sensitive mutations were 1.43, 0.30, 0.07% and of cold-sensitive ones were 0.39, 0.16 and 0.09% in the 1-st, 2-nd and 3-rd chromosomes, respectively. For the first time, dominant cold-sensitive lethals were obtained in chromosome 3. The data from genetic analysis point to the fact that penetrance of such mutations strongly depends on the genetic background. That may be the reason, why they were not obtained using some of the balancer-3 chromosomes. Also, "cryptic" dominant autosome mutants were found which were not conditional but only revealed in the F2 generation. Their possible origin as gonado-somatic mosaics is discussed.     

Developmental Analysis of the Achaete-Scute System of DROSOPHILA MELANOGASTER

The interpretation of the wild-type function of a gene depends on our knowledge of the phenotype caused by its absence. We have first defined the genetic extent of the achaete-scute system by studying the phenotype of different terminal and intercalary deficiencies including these genes. When these deficiencies were lethal, we have defined the phenoeffective phase of lethality and studied their phenotype in genetic mosaics (gynandromorphs and mitotic recombination clones). The achaete-scute system affects two functions, one necessary for the differentiation of the embryonic (central?) nervous system and the other necessary for the differentiation of peripheral nervous elements of the chaetes and sensillae of the adult cuticle. The possibility that these functions correspond to differential expression of a single mechanism is discussed.     

Mutagenicity of hycanthone in Drosophila melanogaster

The schistosomicidal agent hycanthone was tested for mutagenicity in Drosophila melanogaster. The compound was administered either by injection into adult males or by larval feeding. The following types of genetic damage were measured:(1) complete and mosaic sex-linked recessive lethal mutations; (2) II–III translocations; and (3) dominant lethals.In postmeiotic germ cells, especially in late spermatids, a pronounced increase was found in the frequency of sex-linked recessive lethals, both completes and mosaics. By contrast, translocations and dominant lethals were not induced.     

Irregularities in the early development of the Drosophila Melanogaster egg

Summary Polyspermy is normally present in the Drosophila egg.Extreme polyspermy may lead to disturbances which may prevent the egg from further development.Polar body formation may be disorganized by supernumerary sperms.Supernumerary sperms may form unipolar, bipolar and multipolar spindles.In rarer cases sperms will enter into normal spindles and form multipolar spindles. In some cases such interference seems to adjust itself to the original bipolar condition and division figures with the polyploid number of chromosomes will result. Either by the conjugation of two sperms, or by dimegaly, or by the doubling of chromosomes of a single sperm, supernumerary nuclei may develop. The origin of certain mosaics and gynandromorphs may be explained on this basis.     

Control of Female Reproduction in Drosophila: Genetic Dissection Using Gynandromorphs

The sexual behavior of Drosophila melanogaster gynandromorphs was studied to analyze the relationship between different steps in the female reproductive pathway. It was assumed that, in some gynandromorphs, certain female functions are missing because the corresponding control sites (foci) are either composed of male tissue or did not develop. A given gynandromorph can show elements of both male and female reproductive pathways. None of the steps of the female reproductive pathway appeared to be dependent on any other, in contrast to male behavior where, for example, following of females is a prerequisite for attempted copulation. By correlating each of the behaviors with the genotype of the cuticle, we confirmed previous findings that the focus for the female sex appeal is located in the abdomen, but receptivity to copulation is controlled by a site in the head. Many of the gynandromorphs did not lay eggs, presumably because either the focus controlling egg transfer from the ovaries to the uterus or the one controlling egg deposition was composed of male tissue. Many of the nonovipositing gynandromorphs laid eggs while dying or could be induced to deposit eggs after implantation of hormone-producing glands or topical application of a juvenile hormone analog. Some of the noninseminated gynandromorphs laid eggs at the rate characteristic for inseminated females, suggesting that an oviposition focus (mapping in the head region) suppresses oviposition in virgin females, but not in gynandromorphs whose focus is composed of male tissue. Some of the inseminated gynandromorphs oviposited eggs at a low rate, possibly because the focus responsible for detection of insemination could not function properly. Some of the inseminated gynandromorphs laid unfertilized eggs, revealing the importance of the focus controlling sperm release from the seminal receptacle. Foci controlling egg transfer, egg deposition and sperm release are located in the thorax, according to mosaic fate mapping results and studies on the reproductive behavior of decapitated females. The location of egg deposition in the culture vial seems to be controlled by a brain site. Sexual behavior in Drosophila does not depend on the presence (or absence) of the ovary or germ line.     

