MSc Exit Seminar - Shafinaz Eisha (McGowan Lab)
Effect of Maternal High Fat Diet and Milk-Derived Exosome Supplementation on DNA Methyltransferase 1 in the Brain
Abstract
Maternal high fat diet (mHFD) exposure during perinatal life shapes offspring’s neurodevelopment trajectories. mHFD also influences the bioactive composition of maternal milk, including milk-derived exosomes (MDEs), nanovesicles that transport functional microRNAs (miRNA). miRNA-148/152 regulate DNA methyltransferase-1 (DNMT1), enzyme that catalyzes DNA methylation modifications, which in turn downregulates fat-mass and obesity-associated (Fto) gene via promoter hypermethylation. The main objectives of this thesis were: 1) examine the effects of mHFD exposure on DNMTs and Fto expressions in postnatal day 7 rats and 2) investigate MDE-supplementation effects on DNMTs and Fto expressions using a human microglia cell line. DNMT1 and Fto transcript abundances remained unchanged in the PVN and ventral hippocampus but decreased in the retroperitoneal fat of mHFD offspring. In microglia, MDE-supplementation resulted in increased DNMT1 expression only. These findings indicate that DNMT1 and Fto exhibit tissue-specific sensitivity to mHFD exposure in neonates, and that DNMT1 is altered by MDE-supplementation in microglia.
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https://utoronto.zoom.us/j/89443033929
Meeting ID: 894 4303 3929
Host: Patrick McGowan (patrick.mcgowan@utoronto.ca)
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MSc Exit Seminar - Alexander Sullivan (Provart Lab)
GAIA: An “eEcosystem” of Aggregated Information for Plant Biology
Abstract
Currently, there is a plethora of scientific data available online through many different tools, databases, repositories, and websites, making it difficult for researchers to find the information for their prospective investigation. When the Multinational Arabidopsis Steering Committee (MASC) conducted a 2018 bioinformatics survey, they found that researchers have expressed a strong desire to have data centralized. Inspired by this request, the General Agricultural Intelligent Agent (GAIA) was created. Aggregating data for plant biology across many scientific resources, GAIA can create an executive summary that can either summarize a gene or gene product of interest or use Natural Language Processing (NLP) and machine reading (MR) to answer an inputted question. Built with modular design philosophy, GAIA is created with simplicity in mind for its code, allowing new developers to easily access, modify, and create new components for its UI and functionality. GAIA is currently available at https://bar.utoronto.ca/gaia.
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https://utoronto.zoom.us/j/85953765984
Meeting ID: 859 5376 5984
Host: Nicholas Provart (nicholas.provart@utoronto.ca)
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MSc Exit Seminar - Erin Hunt (Harris Lab)
SCAR/WAVE and Arp2/3 Complexes Associate with a Supra-Cellular Actomyosin Cable in Response to Myosin and a Cytohesin Arf-GEF
Abstract
Expansive Arp2/3 actin networks and contractile actomyosin networks were once viewed as antagonistic assemblies with spatially and temporally distinct distributions within the cell. More recently, Arp2/3 and actomyosin networks were found to closely interact during cell migration, adhesion and epithelial remodeling. However, molecular mechanisms responsible for coordinating the networks remain unclear. Here, I show that the Arp2/3 complex and its activator, the SCAR/WAVE complex, are enriched at cell-cell junctions of a supra-cellular actomyosin cable found at the leading edge (LE) of epidermal sheets during dorsal closure (DC) of the Drosophila embryo. Myosin activity is both necessary and sufficient for these accumulations of the SCAR/WAVE complex. Myosin has previously been shown to promote localization of the cytohesin Arf-GEF Steppke (Step) to these sites. Strikingly, I find that Step is required for the enrichment of the SCAR/WAVE complex. Partial SCAR loss does not dramatically alter LE actin enrichment, as might be expected, but does result in LE shape irregularities. Genetic interactions between step mutants and those of SCAR and Arp3 further indicate a role for the SCAR/WAVE and Arp2/3 complexes in DC. My data suggests myosin activity acts through Step to recruit the SCAR/WAVE and Arp2/3 complexes to promote actin network growth that may be important for actomyosin cable function during DC.
