Thesis (M.S.)--University of Wisconsin--Madison, 1962. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaf 13).

Conditional lethals which survive under "permissive" conditions but die under "restrictive" conditions have greatly facilitated genetic and biochemical analyses in micro-organisms. One class of conditional lethals, the so-called temperature-sensitives, has been recovered from ethyl methanesulfonate-induced lethals in Drosophila melanogaster. Such mutants survive at 17°C or 22°C but die at 29°C and appear to map genetically as point mutants. These mutants may be useful in identifying previously undetected loci and in studying mitotic, meiotic and developmental processes. In Drosophila gross developmental studies indicate that recessive temperature-sensitive (ts) lethals may have specific developmental effects. By means of shifting different cultures from one temperature to the other at successive intervals, the period during which the restrictive temperature (29°C) prevented survival could be ascertained. Each ts lethal could be classified with respect to its effective lethal phase(s), its period(s) of temperature-sensitivity and any effect on visible morphological characteristics. Four sex-linked recessive ts lethals were studied extensively with respect to their genetic and developmental action. Each of these mutants is representative of a different class of lethals with specific properties: pupal lethality, indispensability, sexual dimorphism, and parental influence. The potential use of these and other ts lethals in genetic and developmental analyses is discussed. / Science, Faculty of / Zoology, Department of / Graduate

Reported here is the isolation and molecular characterization of two novel alleles of the DIP 1 gene; GE89 and GE77. As well, a third deletion of the DIP 1 gene, EY*4, isolated by our collaborators in France was characterized. PCR and sequencing analysis confirms all three alleles to be molecular deletions of the DIP 1 gene. However, in none of these cases is the entire gene excised. Also, immunohistochemistry of ovaries from each of these strains does not demonstrate a complete lack of DIPl protein expression in any of the deletion strains. Thus, it appears that some protein product is being formed in each case. However, it is not clear whether this protein is functional. An assay was also conducted to investigate a function for DIPl in mechanisms of epigenetic gene silencing. Although the findings of these experiments are incomplete, it appears that DIPl may play a functional role in heterochromatin formation and/or post-transcriptional gene silencing. Interestingly, appendage formation phenotypes were observed in the original P-element insertion line as well as a female sterility phenotype in the GE77 allele. Overall, DIP 1 is an interesting double stranded RNA binding protein. Newly isolated alleles of the DIP 1 gene will be useful tools for further investigation of the functional role of this gene. / Thesis / Master of Science (MSc)

The midline glia (MG) perform an active role in the pioneering and morphogenesis ofthe commissural axons of the central nervous system (CNS) during Drosophila embryogenesis. Following the establishment of these commissural axon pathways, a subset of the MG undergoes apoptosis and the surviving MG remain to ensheath the commissures. Previous studies demonstrated that the pattern ofthis developmental apoptosis is stochastic. The surviving MG exhibit intersegment variability in their final number and position relative to the commissures. Interestingly, argos, a gene which encodes a diffusible extracellular factor with an epidermal growth factor (EGF) motif, is only expressed in MG which survive. argos expression in this subset of MG is initiated at the onset of apoptosis, reflecting the temporal pattern of cell death of the other MG. In argos loss-of-function mutants, there are extra surviving MG in each segment while ectopic over-expression of argos results in increased apoptosis among the MG. Therefore, Argos has a negative regulatory effect on MG survival. Its effects are opposite to the spitz group/Drosophila EGF receptor (DER) pathway, a cassette of genes required for MG survival. However, argos is hypostatic to the spitz group/DER pathway function and its expression requires a certain threshold of spitz group and DER pathway activity. Argos is postulated to act as an EGF receptor antagonist. It attenuates signaling of the DER pathway. Therefore, the regulation of MG survival is mediated by a balance of extracellular inductive and inhibitory signals. How can this signaling pathway be reconciled with the stochastic pattern of MG survival and the observation that only argos expressing MG survive? We propose a model in which the adherens junction along with possible accessory proteins like Rhomboid and Star mediate the close apposition of MG and promote intense Spitz mediated DER pathway signaling. The MG with the most concentrated coupling of this adhesion and signaling pathway network achieve sufficient DER signaling to express argos. These MG are in tum fated to assume a final differentiated identity and will remain to ensheathe the commissures. Argos is secreted and it mediates the apoptosis of MG with reduced levels of adhesion contacts and lower DER signaling (MG not expressing argos). The other mesectodermal cell (MEC) lineages appear to develop independently of argos function. The repercussions of HSargos elicited MG loss to the commissural axons and CNS cytoarchitecture was also examined. Removal of the MG through ectopic expression ofargos results in a loss of ensheathment and the commissural axon tracts are exposed to the haemolymph. Furthermore, the commissural axons are wider and they misexpress Fasciclin II, a phenotype reminiscent of mutations in roundabout. / Thesis / Master of Science (MSc)