Genetic analysis of mutations at the Glued locus and interacting loci in Drosophila melanogaster

A genetic analysis of the dominant mutation Glued that perturbs the development of the normal axonal architecture of the fly's visual system was undertaken. Ten new alleles at this locus were identified and characterized. Two complementation groups that were identified failed to complement the original allele, suggesting that it is a double mutant or that it resides at a complex locus. Several of the new alleles display visual-system abnormalities similar to those of the original mutation. Seven of the eight members of one complementation group are embryonic/early larval lethals, like the original mutation. The other allele in this group is temperature sensitive. Homozygous mutant adults exhibit a temperature-sensitive female sterile phenotype. Unsuccessful attempts to recover genetic mosaics carrying clones of cells homozygous for some of these mutations revealed that they are either essential for the viability of individual cells or that they affect some other fundamental cellular function, such as mitosis or the ability to participate in tissue level organization, which prevents them from being recovered in adult mosaics. This also indicates that these mutations do not specifically affect neural cells. A number of X-ray- and EMS-induced partial and complete phenotypic "revertants" of the original allele have also been isolated as material for a comparative analysis of visual system development. All "revertants" that alter the abnormal eye phenotype towards the wild type have similar impact on the organization of the optic lobe.     

Increment kinetics of recessive lethal mutations and dominant lethals in offspring of Drosophila melanogaster on storage of methyl-methanesulfonate-treated sperm in females

The frequencies of sex-linked recessive lethal mutations in F1 males after feeding adult male Drosophila melanogaster with 0.25 and 0.5 mM methyl methanesulfonate (MMS) orally for 24 h increased approximately linearly with storage of the treated spermatozoa in females, whereas the number of hits of dominant lethals in the sperm after feeding 0.3 and 0.5 mM MMS increased approximately with the square of the storage time. Chromosome losses and mosaics in F1 males also increased with the dose of MMS to males, but their yields were too low to be analyzed quantitatively, only indicating a slight increase of chromosome loses and a slight decrease of mosaics with the time of storage of sperm. Maternal non-disjunctions (or chromosome losses), detected in F1 males, decreased with the dose of MMS to spermatozoa and their yield decreased with the time of storage of sperm of both MMS-treated and the control groups. A unitary model is proposed to explain the effect of storage on the dominant lethals and recessive lethal mutations.     

Control of Male Reproductive Behavior by the Central Nervous System of Drosophila: Dissection of a Courtship Pathway by Genetic Mosaics

In gynandromorphs of Drosophila, a detailed examination was made of the association between male courtship behavior and the chromosomal genotype of various parts of the central nervous system. Mosaic flies that behave as males repeatedly show a shorter courtship than normal males. If there is to be male behavior, the posterior dorsal brain must be haplo-X on at least one side for occurrence of the early courtship events. Tapping, following of females and wing extension. Licking (proboscis extension) has nearly the same focus but is submissive; that is, male tissue must be present in both left and right dorsal brain. The next courtship step, attempted copulation, has a focus (especially for actual genital contact) located in the thoracic ganglia, though apparently not in a discrete region. Attempted copulation, which can occur even in mosaics with a gravid abdomen, may be correlated with the presence of sex combs. The role of courtship foci are interpreted in terms of known sensory inputs to and functions of the major insect ganglia.     

Transplantation of nuclei into the polar plasm of Drosophila eggs

Nuclei of preblastoderm and blastoderm stages of yw strain of Drosophila melanogaster were transplanted into posterior ends of fertilized eggs of the v; bw strain. Several mosaics were obtained, including gynandromorphs with external genitalia of the donor's type. One mosaic was fertile, with gonads derived from the transplanted preblastodermic nuclei. Genetic analysis showed that the genome of these nuclei was not affected by differentiation or by transplantation.     

A male-specific lethal mutation in Drosophila melanogaster that transforms sex.

