PhD Proposal Seminar - Ghyda Hashim (Mounir Lab)
Analysis of early plant gene(s) involved in the viral infectious process and plant defence responses through the dysregulation of plant circular RNAs
Abstract
Unlike linear RNAs, circular RNAs (circRNAs) are covalently closed continuous loops without 5’ and 3’ ends, cap or polyadenylated tail molecules. CircRNAs are generated from RNA splicing followed by back ligation. Although circRNAs have continued to gain attention, our understanding of their function remains limited. We selected the strategy to profile changes (dysregulation up/down) of circRNAs in response to virus (or single viral gene) infection in plants rather than profiling differentially expressed plant proteins (cDNAs).
The overarching goal of my research includes four aims: (i) evaluating circRNAs and their predicted impact on the dysregulation of plant genes due to viral infectious process; (ii) identifying plant circular RNAs generated/dysregulated very early on in the infection process that are responsible for triggering plant cell responses to the virus infectious process; (iii) studying and identifying plant genes (through their circRNAs) involved in plant host (susceptible/compatible interactions) responses to viral infection versus non-host recognition; and (iv) investigating how those circular RNAs are involved in the plant defense mechanisms, against viruses and how some viruses can over-come the defense mechanisms to be able to replicate.
Our current study aims to shed light on the role(s) of circular RNAs in the context of virus infection, virus susceptibility and resistance.
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https://utoronto.zoom.us/j/81695333689
Meeting ID: 816 9533 3689
Host: AbouHaidar Mounir (mounir.abouhaidar@utoronto.ca)
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PhD Proposal Seminar - Yingtian He (Liu Lab)
The Functional Roles of Inhibitory Neuronal Populations in Brainstem Circuit Mediating Optokinetic Reflex
Abstract
The optokinetic reflex (OKR) response is an oculomotor behaviour that helps stabilize vision. It can be elicited in both horizontal and vertical direction. Horizontal OKR is driven by brainstem structures called the nucleus of optic tract and the dorsal terminal nucleus (NOT-DTN). NOT-DTN is abundant in inhibitory neurons, with about 30% of its neurons expressing GAD, a molecular marker for GABAergic inhibitory neurons36. However, what role these functionally different neurons have in OKR response is yet unknown. My preliminary data show that inhibitory neurons projecting to different downstream targets are different in their functional properties. This suggests they may encode different information and play different roles in regulating OKR response. Next, I will investigate how the inhibitory population affects NOT-DTN population activity and how it affects OKR response. Finally, I will study how different inhibitory populations affect the target of the innervated, and how these different populations affect behavioral output of OKR. This study will help us to understand the information encoded by different inhibitory populations in NOT-DTN, and the roles they play in regulating OKR response. Hopefully, these results can shine light on the function of inhibition in reflex regulation.
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https://utoronto.zoom.us/j/87081276866
Meeting ID: 870 8127 6866
Host: Baohua Liu (baohua.liu@utoronto.ca)
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PhD Proposal Seminar - Emily Dong (Chang and Tropepe Lab)
Examining the role of rod development genes in the evolution of transmuted photoreceptors in squamates
Abstract
The evolution of a duplex of highly specialized photoreceptors (rods and cones) in vertebrates has allowed for their invasion into a diversity of environmental niches. Among vertebrates, squamates (a group including snakes and lizards) are unique in having evolved simplex retinas by the repeated secondary losses of either rods or cones as an adaptation to new light environments. The loss of duplexity in squamates appears to have occurred not by the outright loss of one type, but by “transmutation” or the evolutionary transition between rods and cones, whereby one photoreceptor class evolves features of the other.
Transmuted photoreceptors have been identified by molecular and morphological means in adult eyes. However, it remains unclear how these photoreceptors arise during embryonic development, leaving conserved rod vertebrate photoreceptor development genes including NRL, NR2E3, and CRX unexplored in squamates. I hypothesize that this rod-determining developmental network has been a target for selection in squamate lineages, causing adaptive changes in their functional capacity to induce rod identity, leading to transmutation.
To address this, my first Aim is a computational analysis of evolutionary selection pressures on orthologous squamate sequences. My second Aim is focused on the temporal and spatial expression patterns of these same genes in gecko embryos. Lastly, my third aim tests hypotheses developed from the molecular evolution analyses in Aim 1 by expressing squamate proteins in cultured mammalian cells and in the developing zebrafish retina. My research proposal touches on evolutionary, developmental, and functional perspectives that can be united for a holistic view of vertebrate photoreceptor identity, with a particular focus on how squamate groups have evolved photoreceptors that largely lie outside the typical photoreceptor duplex.
