Spontaneous Unfolding and Refolding of FNIII Domains Assayed by Thiol Exchange

Fibronectin (FN) is a large extracellular matrix (ECM) protein that is made up of

type I (FNI), type II (FNII), & type III (FNIII) domains. It assembles into an insoluble

supra-­‐‑molecular structure: the fibrillar FN matrix. FN fibrillogenesis is a cell‐‑mediated process, which is initiated when FN binds to integrins on the cell surface. The FN matrix plays an important role in cell migration, proliferation, signaling & adhesion. Despite decades of research, the FN matrix is one of the least understood supra-­‐‑molecular protein assemblies. There have been several attempts to elucidate the exact mechanism of matrix assembly resulting in significant progress in the field but it is still unclear as to what are FN-­‐‑FN interactions, the nature of these interactions and the domains of FN that

are in contact with each other. FN matrix fibrils are elastic in nature. Two models have been proposed to explain the elasticity of the fibrils. The first model: the ‘domain unfolding’ model postulates that the unraveling of FNIII domains under tension explains fibril elasticity.

The second model relies on the conformational change of FN from compact to extended to explain fibril elasticity. FN contain 15 FNIII domains, each a 7-­‐‑strand beta sandwich. Earlier work from our lab used the technique of labeling a buried Cys to study the ‘domain unfolding’ model. They used mutant FNs containing a buried Cys in a single FNIII domain and found that 6 of the 15 FNIII domains label in matrix fibrils. Domain unfolding due to tension, matrix associated conformational changes or spontaneous folding and unfolding are all possible explanation for labeling of the buried Cys. The present study also uses the technique of labeling a buried Cys to address whether it is spontaneous folding and unfolding that labels FNIII domains in cell culture. We used thiol reactive DTNB to measure the kinetics of labeling of buried Cys in eleven FN III domains over a wide range of urea concentrations (0-­‐‑9M). The kinetics data were globally fit using Mathematica. The results are equivalent to those of H-­‐‑D exchange, and

provide a comprehensive analysis of stability and unfolding/folding kinetics of each

domain. For two of the six domains spontaneous folding and unfolding is possibly the reason for labeling in cell culture. For the rest of the four domains it is probably matrix associated conformational changes or tension induced unfolding.

A long-­‐‑standing debate in the protein-­‐‑folding field is whether unfolding rate

constants or folding rate constants correlate to the stability of a protein. FNIII domains all have the same ß sandwich structure but very different stabilities and amino acid sequences. Our study analyzed the kinetics of unfolding and folding and stabilities of eleven FNIII domains and our results show that folding rate constants for FNIII domains are relatively similar and the unfolding rates vary widely and correlate to stability. FN forms a fibrillar matrix and the FN-­‐‑FN interactions during matrix fibril formation are not known. FNI 1-­‐‑9 or the N-­‐‑ terminal region is indispensible for matrix formation and its major binding partner has been shown to be FNIII 2. Earlier work from our lab, using FRET analysis showed that the interaction of FNI 1-­‐‑9 with a destabilized FNIII 2 (missing the G strand, FNIII 2ΔG) reduces the FRET efficiency. This efficiency is restored in the presence of FUD (bacterial adhesion from S. pyogenes) that has been known to interact with FNI 1-­‐‑9 via a tandem ß zipper. In the present study we

use FRET analysis and a series of deletion mutants of FNIII 2ΔG to study the shortest fragment of FNIII 2ΔG that is required to bind FNI 1-­‐‑9. Our results presented here are qualitative and show that FNIII 2ΔC’EFG is the shortest fragment required to bind FNI 1-­‐‑9. Deletion of one more strand abolishes the interaction with FNI 1-­‐‑9.

Decoding Spontaneous Emotional States in the Human Brain.

Pattern classification of human brain activity provides unique insight into the neural underpinnings of diverse mental states. These multivariate tools have recently been used within the field of affective neuroscience to classify distributed patterns of brain activation evoked during emotion induction procedures. Here we assess whether neural models developed to discriminate among distinct emotion categories exhibit predictive validity in the absence of exteroceptive emotional stimulation. In two experiments, we show that spontaneous fluctuations in human resting-state brain activity can be decoded into categories of experience delineating unique emotional states that exhibit spatiotemporal coherence, covary with individual differences in mood and personality traits, and predict on-line, self-reported feelings. These findings validate objective, brain-based models of emotion and show how emotional states dynamically emerge from the activity of separable neural systems.