Sex-lethal, male-specific allele #1 (Sxl^M#1, 1–19.2) is a dominant, X-linked mutation that is lethal to males. It has no effect in females other than to rescue them from the otherwise lethal maternal effect of the autosomal mutation, daughterless. From a study of the effects of Sxl^M#1 on the development of sexually mosaic flies (gynandromorphs), it was observed that this lethal mutation can cause genetically male (haplo-X) tissue to differentiate as if it were female. With respect to its effect on sexual differentiation, the mutation transformer (tra, 3–45) is epistatic to Sxl^M#1, though the lethal effects of Sxl^M#1 are not modified by tra. In addition to its effect on sexual differentiation, Sxl^M#1 reduces the size of haplo-X imaginal disc and histoblast derivatives in general in a cell autonomous fashion. The viability of gynandromorphs with Sxl^M#1 tissue is very low, and the surviving mosaics have relatively little haplo-X tissue, suggesting that there is no single localized “lethal focus” for this mutation. The relationship between Sxl and daughterless is discussed, as well as the possible involvement of the Sxl locus in X-chromosome dosage compensation.     

Preblastoderm mosaics of mutants of the bithorax-complex

Summary InDrosophila melanogaster, segmental specification takes place in groups of cells around the blastoderm stage. This segmental specification requires the function of the genes of the bithorax-complex. We have studied preblastoderm mosaics (gynandromorphs) of mutant (bx ³,pbx, Ubx, Ubx ⁸⁰) and wildtype (heterozygotes for these alleles) cells. The results show a total cell autonomy in the differentiation of both wildtype and homoeotially transformed cells. However, several unexpected phenotypes were found. They are discussed in terms of the function of the bithorax genes and early interactions between mutant and wildtype territories.     

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.     

Cell lineage restrictions in the genital disc ofDrosophila revealed byMinute gynandromorphs

Summary The genital imaginal disc ofDrosophila differentiates the terminalia, i.e. the genitalia and analia, of both sexes. It represents a composite anlage, containing a female genital primordium, a male genital primordium and an anal primordium. In normal males and females, only one of the two genital primordia differentiates; the other is developmentally repressed. Therefore, cell-lineage relationships between the male and female genital primordia can only be studied in sexual mosaics which differentiate female and male cells. We producedMinute (M)non-Minute(M⁺) gynandromorphs and selected those with sexually mosaic terminalia for a cell-lineage analysis. In these mosaics, either the male (XO) or female (XX) cells wereM ⁺ and thus had a growth advantage. The differential growth rates served as a tool to detect clonal restrictions. In control gynandromorphs (M ⁺M ⁺), the amount of female genitalia differentiated was largely independent of the amount of male genitalia present. In contrast, male and female anal structures, as a rule, added up to one full set. The same was true for the experimentalMM ⁺ gynandromorphs, but the contribution ofXX andXO cells to mosaic terminalia changed drastically due toM ⁺ cells competing successfully against the more slowly growingM cells. Specific subsamples ofMM ⁺ gynandromorphs showed thatM cells in a non-mosaic primordium are shielded from cell competition taking place in the neighbouring mosaic primordium. We conclude that the three primordia of the genital disc represent developmental compartments. In the genital primordia, even developmentally repressedM ⁺ cells compete successfully against developmentally activeM cells.     

Mosaic Analysis Gives an Estimate of the Extent of Genomic Involvement in the Development of the Visual System in Drosophila Melanogaster

To investigate the role of vital loci in the development of the visual system of Drosophila, we induced mitotic recombination in individuals heterozygous for recessive organismal lethals and selected for analysis the resulting mosaics with homozygous mutant eye clones. Heads bearing clones were serially sectioned, silver-stained and examined for aberrations in the ommatidia and the neural structures to which they project. In our screen of 68 lines bearing diepoxybutane-induced X-linked lethals, 26 yielded few or no homozygous mutant clones (putative cell-lethals). Of the rest, 20 lines produced individuals with morphologically abnormal eye clones showing various degrees of aberrations in the ommatidial architecture. In 14 of these 20, the laminar cartridges innervated by the mutant clones were also disorganized. Clones with normal structure were found in 18 of the lines, and three lines were resistant to the induction of mitotic recombination. In a single line, comparatively normal clones in the eye projected to a lamina with subtle but consistent abnormalities. To the extent that we have a representative sample, these results suggest that about two-thirds of all vital genes may be essential for the normal assembly and neural connectivity of the eye. This points to a high degree of pleiotropy in the manner in which information in the genome of the fly is used in development.