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https://utoronto.zoom.us/j/85682403987
Meeting ID: 856 8240 3987
Host: Tony Harris (tony.harris@utoronto.ca)
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MSc Exit Seminar- Zobia Anwar (Rhee Lab)
Investigating the Role of Onecut1 in Establishment of Cell-type-Specific Enhancers during Mouse Motor Neuron Differentiation
Thursday, October 22nd, 2020 at 2:00pm
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https://utoronto.zoom.us/j/81258699707
Meeting ID: 812 5869 9707
Passcode: 148248
Abstract:
Enhancers are short regulatory DNA sequences that act to ‘enhance’ associated gene transcription. The immense complexity and diversity within the central nervous system provides an excellent opportunity to study the role of enhancers during cell fate specification. The identity of spinal motor neurons is established by three programming transcription factors (TFs), also known as lineage-determining TFs: Ngn2, Isl1 and Lhx3. Two LIM homeodomain TFs, Isl1 and Lhx3, bind to enhancers in nascent motor neurons regulating the expression of many effector genes, but Lhx3 is downregulated as nascent motor neurons mature. Then, Isl1 is relocated to the enhancers bound by Onecut1 TF in maturing motor neurons ensuring stable motor neuron gene expression. The objective of this thesis was to characterize the role of Onecut1 in maintenance of gene expression in maturing spinal motor neurons. I employed the CRISPR-Cas9 system to generate embryonic stem (ES) cell line containing Onecut1 knockout mutation, followed by in-vitro differentiation of these ES cells into spinal motor neurons. I performed analyses for gene expression changes and genomic mapping in wildtype motor neurons and differentiated cells from the Onecut1 knockout mutant. My gene expression analysis revealed attenuation of expression of a subset of motor neuron genes in Onecut1 mutant motor neurons. Chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) analysis showed a global redistribution of Isl1 binding and active enhancers in the absence of Onecut1. These results suggest that Onecut1 plays a role in the maintenance of Isl1-bound enhancers linked to motor neuron genes. Together, my analyses revealed that Onecut1 is associated with the expression of a subset of neuronal genes by binding with Isl1 at motor neuron enhancers.
MSc Exit Seminar -Claresta Adityani (Lumba/McCourt Lab)
Characterizing Striga hermonthica Gibberellin Receptors and Investigating their Crosstalk with Strigolactone Receptors
Wednesday, September 30th, 2020 at 2:00pm
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https://utoronto.zoom.us/j/92018414970
Meeting ID: 920 1841 4970
Passcode: 255133
Abstract:
Gibberellic acids (GAs) are a group of phytohormones known to stimulate germination of land plants. However, a root parasitic plant called Striga hermonthica (Striga) germinates poorly on GA, instead utilizes host root-exuded phytohormones strigolactones (SLs) to initiate germination. It remains unknown why GA signalling is not sufficient to germinate Striga. Here I report the identification and functional analysis of three Striga GA receptors, GA INSENSITIVE DWARF1-Like (GIL), and explored the crosstalk between GA and SL signalling pathways. ShGILs interacted with GA signalling targets, ShDELFAs, and overexpression of ShGIL complemented the germination defect of Atgid1ac loss-of-function mutant, indicating ShGILs may function as GA receptors. In addition, two ShGILs interacted with Striga SL receptors, ShHTLs, and overexpression of ShHTLs increased the GA requirement in GA auxotrophic Arabidopsis (ga1-10) suggesting GA-SL crosstalk and its role in regulating Striga germination. This study provides an insight in understanding GA signalling during Striga seed germination.
MSc Exit Seminar- Diana Bonea
Investigation of proteasome architecture, activity, and interactors in Arabidopsis thaliana under abiotic stresses
Abstract:
The 26S proteasome plays a critical role in protein homeostasis via turnover of cellular proteins. Changes in subunit expression, assembly/disassembly of the holoenzyme, and association of non-canonical activators or inhibitors can fine-tune proteasome activity, although such pathways have not been explored in plants. This thesis examines the influence of abiotic stresses on structure, function, and interactors of the Arabidopsis thaliana proteasome. I found that oxidation and salinity decreased the cellular ratio of 26S to 20S proteasomes, altered proteasome subunit composition, and reduced binding of proteasome-associated proteins (PAPs) such as heat-shock and assembly chaperones. Additionally, oxidative stress specifically activated the 20S proteasome; a process which might be promoted by ATP deficiency. Further characterization of PAPs revealed that PBAC1, a 20S assembly chaperone, dampens oxidative damage to the germinating seed. Overall, findings suggest that regulation of proteasome activity, either through PAPs or ATP availability, is necessary for appropriate stress responses in Arabidopsis.