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abstract: Anoxia tolerance is strongly correlated with tolerance to heat, desiccation, hyperosmotic shock, freezing, and other general stressors, suggesting that anoxia tolerance is broadly related to stress tolerance. Age affects the capacity of many animals to survive anoxia, but the basis to this ontogenic variation is poorly understood. We exposed adult Drosophila, 1, 3, 5, 7, 9, and 12 days past eclosion, to six hours of anoxia and assessed survival 24-hours post-treatment. Survival of anoxia declined strongly with age (from 80% survival for one-day-old flies to 10% survival for 12 day-old-flies), a surprising result since adult fly senescence in Drosophila is usually observed much later. In anoxia, adenosine triphosphate (ATP) levels declined rapidly (< 30 min) to near-zero levels in both 1 and 12-day old adults; thus the higher anoxia-tolerance of young adults is not due to a better capacity to keep ATP elevated. Relatively few physiological parameters are reported to change over this age range in D. melanogaster, but gut bacterial content increases strongly. As a partial test for a causal link between bacterial load and anoxia tolerance, we replaced food daily, every third day, or every sixth day, and assayed survival of six hours of anoxia and bacterial load at 12 days of age. Anoxia tolerance for 12-day old flies was improved by more food changes and was strongly and negatively affected by bacterial load. These data suggest that increasing bacterial load may play an important role in the age-related decline of anoxia tolerance in Drosophila. / Dissertation/Thesis / Masters Thesis Biology 2020

Malpighian (renal) tubules of the fruit fly Drosophila melanogaster consist of three functional domains: a non-secretory distal segment, a secretory main segment and a reabsorptive lower segment. In this study a computer-controlled micropositioning system and a self-referencing K^+ microelectrode were used to measure K^+ concentration gradients of extracellular unstirred layers associated with specific epithelial domains. K^+ fluxes were calculated from the measured gradients. This is the first time: that an accurate assessment of the concentration gradients of the unstirred layer of Drosophila melanogaster could be assessed due to the enhanced sensitivity of this self-referencing technique over conventional ion-selective microelectrodes. The technique permits high resolution spatial and temporal mapping of the flux patterns in response to stimulation or inhibition of ion transport. Variations in K^+ transport over time and at different sites suggest that transport is non-uniform within any one functional domain. / Thesis / Master of Science (MSc)

HSP27, the Drosophila homolog of mammalian HspB1, is a nuclear sHsp that is both stress induced and developmentally regulated with a conserved cyto-protective function. It is multiply phosphorylated in vivo through an unconfirmed mechanism at unidentified residues. The effect of phosphorylation on its localization, oligomerization, and function is also not well understood. Here we report a genetic investigation into the role of Hsp27 in Drosophila development, and a preliminary investigation into the effect of phosphorylation on HSP27 localization and function in Drosophila S2 cells. Through a proteomic screen, a pro-apoptotic role for Hsp27 in embryonic developmentally regulated programmed cell death was suggested and supported by RNAi experiments, but not replicated using Hsp27null mutant stocks. These stocks were complicated by the intriguing appearance of multiple background mutations. Specific developmental defects in transgenic lines overexpressing phospho-mutant isoforms were then investigated. These too were subject to multiple independent incidences of background genetic mutation, which we believe may be related to Hsp27 mis-expression. We also studied the endogenous expression and localization pattern of HSP27 in stressed and unstressed Drosophila S2 cells. We found evidence that wild-type protein localization is influenced by stress. Finally, we took a first step toward understanding how phosphorylation might regulate HSP27 localization by examining the effect of targeted mutations of serine residues (S58, S71, and S75) on the localization pattern of exogenous HSP27. By characterizing the expression of endogenous and overexpressed HSP27 in Drosophila cells, we provide a foundation for future investigation into the regulated localization and function of HSP27 that can be extended to address the regulatory mechanisms that govern the protective capacities and oligomeric properties of phosphorylated HSP27 in Drosophila.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2006. / Härtill 4 uppsatser.