Creating and Sustaining Community: An Analysis of LGBTQ Community in London, Ontario

There has been an increase in literature over the last decade on lesbian, gay, bisexual, trans, queer (LGBTQ) communities. However, aside from health-related studies, little has been published pertaining to LGBTQ communities in London, Ontario. This dissertation seeks to answer the following research questions: what are the constitutive elements that make up London’s LGBTQ communities? What forms of community-making prove to be viable and effective in a smaller urban setting? Does the practice of aesthetics/artistic performance lead to socio-political change among members of London’s LGBTQ communities? This is a multidisciplinary research project that utilizes archival, theoretical, and ethnographic-informed qualitative research methods. I argue that LGBTQ community in London is based on four major tenets: they require social supports; there are common or shared visions or goals; they require physical spaces such as bars; and there must be LGBTQ-specific events in these spaces. While some argue that bars and events provide superficial or short-lived communities, London’s LGBTQ history suggests that these spaces are a requirement for communities to flourish. I analyze the ways in which the Homophile Association of London Ontario (HALO) functioned historically as a community and both the promises and limitations of community in three contemporary London LGBTQ venues: The APK, Buckwild’s, and Lavish. I examine the arts and community building of an event called Gen Pop, and argue that it is the most effective contemporary example of LGBTQ community. I discuss the aesthetics of Gen Pop poster advertisements and how this queer aesthetic, through a utilization of camp and raunch, contests normative categories and boundaries. I extend this analysis of a queer aesthetics in discussing how they function at Gen Pop events. I argue that drag performance and other forms of art at Gen Pop utilize a queer aesthetic to challenge social norms. These queer arts are important for community as they expand our understandings and practices of gender and sexuality. Through challenging norms, the arts at Gen Pop help to build a more inclusive and political community where they engage people in dialogue and practice that can lead to socio-political change. I conclude by highlighting the importance of queer arts events for community and the negative consequences for the closure of LGBTQ-specific spaces in London

Organized Chaos: Scatter in the relation between stellar mass and halo mass in small galaxies

We use Local Group galaxy counts together with the ELVIS N-body simulations to explore the relationship between the scatter and slope in the stellar mass vs. halo mass relation at low masses, $M_\star \simeq 10^5 - 10^8 M_\odot$. Assuming models with log-normal scatter about a median relation of the form $M_\star \propto M_\mathrm{halo}^\alpha$, the preferred log-slope steepens from $\alpha \simeq 1.8$ in the limit of zero scatter to $\alpha \simeq 2.6$ in the case of $2$ dex of scatter in $M_\star$ at fixed halo mass. We provide fitting functions for the best-fit relations as a function of scatter, including cases where the relation becomes increasingly stochastic with decreasing mass. We show that if the scatter at fixed halo mass is large enough ($\gtrsim 1$ dex) and if the median relation is steep enough ($\alpha \gtrsim 2$), then the "too-big-to-fail" problem seen in the Local Group can be self-consistently eliminated in about $\sim 5-10\%$ of realizations. This scenario requires that the most massive subhalos host unobservable ultra-faint dwarfs fairly often; we discuss potentially observable signatures of these systems. Finally, we compare our derived constraints to recent high-resolution simulations of dwarf galaxy formation in the literature. Though simulation-to-simulation scatter in $M_\star$ at fixed $M_\mathrm{halo}$ is large among separate authors ($\sim 2$ dex), individual codes produce relations with much less scatter and usually give relations that would over-produce local galaxy counts.Comment: 15 pages, 6 figures, 1 table. Accepted for publication into MNRA

A conserved protein interaction network involving the yeast MAP kinases Fus3 and Kss1

The Saccharomyces cerevisiae mitogen-activated protein kinases (MAPKs) Fus3 and Kss1 bind to multiple regulators and substrates. We show that mutations in a conserved docking site in these MAPKs (the CD/7m region) disrupt binding to an important subset of their binding partners, including the Ste7 MAPK kinase, the Ste5 adaptor/scaffold protein, and the Dig1 and Dig2 transcriptional repressors. Supporting the possibility that Ste5 and Ste7 bind to the same region of the MAPKs, they partially competed for Fus3 binding. In vivo, some of the MAPK mutants displayed reduced Ste7-dependent phosphorylation, and all of them exhibited multiple defects in mating and pheromone response. The Kss1 mutants were also defective in Kss1-imposed repression of Ste12. We conclude that MAPKs contain a structurally and functionally conserved docking site that mediates an overall positively acting network of interactions with cognate docking sites on their regulators and substrates. Key features of this interaction network appear to have been conserved from yeast to humans

