PhD Exit Seminar
Tuesday, January 16, 2018 at 2:10 pm, SW 403 – University of Toronto at Scarborough
Mithunah Krishnamoorthy (Treanor Lab)
“The Role of Novel Ion Channel TRPM7 in B Cell Development and Function”
The channel-kinase Transient Receptor Potential Subfamily M7 (TRPM7) is known to regulate magnesium homeostasis and was first channel implicated in the survival of a B cell line. Our study is the first to show that B cells require TRPM7 for development in a murine model. By using a mouse model where TRPM7 is specifically deleted in B cells under the control of the mb1 promotor, we show that B cells are absent in all peripheral lymphoid tissues due to apoptosis of pre B cells. By using an in vitro stromal cell line system, we demonstrate that B cell development can be partially rescued by high levels of extracellular magnesium. Interestingly, the lack of B cells is accompanied by an expanded granulocyte population in the spleen. In addition to identifying TRPM7 as an essential factor for B cell development, we show that TRPM7 is also an important regulator of B cell activation. DT40 B cells lacking TRPM7 fail to contract and gather antigen when activated. To investigate the role of the kinase domain of TRPM7 we made use of B cells expressing a kinase dead point mutant. These cells were also unable to gather antigen, showing that the kinase domain is an important regulator of this process. We also show that the kinase domain may potentially interact with another important regulator of B cell activation, PLCγ2 to mediate antigen collection and cell contraction. Importantly, primary murine B cells expressing only one allele of TRPM7 or treated with a TRPM7 inhibitor both displayed defects in antigen gathering, confirming our results in the DT40 cell line. Lastly, we show that TRPM7 is essential for antigen internalization, a process that is important for the recruitment of T cell help and ultimately, antibody production.
PhD Exit Seminar
Thursday, December 14th at 10:10am – Ramsay Wright Building, Room 432
Zahra Dargaei (Woodin lab)
Aberrant chloride homeostasis and inhibitory synaptic transmission in Huntington’s disease
Huntington’s disease (HD) is primarily characterized by progressive motor incoordination and involuntary movements that result from neurodegeneration of the striatum. However, cognitive impairments and learning and memory deficits involving the hippocampus emerge in the early stages of the disease and precede the motor impairments by ~15 years. Despite the critical role of GABAergic inhibition in learning and memory, inhibition has not been studied in detail in the HD hippocampi. In my PhD thesis, I have presented three novel pieces of evidence that collectively demonstrate the causative role of Cl– homeostasis in hippocampal-related learning and memory deficits in HD. First, the reduced expression of the Cl–-extruding cotransporter KCC2, and the increased expression in the Cl–-importing cotransporter NKCC1 together result in excitatory GABAergic transmission in the hippocampi of the R6/2 transgenic mouse model of HD. Second, inhibition of the Cl–-importing transporter NKCC1 with the FDA-approved drug bumetanide restores hyperpolarizing GABAergic inhibition and rescues the performance of R6/2 mice on hippocampal-associated behavioral tests. Third, the strength of overall hippocampal inhibition is altered at both the presymptomatic and symptomatic stages of the disease, and involves the disruption of both presynaptic and postsynaptic components. Taken together, my PhD work not only significantly increases our understanding of a less recognized aspect of HD, but also for the first time describes the involvement of Cl– homeostasis in a neurodegenerative disease.
PhD Exit Seminar
Wednesday, November 22, 2017 at 2:10 pm, SW 403 – University of Toronto at Scarborough
Wilfred Carlo de Vega (McGowan Lab)
“DNA Methylation Modifications Associated with Glucocorticoid Sensitivity and Clinical Subtypes of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome”
Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a complex chronic disease with an unknown etiology that is primarily characterized by the presence of a highly debilitating fatigue that fails to resolve after sufficient rest. Additionally, patients must present other nonspecific symptoms relating to pain, unrefreshing sleep, cognitive impairment, and autonomic dysregulation in order to be diagnosed with ME/CFS, resulting in a population with highly heterogeneous symptom profiles. Previous research has consistently observed long-lasting changes in the immune system and hypothalamic-pituitary-adrenal (HPA) axis, a neuroendocrine system that regulates stress response. As a prototypical medically unexplained disease, environmental factors are believed to play a major role in the onset and manifestation of ME/CFS symptoms. These enduring differences may be partly explained through epigenetic modifications, referring to heritable gene expression alterations in the absence of mutations, which are known to reflect environmental, genetic, and stochastic influences on gene expression. This thesis explored the epigenetic modifications associated with the immune and neuroendocrine differences in ME/CFS. Specifically, DNA methylation across the genome was examined in peripheral blood mononuclear cells of ME/CFS patients, and immune response was tested by measuring in vitro sensitivity to glucocorticoids, a class of hormones that serve as an HPA axis effector. Differential methylation in ME/CFS was significantly enriched in immune response and cellular signaling genes, and was localized to 4,699 sites, which may serve as potential biomarkers. Two ME/CFS immune subtypes were observed according to glucocorticoid sensitivity. Integration of methylation and clinical data via machine learning discovered 4 clinical subtypes that were differentiated by T cell response genes, physical functioning, and post-exertional malaise. These results suggest that DNA methylation modifications are a feature of ME/CFS pathology. The identified potential biomarkers and clinical subtypes may be implemented in future work to better understand and clarify the biological differences related to specific ME/CFS symptoms.