A series of experiments were conducted to observe the effect of various environments on the duration of life of wild-type Drosophila melanogaster. Natural duration of life curves were prepared for these insects from 10.5 days of age and it was found that the average life expectancy of the males was greater than that of the females. A 10ˉ⁴ M concentration of the mutagen acridine orange in the medium markedly reduced the average life span of Drosophila males and females. The 10ˉ⁵ M concentration reduced the male life expectancy slightly but had no effect on the females. A method utilizing a separatory funnel type population container was developed to test the effect of a chemical stress on Drosophila populations. Some preliminary experiments were performed in order to evaluate the effect of 10ˉ³ M 2, 4-dinitrophenol, a metabolic inhibitor, combined with various food environments on life duration. The sexes differed in their response to these environments and the relative durations of life of the flies were compatible with a priori considerations. From the results a non-linear relationship is suggested between the time of 50% population death and the rate of death. Starvation death curves were compiled for the Drosophila after they were exposed to various conditions which were thought to be representative of the natural environment. Following exposure to various media conditions, female Drosophila under starvation conditions outlived their male counterparts. The difference between the starvation life expectancies of the females and males tended to become greater after the insects had been kept under optimum conditions, due predominantly to an increase in female life duration. Utilizing a combination of a rapid 'quantitized' heat-shock and starvation conditions it was found that only the male starvation life duration was significantly decreased compared with the unshocked starved controls. The female starvation life duration did not appear to be significantly decreased by the heat-shock. Under starvation conditions wholly female populations had a life duration similar to female populations mixed with an equal number of males. Some explanation is given for the results obtained in this thesis and recommendations for further experiments are made. / Science, Faculty of / Botany, Department of / Zoology, Department of / Graduate

The sex-linked Notch locus plays an important role in embryogenesis and determination of many adult structures of the fruit fly, Drosophila melanogaster. Mutation at this locus can cause lethality in embryonic or later stages, as well as morphological abnormalities of the adult eyes, wings, bristles and legs. Alleles of the Notch locus can be broadly grouped into three classes: 1) recessive lethal Notch (N) alleles, which may be deficiencies or point mutations, 2) Abruptex (Ax) alleles, which are probably point mutations and may be either lethal or viable, and 3) viable recessive alleles with visible phenotypes, which affect a variety of traits and are point mutations. The present investigation was initiated with a view to understanding the relationships between N and Ax alleles and the nature of their role in development, and has consisted mainly of the following approaches: 1) an examination of the phenotypes of certain unusual N alleles and the phenotypic responses to alteration of the dosage of these alleles in relation to wild-type (N+), 2) an examination of the interaction of Ax alleles with N alleles and with one another, and 3) developmental studies of the conditional (temperature-sensitive) phenotypes associated with certain Notch-locus genotypes. The results of the N-allele dosage study indicate that a single mutation in the Notch locus can affect different functions associated with this locus in fundamentally different ways. Depending on the genotype and phenotype examined, the responses of various N alleles to dosage changes suggest that mutation at the Notch locus may result in reduced, increased or novel activity at the locus. Four ethyl methanesulfonate-induced Ax alleles have been examined, none of which is cytologically abnormal in salivary gland chromosome preparations, and at least three of which map within the Notch locus. Depending on culture conditions and the alleles involved, Ax/N heteroallelic combinations may be viable or lethal. All Ax/N combinations studied exhibited less severe Abruptex phenotypes (bristle loss and wing vein gapping) than the respective Ax/Ax homozygotes. However, the Ax alleles differed from one another in their effects on the wing nicking of the N alleles, in that the viable allele Ax9B2 and the semi- lethal allele AxE1 both suppressed wing nicking, whereas the two viable alleles AxE2 and AX16172 both enhanced wing nicking. Furthermore, heteroallelic combinations of Ax alleles which affected nicking in different direction, were lethal (AxE1/AxE2, AxE1/Ax16172, Ax9B2/Ax16172), whereas combinations of Ax alleles with similar effects on nicking were viable (AxE1/Ax9B2, AxE2/ Ax16172). The temperature-shift experiments have revealed an interesting pattern of temperature-sensitive periods (TSPs) for lethality or adult morphological abnormalities associated with various Notch-locus genotypes. TSPs for lethality may be monophasic occurring in the embryo (N60g11/N6g11;Dp51b7), or the second larval instar (Ax16172/n264-40), or they may be polyphasic, occurring in embryo, larval and pupal stages (N264-103/fano). On the other hand, the TSPs for all the adult morphological abnormalities examined occur during the third larval instar, including rough eyes and wing nicking (N60gll/+,N264-103/spl), leg segment fusion (N264-103/+, N264-103/spl), wing vein gapping (Ax16172/+) and disturbance of bristle numbers (N264-103/spl, Ax16172/+). Several molecular models are discussed in relation to the observations on N-allele dosage and interactions of the Ax and N alleles. The results are consistent with the hypothesis that the Notch locus is a regulator gene influencing many developmental processes, that mutations can affect the activity either of the entire gene or of various parts of the gene individually, and that N and Ax mutations usually affect this regulatory system in opposite ways from one another. / Science, Faculty of / Zoology, Department of / Graduate