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https://us02web.zoom.us/j/86156218181?pwd=YUcxMVN6dUxRY1FWaHpQSENFUGlhUT09
Meeting ID: 861 5621 8181
Host: Belinda Chang (belinda.chang@utoronto.ca)
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PhD Proposal Seminar - Zachary Kileeg (Mott Lab)
Investigation and identification of immune-related receptor-kinases and their adaptive evolution
Abstract
Plant disease is a major contributor to crop loss around the world. Although many methods are in use to reclaim these losses, tuning basal immunity through receptor-kinases offers a robust method for crop protection. Receptor-kinases play an important role in plant immune responses, yet our understanding of them is limited. My research leverages evolutionary methods to detect possible immune related receptor-kinases. To achieve this, I identified all receptor kinases across 26 ecotypes of Arabidopsis thaliana and clustered them into orthogroups. With this, I found a subset of receptor-kinase families showing expansion of their accessory genomes and evidence of tandem duplication – both indicative of adaptive evolution. I have also calculated the rates of positive and negative selection across the leucine-rich repeat receptor-kinase (LRR-RK) family to fingerprint potential selection events. My project moving forward sets out to complete these analyses for all receptor-kinase families and to examine the evolution and diversity of the receptor-kinases generally. Following this, I will use phylogenetic methods to analyze rates of expansion of the receptor-kinases across different plant species and identify protein domain architectures. These analyses will discover novel immune-related receptors in Arabidopsis thaliana, which I will then confirm using high-throughput immune assays.
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https://utoronto.zoom.us/j/82827947881
Meeting ID: 828 2794 7881
Host: Adam Mott (adam.mott@utoronto.ca)
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PhD Proposal Seminar - Tara McDonnell (Tropepe Lab)
Examining the role of Ehmt1/2-dependent gene regulation in zebrafish retinal progenitor cells
Abstract
During retinal development, retinal progenitor cells (RPCs) undergo a series of multiplication and differentiation events across a concise timeline to form the stratified retina. Epigenetic silencing is crucial when RPCs differentiate to coordinate the downregulation of progenitor genes. A key complex responsible for initiating the first repressive modifications is the Ehmt1/2 complex, which contains four known proteins: Ehmt1, Ehmt2, WIZ and ZNF644. The Ehmt1/2 complex is most well-known for depositing a repressive dimethylation mark at Histone 3 Lysine 9 (H3K9me2) at gene promotors to recruit other repressive complexes and promote gene silencing. However, research over the last decade indicates that in addition to gene silencing, Ehmt2 possesses a multifaceted role in neural tissues including direct and indirect transcriptional activation, and exon splicing, to encourage proper neural cell differentiation and survival. Despite significant research into the Ehmt1/2 complex across various species and tissues, many questions remain, particularly surrounding the discrete roles for different Ehmt1/2 components in its various functions.
Hence, my thesis research is focussed on delineating the distinct roles of different components of the Ehmt1/2 complex in RPC function, and ultimately, retinal development. Our lab generated a zebrafish Ehmt2 loss of function mutant (Ehmt2∆4/∆4) which mimics Ehmt2 loss of function models in other species, however it is unique in that it is also adult viable and fertile. The first aim of my project involves characterising these Ehmt2∆4/∆4 mutant embryos and determining the effects of Ehmt2 loss of function on RPC cell cycle regulation. The Ehmt2∆4/∆4 mutant demonstrates a global loss of H3K9me2 in embryonic cells. Thus, the second aim will focus on understanding the genome wide impact of the loss of this repressive mark on global and RPC-specific gene expression. Finally, the third aim uses CRISPR gene targeting to generate four other Ehmt1/2 complex loss of function mutants. These mutants will then be compared with the Ehmt2∆4/∆4 mutants, and with each other, to examine the distinct roles of different complex components and investigate potential forms of compensation between different Ehmt1/2 mutant alleles. Understanding the roles of different Ehmt1/2 complex components will provide insight for future research into tissue development, developmental disorders, illnesses with aberrant Ehmt1/2 function such as some cancers, and in developing drugs to target these illnesses.