(Zhao & Gazzarrini Lab)
MSc Exit Seminar-Reuben Philip
Characterizing the Microtubule Organizing Centres in Osteoclasts
The skeleton is a metabolically active organ that undergoes continuous remodeling in order to uphold structural integrity and to repair bone following injury. Osteoclasts are highly specialized, multinucleated cells responsible for the selective resorption of bone matrix components, however, they are also responsible for the pathological bone destruction found in microgravity environments, periodontitis, and osteoporosis. Our study investigates the origin of the microtubule cytoskeleton during differentiation and bone resorption. Microtubule nucleation is generally restricted to specific subcellular sites called microtubule organizing centres (MTOCs) and is primarily fulfilled by the centrosome in mitotic animal cells. While mononuclear osteoclast precursor cells contain centrosomal MTOCs, previous research has suggested that functional centrosomal MTOCs do not exist in osteoclasts. To revisit and characterize the MTOCs in osteoclasts, both cell line and primary cell-derived murine osteoclasts were subjected to high-resolution imaging to track centrosome behaviour and their ability to organize microtubules. Through live-cell imaging, fixed immunofluorescence, and ultrastructural analyses, we observed that most, if not all centrosomes donated from precursor cells clustered early in osteoclastogenesis and persisted post-differentiation in non-resorbing and resorbing osteoclasts. Drug-induced microtubule regrowth assays revealed that centrosomes remained individually functional post-differentiation but clustered in a microtubule-dependent manner in order to organize microtubules. Quantification of microtubules emanating from centrosome clusters showed that they were capable of nucleating more microtubules compared to lone centrosomes. Finally, by visualizing Golgi reorganization and the nucleation of Golgi-derived microtubules, we identified the Golgi as a possible non-centrosomal MTOC that potentially facilitates the production of polarized microtubule arrays in osteoclasts. Together these findings show that multinucleated osteoclasts employ unique centrosomal and non-centrosomal MTOCs to organize microtubules.
Supervisor: Prof. Rene Harrison
MSc Exit Seminar-Troy Collins
Adult Neurogenesis in Naked Mole-Rats: Time Course of Cell Maturation and The Effects of Social Novelty
Abstract:
Naked mole-rats are a one of the only eusocial mammalian species. Their complex social structure makes them a unique model for the study of interactions between adult neurogenesis and social behavior. However, foundational knowledge of how cells mature in this species has yet to be established. In this thesis, I explored the time course of maturation of newly born cells in neurogenic regions of the adult naked mole-rat brain. The data demonstrate that time to maturation in this species takes significantly longer than in rats and mice with the majority of newly born cells taking at least 5 months to reach maturity. Then, to test the hypothesis that cell proliferation is related to processing of social cues, I examined the effects of a novel social stimulus on cell proliferation in two subordinate subcastes: workers and soldiers. Soldiers produced more new cells in the subventricular zone than workers. Naked mole-rats exposed to a novel social stimulus had fewer new hippocampal cells, were more aggressive, and investigated the face and body of their counterpart more than those paired with a member of their own colony. Soldiers in novel pairs, but not workers, spent more time sniffing the genitals of their counterpart than those in familiar pairs. Thus, caste and social context play a role in the rate of proliferation and the behavior of naked mole-rats.
Supervisor: Prof. Melissa Holmes
MSc Exit Seminar- Aditi Aggarwal
Effect of thermal stress on the intracellular localization of constitutively expressed heat shock protein HSPA8 (Hsc70) in differentiated and undifferentiated cultured human neuronal cells
Heat shock protein HSPA8 (Hsc70) is a constitutively expressed member of the Hsp70 multigene family that is abundantly expressed in neuronal cells. Previous studies in our laboratory suggest that HSPA8 may play an important role in neuronal pre-protection against cellular stress. This thesis highlights the importance of HSPA8 and its role as a fast responder to cellular stress in differentiated human SH-SY5Y neuronal cells. The effect of thermal stress on the intracellular localization of HSPA8 is compared in differentiated and undifferentiated neuronal cells. HSPA8 rapidly translocated into the nuclei of differentiated neuronal cells after heat shock and co-localized at transcription sites with DNAJB1 (Hsp40) and HSPH1 (Hsp105α) components of the protein disaggregation/ refolding machine. The rapid targeting and assembly of an HSPA8 based disaggregation/ refolding machine acts as a nuclear protective mechanism in differentiated neurons without the time lag needed to induce stress-inducible Hsp70.
Supervisor: Prof. Ian Brown
MSc Exit Seminar- Stefan Schuetz
“Characterizing the basis of strigolactone perception by HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2”
Abstract:
The HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (HTL/KAI2) α/β hydrolases likely play a critical role in the life cycle of parasitic plants of the genus Striga. Several of these hydrolases serve as receptors for strigolactones (SLs), a class of compounds exuded by the roots of some plants, including many economically significant crops. Upon perception of minute levels of SLs, Striga seeds germinate and parasitize the nearby host, often leading to massive losses in crop yields in affected regions. We have produced a series of mutant variants of the Arabidopsis thaliana homolog of HTL/KAI2, which is only weakly responsive to SL. By substituting certain key amino acids in the protein’s active site, we have created a receptor conferring increased Arabidopsis germination under inhibitory conditions in the presence of SL. Additional study of this mutant receptor may offer insight into the biochemical basis of Striga’s SL sensitivity.
Supervisor: Prof. Shelley Lumba