MSc Exit Seminar
Tuesday November 14th, 2:30 pm – Earth Sciences Centre, Rm. 3087
Alexa Chioran (Ringuette lab)
“SPARC promotes proper nucleation and assembly of collagen IV from Drosophila melanogaster embryonic haemocytes”
Basement membranes (BMs) are a hallmark feature of all multicellular organisms in which they support essential developmental events. Collagen IV is the major structural component of the BM, however, its spatiotemporal assembly into stereotypic lattice networks is poorly understood. SPARC (Secreted Protein Acidic and Rich in Cysteine), a collagen IV-binding glycoprotein, is enriched in the BMs of Drosophila embryos and larvae. Fat body cells of Drosophila larvae and egg chamber follicular epithelial cells of the adult deficient in SPARC expression exhibit a fibrotic-like accumulation of collagen IV and other BM components in the pericellular space, resulting in larval lethality. Thus, SPARC is hypothesized to be an extracellular chaperone-like protein, whereby its association with collagen IV prevents aberrant collagen IV polymerization. In this study, I utilized molecular markers distinguishing secretory pathway compartments to demonstrate that SPARC and collagen IV co-localize primarily in the trans-Golgi network prior to secretion. Moreover, knockdown of SPARC expression in cultured embryonic haemocytes supports a role for SPARC in preventing premature nucleation of collagen IV molecules in embryonic haemocytes adherent to a laminin matrix. In order to gain further mechanistic insight on the functional relationship between SPARC and collagen IV, CRISPR gene editing technology was used to generate a sparc-null mutant. This genetic tool will be used in future experiments to unmask the structural domains of SPARC responsible for collagen IV assembly, maturation, and BM homeostasis.
MSc Exit Seminar
Wednesday, September 27, 2017 at 10:10 am, Ramsay Wright Building, Room 432
Cassandra Marie D’Amata (Tropepe Lab)
Characterizing the role of ice1 in maintaining zebrafish neural stem cells
Maintenance of neural stem cell (NSC) niches is required for the continued growth of the zebrafish retina and forebrain after embryogenesis. The zebrafish mutant kess564, which maps to the ice1 locus, exhibits reduced NSC niches. RNA polymerase II-dependent snRNA transcription requires the little elongation complex (LEC) for which ICE1 is an essential scaffolding component. Mutant NSCs which are normally active no longer express markers of cycling cells and become apoptotic. Furthermore, quiescent NSCs of the mutant retina are unable to proliferate in response to UV lesion. Whole-transcriptome analysis of ice1 mutant larvae show a downregulation of CNS and cell cycle genes, and an upregulation of splicing genes indicative of a possible compensatory mechanism. snRNA production appears to be unaffected in a subset of NSCs but reduced in differentiated neurons. This work demonstrates that ice1 is essential for NSC maintenance in an in vivo loss of function model.
MSc Exit Seminar
Monday, September 25, 2017 at 11:10 am, Ramsay Wright Building, Room 432
Stefan Vujadinovic (Tepass Lab)
The Okapi (Oka) FERM Domain Protein Regulates Somatic Stem Cell Numbers and Cell Intercalation during Drosophila Oogenesis
Okapi (Oka) is a Drosophila melanogaster FERM domain protein that is essential for oogenesis. oka (CG34347) null mutants are viable but display multiple defects during oogenesis. Mutant ovaries showed greatly enlarged and multilayered interfollicular stalks, compromised follicular epithelia, and germline disorganization, leading to reduced female fertility. I generated both endogenously and exogenously GFP-tagged Oka protein to examine the distribution of Oka.
Oka was present is all somatic cells of the ovary and localized to adherens junctions (AJs) in the germarium and follicles, including the cap cells, follicular epithelium, intercalating cells that form the interfollicular stalk, and in the migrating border cell cluster and centripetal cells. I also found that oka mutant ovaries contain an increased number of follicle stem cells, a defect that could explain the increased somatic cell numbers observed, and is reminiscent of the phenotypes of several overactive signalling pathways such as Hippo and Hedgehog. Examination of a series of five oka alleles that display different phenotypic strength suggest that interfollicular stalk formation is more sensitive to the reduction in Oka function than other observed defects. Together, my findings suggest that Oka is a component of AJs that negatively regulates follicle stem cell numbers and is required for the intercalation of interfollicular stalk cells.