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https://utoronto.zoom.us/j/85249561868
Meeting ID: 852 4956 1868
Host: Vincent Tropepe (v.tropepe@utoronto.ca)
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PhD Proposal Seminar - Sonia Ehi-Eromosele (Phillips Lab)
Control of substrate supply in the chloroplast 2C-methyl-D-erythritol-4-phosphate pathway
Abstract
The chloroplast 2C-methyl-D-erythritol-4-phosphate (MEP) pathway supplies precursors for terpenoid cofactors essential to photosynthesis and primary metabolism. In specialized contexts, it produces multitudes of high value secondary metabolites. This precursor pathway generates the universal terpenoid intermediates isopentenyl and dimethylallyl diphosphate (IDP and DMADP) from the central metabolic intermediates D-glyceraldehyde-3-phosphate (GAP) and pyruvate. While GAP is supplied via the Calvin-Benson cycle, the source of pyruvate in the chloroplast remains poorly understood. Pyruvate entering the MEP pathway may originate through the reimport of glycolytically derived phosphoenolpyruvate (PEP) from the cytosol due to downregulation of glycolysis in chloroplasts. Endogenous pyruvate production with contributions from the cytosolic pentose phosphate pathway (PPP) provides an alternative explanation. My dissertation aims to distinguish between these alternative carbon sources supplying the chloroplast MEP pathway and evaluate their relative contributions in Arabidopsis thaliana. This research line will provide insight to the following questions: What is the role of the PEP reimport shunt in MEP substrate supply in photosynthetic tissue? Can chloroplasts support endogenous pyruvate production? Can the Entner-Doudoroff pathway be engineered in plants to enhance pyruvate and GAP availability for the MEP pathway? To address the above questions, I will rely on 13C isotopic labeling studies in Arabidopsis mutants (cue1 or xpt2) deficient in a carbon import route. Mutant labeling time courses will be complemented by those of stable transgenic lines capable of pyruvate production in the chloroplast through constitutive expression of plastidic enolase and phosphoglycerate mutase. Mutants and transgenics altered in carbon metabolism or transport (and wild-type) will be examined using a 13CO2 kinetic labeling approach and mass spectrometry analysis. This research will provide a detailed understanding of substrate control for the plastid terpenoid precursor pathway.
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https://utoronto.zoom.us/j/87858737038
Meeting ID: 878 5873 7038
Host: Michael Phillips (michaelandrew.phillips@utoronto.ca)
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PhD Proposal Seminar - Jerrica Jamison (Welch Lab)
A Comparative Investigation of Fructose Metabolism in Flying Nectivorous Vertebrates
Abstract
Fructose is a sugar found in many foods but poorly metabolized by most vertebrates. Over consumption of fructose has been linked to a number of diseases including diabetes and chronic kidney disease. However, up to half of the diet hummingbirds and nectar bats consume is made of fructose, but they do not show signs of metabolic disease. In hummingbirds, unique adaptations regarding fructose transport and metabolism have already been uncovered, but fructose metabolism in nectar bats is still poorly understood. I plan to use a combination of stable isotope respirometry and blood metabolite analyses to uncover metabolic adaptations regarding fructose use in nectar bats. As there are a number of independent lineages of nectivorous bats, I will compare adaptations between these bat lineages to look for signs of convergent evolution. Additionally, I will examine seasonal variation in fructose metabolism in a bat that opportunistically feeds on nectar during part of the year, exploring plasticity in animals’ ability to metabolize fructose. I will also continue previous work on hummingbird fructose metabolism during exercise and expand the knowledge of fructose as fuel for exercise to nectar bats. Finally, I will look at potential glycation stress in hummingbirds and nectar bats due to fructose consumption using mass spectrometry. This work is the first to look at fructose metabolism in nectar bats, and will give insight into how they can ingest so much of this disease causing sugar without suffering the consequences other animals do.
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Wednesday, June 2nd, 2021 at 10:00am
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https://utoronto.zoom.us/j/83033665879
Meeting ID: 830 3366 5879
Host: Kenneth Welch (kwelch@utsc.utoronto.ca)
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PhD Proposal Seminar - Aparna Haldar (Mott Lab)
Investigating immune functions of unknown LRR-RLKs in Arabidopsis thaliana
Abstract
To receive environmental stimuli, plants rely on the activities of numerous cell surface receptors. The largest group of cell surface receptors is the approximately 225 leucine rich repeat receptor-like kinases (LRR-RLKs) present within the genome of the model plant Arabidopsis thaliana. These receptors have been shown to have important roles in both plant growth and immunity, but most are not well characterized.