Fermion Mass Generation without Spontaneous Symmetry Breaking

The conventional mechanism of fermion mass generation in the Standard Model involves Spontaneous Symmetry Breaking (SSB). In this thesis, we study an alternate mechanism for the generation of fermion masses that does not require SSB, in the context of lattice field theories. Being inherently strongly coupled, this mechanism requires a non-perturbative approach like the lattice approach.

In order to explore this mechanism, we study a simple lattice model with a four-fermion interaction that has massless fermions at weak couplings and massive fermions at strong couplings, but without any spontaneous symmetry breaking. Prior work on this type of mass generation mechanism in 4D, was done long ago using either mean-field theory or Monte-Carlo calculations on small lattices. In this thesis, we have developed a new computational approach that enables us to perform large scale quantum Monte-Carlo calculations to study the phase structure of this theory. In 4D, our results confirm prior results, but differ in some quantitative details of the phase diagram. In contrast, in 3D, we discover a new second order critical point using calculations on lattices up to size $ 60^3$. Such large scale calculations are unprecedented. The presence of the critical point implies the existence of an alternate mechanism of fermion mass generation without any SSB, that could be of interest in continuum quantum field theory.

Analyse cinétique et cinématique de la locomotion chez les enfants atteints d'un déficit moteur cérébral : comparaison de l'orthèse tibiale fixe et articulée

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Comparaison des paramètres cinématiques et cinétiques lors de la locomotion chez des enfants obèses et non-obèses

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Modulation de l'activité respiratoire par la locomotion chez la lamproie

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Contribution du cortex moteur et des afférences cutanées dans le contrôle et la plasticité de la locomotion chez le chat

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

Quantum-limited estimation of continuous spontaneous localization

We apply the formalism of quantum estimation theory to extract information about potential collapse mechanisms of the continuous spontaneous localisation (CSL) form.
In order to estimate the strength with which the field responsible for the CSL mechanism couples to massive systems, we consider the optomechanical interaction
between a mechanical resonator and a cavity field. Our estimation strategy passes through the probing of either the state of the oscillator or that of the electromagnetic field that drives its motion. In particular, we concentrate on all-optical measurements, such as homodyne and heterodyne measurements.
We also compare the performances of such strategies with those of a spin-assisted optomechanical system, where the estimation of the CSL parameter is performed
through time-gated spin-like measurements.

Multicentric Genome-Wide Association Study for Primary Spontaneous Pneumothorax

Despite elevated incidence and recurrence rates for Primary Spontaneous Pneumothorax (PSP), little is known about its etiology, and the genetics of idiopathic PSP remains unexplored. To identify genetic variants contributing to sporadic PSP risk, we conducted the first PSP genome-wide association study. Two replicate pools of 92 Portuguese PSP cases and of 129 age- and sex-matched controls were allelotyped in triplicate on the Affymetrix Human SNP Array 6.0 arrays. Markers passing quality control were ranked by relative allele score difference between cases and controls (|RASdiff|), by a novel cluster method and by a combined Z-test. 101 single nucleotide polymorphisms (SNPs) were selected using these three approaches for technical validation by individual genotyping in the discovery dataset. 87 out of 94 successfully tested SNPs were nominally associated in the discovery dataset. Replication of the 87 technically validated SNPs was then carried out in an independent replication dataset of 100 Portuguese cases and 425 controls. The intergenic rs4733649 SNP in chromosome 8 (between LINC00824 and LINC00977) was associated with PSP in the discovery (P = 4.07E-03, ORC[95% CI] = 1.88[1.22-2.89]), replication (P = 1.50E-02, ORC[95% CI] = 1.50[1.08-2.09]) and combined datasets (P = 8.61E-05, ORC[95% CI] = 1.65[1.29-2.13]). This study identified for the first time one genetic risk factor for sporadic PSP, but future studies are warranted to further confirm this finding in other populations and uncover its functional role in PSP pathogenesis.