Conditional lethal mutants which die under restrictive conditions but are viable in a permissive environment provide useful tools for the genetic and developmental analysis of certain loci. One class of conditional mutants, temperature-sensitive recessive lethals which die at 29°C but survive at 23°C has been reported for an extensive number of loci in Drosophila melanogaster. However, loci may exist which are highly redundant, or concerned with functions requiring the total output of both wild type alleles or with synthesis of a structural component which would pre-exempt their ready detection as recessive mutations. Mutations within such regions could, however, be recovered by selecting temperature-sensitive mutations which behave as dominant lethals at the restrictive temperature. In addition, such mutants would permit the first genetic characterizations of dominant lethals. Ethyl methanesulfonate-induced dominant temperature-sensitive lethal and semi lethal mutations were induced in chromosome 2. Twenty-one lethals of this type were isolated from 6,130 tested chromosomes and sixteen were characterized with respect to their genetic localization and developmental effects. Unexpectedly, eleven of the mutants were found to be closely linked to the dumpy, dp, locus. All eleven were recessive lethals at room temperature and were functionally allelic. The temperature-sensitive period (TSP) was similar for all cluster mutants although the effective lethal phase (LP) at 29°C differed. It was concluded that the cluster lethals are, in fact, genetically allelic. Three other loci were demonstrated by genetic recombination and each had a characteristic TSP and LP. In addition, two mutants caused sterility of females and could not be localized. The recovery of mutations which map genetically within a segment and are dominant lethals, proves that dominant lethality need not reflect gross chromosomal alterations in higher organisms. / Science, Faculty of / Zoology, Department of / Graduate

The antibiotic, Mitomycin-C (MC), increases recombination in Drosophila females, yet has no effect on crossing over in males. This sexual difference in recombinagenic response to MC cannot be attributed to a sex difference in permeability to MC, since MC is mutagenic in both sexes. MC increases the frequency of exchange events involving X and Y chromosomes as well as those involving the two arms of the Y in both sexes. On the basis of brood analysis it was found that those cells which are most sensitive to the recombinagenic and X-Y and Y-Y exchange effects of MC were in a premeiotic stage of gametogenesis at the time of treatment. It is proposed that the apparent inability of MC to induce crossovers in males is related to the normal absence of spontaneous meiotic crossing over in males. MC can act as a monofunctional or as a bifunctional alkylating agent which primarily reacts with guanine residues of DNA. Bifunctional alkylation results in the crosslinking of the two strands in the DNA double helix. It is postulated that prior to meiosis, alkylated guanine residues are excised from the DNA molecule through the action of natural repair mechanisms. Depending on whether mono or bifunctional alkylation has taken place, either single strand nicks or double strand cuts will be induced in the DNA. Double strand cuts may be repaired as a mutation or as a non-reciprocal exchange event whereas single strand nicks might act as the natural pre-condition for genuine meiotic crossovers and could thus form crossovers when the necessary "crossover substances" are present. This model predicts that an alkylating agent such as ethyl methanesulfonate (which alkylates guanine monofunction-ally, and therefore would give rise to single strand nicks following excision), should be recombinagenic in females, but not in males but should not effect the frequency of X-Y and Y-Y exchange events in either sex. This prediction was indeed demonstrated thus supporting the model. / Science, Faculty of / Zoology, Department of / Graduate

Cold-sensitive mutants of E. coli have been valuable in the study of the structure and assembly of bacterial ribosomes. Some cold-sensitive mutations of E. coli have been mapped to loci coding for ribosomal proteins. Other cold-sensitive mutations have been shown to alter the regulatory properties of bacterial enzymes. In contrast, little was known about the genetics and biochemistry of cold-sensitive mutants of Drosophila and other eukaryotes. Also, the genetic loci of ribosomal proteins of D. me1anogaster were unknown although the ribosomal RNA genes have been located on the X and 2nd chromosomes. Therefore the following questions were asked: Can cold-sensitive lethals of the X chromosomes of Drosophila melanogaster be isolated? If so, what are their genetic properties? Do some have altered ribosomal proteins as in cold-sensitive mutants of bacteria? The study was part of a general search for and characterization of cold-sensitive lethals of all the chromosomes of D. melanogaster. Among 3,919 EMS-treated X chromosomes, 25 were retained as cold-sensitive lethals or semi-lethals. That is, more than 20% of the flies carrying a cold-sensitive lethal survived at 22°C and none at 17°C, and for cold-sensitive semi-lethals, > 30% survived at 22°c and <13% at 17°c. The cold-sensitive mutations were not randomly distributed, 7 being located at the X tip and 3 being alleles to the right of car. Over half exhibited female sterility at the permissive temperature and 7 exhibited visible phenotypes characteristic of bobbed and Minute mutations. As the presence of altered ribosomal proteins has been demonstrated in cold-sensitive mutants of bacteria, the ribosomal proteins from Drosophila mutants and controls were studied by 2-dimensional gel electrophoresis. By this method, it was estimated that 69-72 proteins are present in the ribosomes of Drosophila. No qualitative differences were observed in the gel patterns of ribosomal proteins from 5 cold-sensitive, sex-linked mutants and the controls. A summary of the biochemical testing of the ribosomes from other cold-sensitive mutants of Drosophila is also given. No biochemical evidence to support the hypothesis that cold-sensitive mutants of Drosophila have altered ribosomes was obtained. The limitations of the study and recommendations for future research are discussed. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

Undergraduate honors paper--Mount Holyoke College, 2006. Dept. of Biological Sciences. / Includes bibliographical references (leaves 75-82).

Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 157-170).

No description available.

Courtship and copulation behaviors in Drosophila melanogaster males are regulated by sex-specific products from the gene fruitless (fru). Male-specific FRU proteins (FRU[superscript M]) are putative transcription factors of the BTB-ZnF family that likely act by controlling development and maintenance of the neural circuitry used during male sexual behavior. However, which neuronal characteristics are regulated by FRU[superscript M] is mostly unknown and how FRU[superscript M] neurons are grouped into circuits and the role that specific neuronal circuits play in sexual behavior has not been elucidated. I have identified a subset of FRU[superscript M] neurons that co-express the transcription factor, Engrailed (En). After fru[superscript M]-RNAi-induced targeted removal of FRU[superscript M] proteins from FRU[superscript M]/En neurons, males were impaired in their ability to initiate or maintain copulation. Further, I examined two characteristics, the initial projections and neurotransmitters used by FRU[superscript M]/En neurons. Males and females showed a difference in the neurochemistry of FRU[superscript M]/En neurons in the thoracic ganglia; this neurochemistry is disrupted in fru mutant males. For one cohort of serotonergic neurons in the abdominal ganglion that were previously shown to be dependent on FRU[superscript M] for expression of serotonin, I determined that FRU[superscript M] works in conjunction with other sex-specific genes, TAKEOUT (TO) and DOUBLESEX (DSX), to induce of serotonin expression in males; in females serotonin expression is repressed by DSX and TO. Finally, I performed a genetic screen for genes that interact with, or are downstream targets of, fru, dsx, or dissatisfaction (dsf). I assessed fertility, copulation success, and abdominal muscle development of EMS-mutagenized flies, resulting in one fly line in which homozygous mutant animals had a novel muscle phenotype. By genetic tests, the mutation was found to be allelic to string, which encodes a Cdc25- like phosphatase. Taken together, my research demonstrates that subsets of FRU[superscript M] neurons function in circumscribed circuits to regulate specific portions of sexual behavior, and that FRU[superscript M], along with other sex-specific genes, controls development of these neurons in part by determining neurochemistry. Further, FRU[superscript M] likely directs multiple downstream targets, in different subsets of neurons in which it is expressed, which collectively provide correct development of neural circuits underlying courtship and copulation behavior. / Graduation date: 2006

The fruitless (fru) gene in Drosophila melanogaster is a multifunctional gene having sex-specific functions in the regulation of male sexual behavior and sex-nonspecific functions affecting adult viability and external morphology. While much attention has focused on fru's sex-specific roles, little is known about its sex-nonspecific functions. The embryonic central nervous system (CNS) is a prime model system in which to study the genetic control of axonal outgrowth and proper CNS formation. I have examined fru's sex-nonspecific role in embryonic neural development. fru transcripts and FRU proteins from sex-nonspecific promoters are expressed beginning at the earliest stages of neurogenesis and subsequently in both neurons and glia. In embryos that lack most or all fru function, Fasciclin II- and BP102-positive axons appeared to defasciculate from their normal pathway and fasciculate along aberrant neuronal pathways, suggesting that one of fru's sex-nonspecific roles is to regulate axonal differentiation. I next examined whether the loss of fru function in FRU-expressing neuronal precursors causes neuronal fate change. Analysis of fru mutant embryos revealed a lack of Even-skipped (Eve) staining in Eve-expressing neurons, ectopic Eve staining in non-Eve-expressing neurons and mispositioned dorsal Eve-expressing neurons, which suggests that fru functions to maintain neuronal identity rather than to specify neuronal fate. In fru mutants these defects in axonal projections and in Eve staining were rescued by the expression of specific fru transgenes. To better understand fru's function in the formation of the embryonic CNS, I dissected out fru's function in neuron and glia through a genetic interaction study. fru genetically interacts in neurons with longitudinal lacking to make proper axonal projections. In addition, fru might be in the same genetic pathway as roundabout (robo), a repulsive guidance receptor, and commissureless, a downregulator of Robo, to ensure proper axonal pathfinding. Surprisingly, fru interacts with tramtrack and glial cells missing to repress neuronal differentiation in the lateral glia and with single-minded for the development of midline glia. Taken together, fru function is required for proper axonal pathfinding in neurons and for proper development of lateral and midline glia. / Graduation date: 2002