A metabolite binding protein moonlights as a bile- responsive chaperone

Bile salts are secreted into the gastrointestinal tract to aid in the absorption of lipids. In addition, bile salts show potent antimicrobial activity in part by mediating bacterial protein unfolding and aggregation. Here, using a protein folding sensor, we made the surprising discovery that the Escherichia coli periplasmic glycerol- 3- phosphate (G3P)- binding protein UgpB can serve, in the absence of its substrate, as a potent molecular chaperone that exhibits anti- aggregation activity against bile salt- induced protein aggregation. The substrate G3P, which is known to accumulate in the later compartments of the digestive system, triggers a functional switch between UgpB’s activity as a molecular chaperone and its activity as a G3P transporter. A UgpB mutant unable to bind G3P is constitutively active as a chaperone, and its crystal structure shows that it contains a deep surface groove absent in the G3P- bound wild- type UgpB. Our work illustrates how evolution may be able to convert threats into signals that first activate and then inactivate a chaperone at the protein level in a manner that bypasses the need for ATP.SynopsisThe periplasmic glycerol- 3- phosphate binding protein, UgpB, was found to have dual functions, as a metabolite binding protein and as a bile- responsive molecular chaperone. Stomach- acid induced stripping of its glycerol- 3- phosphate substrate functions as a switch that activates the chaperone activity of UgpB.A tripartite periplasmic protein folding sensor and Tn- Seq uncover UgpB as a new chaperone.UgpB prevents bile- induced protein aggregation when in its G3P- free form.Stomach acid- induced G3P stripping activates UgpB chaperone function.Crystal structure of a G3P- nonbinding variant of UgpB reveals opening of a deep surface groove when compared to the structure of G3P- bound wild- type UgpB.A periplasmic folding sensor reveals a mechanism by which stomach acid- induced G3P stripping remodels UgpB into a chaperone that prevents bile- induced bacterial protein aggregation.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/6/embj2019104231.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/5/embj2019104231-sup-0002-EVFigs.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/4/embj2019104231-sup-0006-SDataFig3.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/3/embj2019104231.reviewer_comments.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/2/embj2019104231_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163430/1/embj2019104231-sup-0005-SDataFig2.pd

Elaborating a coiledâ coilâ assembled octahedral protein cage with additional protein domains

De novo design of protein nanoâ cages has potential applications in medicine, synthetic biology, and materials science. We recently developed a modular, symmetryâ based strategy for protein assembly in which short, coiledâ coil sequences mediate the assembly of a protein building block into a cage. The geometry of the cage is specified by the combination of rotational symmetries associated with the coiledâ coil and protein building block. We have used this approach to design wellâ defined octahedral and tetrahedral cages. Here, we show that the cages can be further elaborated and functionalized by the addition of another protein domain to the free end of the coiledâ coil: in this case by fusing maltoseâ binding protein to an octahedral protein cage to produce a structure with a designed molecular weight of ~1.8 MDa. Importantly, the addition of the maltose binding protein domain dramatically improved the efficiency of assembly, resulting in ~ 60â fold greater yield of purified protein compared to the original cage design. This study shows the potential of using small, coiledâ coil motifs as offâ theâ shelf components to design MDaâ sized protein cages to which additional structural or functional elements can be added in a modular manner.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/146469/1/pro3497.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/146469/2/pro3497_am.pd

Organized Chaos: Scatter in the relation between stellar mass and halo mass in small galaxies

We use Local Group galaxy counts together with the ELVIS N-body simulations to explore the relationship between the scatter and slope in the stellar mass versus halo mass relation at low masses, M⋆ ≃ 10^5–10^8 M⊙. Assuming models with lognormal scatter about a median relation of the form M⋆ ∝ M^α_(halo), the preferred log-slope steepens from α ≃ 1.8 in the limit of zero scatter to α ≃ 2.6 in the case of 2 dex of scatter in M⋆ at fixed halo mass. We provide fitting functions for the best-fitting relations as a function of scatter, including cases where the relation becomes increasingly stochastic with decreasing mass. We show that if the scatter at fixed halo mass is large enough (≳ 1 dex) and if the median relation is steep enough (α ≳ 2), then the ‘too-big-to-fail’ problem seen in the Local Group can be self-consistently eliminated in about ∼5–10 per cent of realizations. This scenario requires that the most massive subhaloes host unobservable ultra-faint dwarfs fairly often; we discuss potentially observable signatures of these systems. Finally, we compare our derived constraints to recent high-resolution simulations of dwarf galaxy formation in the literature. Though simulation-to-simulation scatter in M⋆ at fixed M_(halo) is large among different authors (∼2 dex), individual codes produce relations with much less scatter and usually give relations that would overproduce local galaxy counts

Genetic Selection for Enhanced Folding In Vivo Targets the Cys14-Cys38 Disulfide Bond in Bovine Pancreatic Trypsin Inhibitor