Spontaneous rupture of a giant hepatic hemangioma. Sequential treatment with preoperative transcatheter arterial embolization and conservative hepatectomy

Hemangioma is the most common benign tumor of the liver and it is often asymptomatic. Spontaneous rupture of liver hemangiomas is a rare but potentially lethal complication. Emergent hepatic resection has been the treatment of choice but carries high operative morbidity and mortality. Recently, preoperative transcatheter arterial embolization (TAE) has been used successfully for the management of bleeding ruptured liver tumors and non-operative treatment of symptomatic giant liver hemangiomas. We report a case of spontaneous rupture of a giant hepatic hemangioma that presented with thoracic and abdominal pain and shock due to hemoperitoneum. Once proper diagnosis was made the patient was successfully managed by TAE, followed by conservative hepatic resection.

Liver resection for hemoperitoneum caused by spontaneous rupture of unrecognized hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is an increasingly common form of cancer. Although its spontaneous rupture is rare in Western countries, it constitutes a surgical emergency and is associated with high mortality. There is a lack of consensus as to the best approach and what parameters to use in choosing it. The three main approaches are conservative, endovascular and resection - the treatment of choice for acute abdominal bleeding. We report a case of hemoperitoneum following the spontaneous rupture of an unrecognized HCV-related HCC in a patient with no history of liver disease. The patient was successfully treated by emergency surgery, with resection of two segments of the left liver.

Role of emergency thoracic ultrasonography in spontaneous pneumomediastinum. Two case report

Early identification of spontaneous pneumomediastinum in an Emergency Department is possible with thoracic ultrasound. We report two cases of spontaneous pneumomediastinum, diagnosed in a 26-year old man with chronic asthma and a 19-year old athlete, and discuss the role of thoracic US alongside conventional X-ray and thoracic CT in emergency medicine. The patients were transferred to an Emergency Department, where conservative treatment produced a good outcome. The greater sensitivity and specificity of thoracic US over conventional supine X-ray in the detection of occult pneumothorax is ever more appreciated. However, training in the diagnosis of pneumomediastinum is required.

Gait Regulation for Bipedal Locomotion

This work explores regulation of forward speed, step length, and slope walking for the passive-dynamic class of bipedal robots. Previously, an energy-shaping control for regulating forward speed has appeared in the literature; here we show that control to be a special case of a more general time-scaling control that allows for speed transitions in arbitrary time. As prior work has focused on potential energy shaping for fully actuated bipeds, we study in detail the shaping of kinetic energy for bipedal robots, giving special treatment to issues of underactuation. Drawing inspiration from features of human walking, an underactuated kinetic-shaping control is presented that provides efficient regulation of walking speed while adjusting step length. Previous results on energetic symmetries of bipedal walking are also extended, resulting in a control that allows regulation of speed and step length while walking on any slope. Finally we formalize the optimal gait regulation problem and propose a dynamic programming solution seeded with passive-dynamic limit cycles. Observations of the optimal solutions generated by this method reveal further similarities between passive dynamic walking and human locomotion and give insight into the structure of minimum-effort controls for walking.

Evaluating alternative gait strategies using evolutionary robotics

Evolutionary robitics is a branch of artificial intelligence concerned with the automatic generation of autonomous robots. Usually the form of the robit is predefined an various computational techniques are used to control the machine's behaviour. One aspect is the spontaneous generation of walking in legged robots and this can be used to investigate the mechanical requiements for efficient walking in bipeds. This paper demonstrates a bipedal simulator that spontaneously generates walking and running gaits. The model can be customized to represent a range of hominoid morphologies and used to predict performance paramets such as preferred speed and metabolic energy cost. Because it does not require any motion capture data it is particularly suitable for investigating locomotion in fossil animals. The predictoins for modern humans are highly accurate in terms of energy cost for a given speend and thus the values predicted for other bipeds are likely to be good estimates. To illustrate this the cost of transport is calculated for Australopithecus afarensis. The model allows the degree of maximum extension at the knee to be varied causing the model to adopt walking gaits varying from chimpanzee-like to human=like. The energy costs associated with these gait choices can thus be calculated and this information used to evaluate possible locomotor strategies in early hominids