The goal of my research is to assign biological functions to orphan LRR-RLKs and examine the diversity and evolution of these receptors across the diversity of plants. Network analysis was used to predict biological functions for LRR-RLKs by constructing biologically relevant spatial and temporal protein-protein interaction networks. These networks show the unique interactions observed in plant tissues relevant to pathogen infection and induced upon pathogen challenge. Based on these networks two candidate genes were chosen for further study, SRF-9 and MUL. To better understand the evolutionary context in which these receptors emerged, LRR-RLKs were divided into smaller sub-families to examine expansion and contraction patterns. Preliminary results have indicated that expansion is observed in sub-family I of the LRR-RLKs specifically within the Brassicaceae family.
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https://utoronto.zoom.us/j/81206557657
Meeting ID: 812 0655 7657
Host: Adam Mott (adam.mott@utoronto.ca)
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PhD Proposal Seminar - Eduardo Ramirez Rodriguez (McFarlane Lab)
Decoding the signals that shape plant cell walls
Abstract
The plant cell wall is a polysaccharide-based extracellular matrix that surrounds and protects plant cells. As highly dynamic structures, the cell walls balance between rigidity for protection and structure, and flexibility to grow and respond to environmental cues. These properties make it an ideal source for renewable materials and bioenergy. However, attempts to overhaul the plant’s finely tuned polysaccharide deposition ability by increasing the expression of polysaccharide biosynthetic enzymes in plants have had limited success. These results hint at the existence of an underlying regulatory mechanism, called ‘cell wall integrity’ (CWI) signaling that perceives changes outside the cell, and in turn, remodels the cell wall and/or regulates plant growth. To date, several cell membrane-bound kinases have been implicated as the signal perception component of CWI, yet the downstream players remain undefined. We employed a proteomics approach by treating plants to induce cell wall stress and analyzing phosphorylated peptides through high resolution mass spectrometry. Altogether, our data from 3 independent experiments (n=9) shows 242 differentially phosphorylated phosphopeptides, corresponding to 241 proteins. We conducted bioinformatic analysis of all 241 candidates integrating gene ontology terms, protein-protein interaction networks, phosphomotif analysis, and comparative analysis with other stress-induced phosphoproteomes. These results guided selection of 30 candidates for a reverse genetics screen under cell wall stress conditions. Results from this screen implicate several intracellular kinase candidates in CWI. I propose to characterize several of these candidates within the context of CWI, using a combination of biochemistry, genetics, live cell imaging and proteomics.
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https://utoronto.zoom.us/j/84659195735
Meeting ID: 846 5919 5735
Host: Heather McFarlane (h.mcfarlane@utoronto.ca)
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PhD Proposal Seminar - Miranda de Saint-Rome (Woodin Lab)
Characterizing hyperexcitability in the C9orf72 mouse model of ALS
Abstract
Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease in humans, whereby upper motor neurons in the motor cortex and lower motor neurons in the spinal cord degenerate, eventually resulting in death. A key mechanistic determinant of neurodegeneration in ALS is postulated to be aberrations in neuronal excitability and subsequent excitotoxicity in the primary motor cortex, promoting a cascade of pathological events that facilitate cell death. Additionally, previous research has identified the GGGGCC hexanucleotide repeat expansion in the C9orf72 gene as the most common genetic cause of ALS. However, little is known about the contribution of the C9orf72 gene to neuronal excitability in the primary motor cortex. Therefore, using a C9orf72 mouse model, my aims are to (1) Characterize synaptic transmission and neuronal excitability in C9orf72 mice; (2) Elucidate the mechanisms underlying changes in transmission; and (3) Assess the ability of Riluzole, a clinically approved treatment for ALS whose mechanism of action has not been completely resolved, to work synergistically with chemogenetics to rescue hyperexcitability in C9orf72 mice. Results from this study will be the first to characterize hyperexcitability using electrophysiological recordings in the cortex and hippocampus of the C9orf72 mouse, thereby revealing essential information about the neurophysiological mechanisms underlying neurodegeneration in C9orf72 ALS patients.
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Tuesday, May 11th, 2021 at 1:00pm
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https://utoronto.zoom.us/j/85831754165
Meeting ID: 858 3175 4165
Host: Melanie Woodin (m.woodin@utoronto.ca)
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