The periplasm provides a strongly oxidizing environment; however, periplasmic expression of proteins with disulfide bonds is often inefficient. Here, we used two different tripartite fusion systems to perform in vivo selections for mutants of the model protein bovine pancreatic trypsin inhibitor (BPTI) with the aim of enhancing its expression in Escherichia coli. This trypsin inhibitor contains three disulfides that contribute to its extreme stability and protease resistance. The mutants we isolated for increased expression appear to act by eliminating or destabilizing the Cys14-Cys38 disulfide in BPTI. In doing so, they are expected to reduce or eliminate kinetic traps that exist within the well characterized in vitro folding pathway of BPTI. These results suggest that elimination or destabilization of a disulfide bond whose formation is problematic in vitro can enhance in vivo protein folding. The use of these in vivo selections may prove a valuable way to identify and eliminate disulfides and other rate-limiting steps in the folding of proteins, including those proteins whose in vitro folding pathways are unknown. Antioxid. Redox Signal. 14, 973-984.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90494/1/ars-2E2010-2E3712.pd

Symmetryâ Directed Selfâ Assembly of a Tetrahedral Protein Cage Mediated by de Novoâ Designed Coiled Coils

The organization of proteins into new hierarchical forms is an important challenge in synthetic biology. However, engineering new interactions between protein subunits is technically challenging and typically requires extensive redesign of proteinâ protein interfaces. We have developed a conceptually simple approach, based on symmetry principles, that uses short coiledâ coil domains to assemble proteins into higherâ order structures. Here, we demonstrate the assembly of a trimeric enzyme into a wellâ defined tetrahedral cage. This was achieved by genetically fusing a trimeric coiledâ coil domain to its C terminus through a flexible polyglycine linker sequence. The linker length and coiledâ coil strength were the only parameters that needed to be optimized to obtain a high yield of correctly assembled protein cages.Geometry lesson: A modular approach for assembling proteins into largeâ scale geometric structures was developed in which coiledâ coil domains acted as â twist tiesâ to facilitate assembly. The geometry of the cage was specified primarily by the rotational symmetries of the coiled coil and building block protein and was largely independent of protein structural details.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/1/cbic201700406_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/2/cbic201700406.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/3/cbic201700406-sup-0001-misc_information.pd

Protein Scaffolds Can Enhance the Bistability of Multisite Phosphorylation Systems

The phosphorylation of a substrate at multiple sites is a common protein modification that can give rise to important structural and electrostatic changes. Scaffold proteins can enhance protein phosphorylation by facilitating an interaction between a protein kinase enzyme and its target substrate. In this work we consider a simple mathematical model of a scaffold protein and show that under specific conditions, the presence of the scaffold can substantially raise the likelihood that the resulting system will exhibit bistable behavior. This phenomenon is especially pronounced when the enzymatic reactions have sufficiently large KM, compared to the concentration of the target substrate. We also find for a closely related model that bistable systems tend to have a specific kinetic conformation. Using deficiency theory and other methods, we provide a number of necessary conditions for bistability, such as the presence of multiple phosphorylation sites and the dependence of the scaffold binding/unbinding rates on the number of phosphorylated sites

Arousal frequency is associated with increased fatigue in obstructive sleep apnea

Fatigue is an important and often underemphasized symptom in patients with obstructive sleep apnea (OSA). Sleep fragmentation, i.e., arousals and disruptions in sleep architecture, is common in patients with OSA and may potentially contribute to their fatigue. We hypothesized that arousal frequency and changes in sleep architecture contribute to the fatigue experienced by patients with OSA. Seventy-three patients with diagnosed but untreated OSA (AHI ≥ 15) were enrolled in this study. A baseline polysomnogram was obtained, and fatigue was measured with the Multidimensional Fatigue Symptom Inventory-short form (MFSI-sf). We evaluated the association between fatigue and arousals and various polysomongraphic variables, including sleep stages and sleep efficiency. Significant correlations between MFSI-sf subscale scores and various arousal indices were noted. Emotional fatigue scores were associated with total arousal index (r = 0.416, p = .021), respiratory movement arousal index (r = 0.346, p = .025), and spontaneous movement arousal index (r = 0.378, p = .025). Physical fatigue scores were associated with total arousal index (r = 0.360, p = .033) and respiratory movement arousal index (r = 0.304, p = .040). Percent of stage 1 sleep and REM sleep were also associated with physical and emotional fatigue scores. Hierarchal linear regression analysis demonstrated that emotional fatigue scores were independently associated with spontaneous movement arousals after controlling for age, body mass index, depression, and sleep apnea severity. These findings suggest that arousals may contribute to the fatigue seen in patients with OSA