From nucleus to mitochondria : a ubiquitination study

Ubiquitination is a post-translational modification achieved by an enzymatic cascade. This post-translational modification is involved in many crucial cellular processes across different cellular compartments such as protein turnover via the ubiquitin proteasome system and various signaling pathways from the DNA damage response to the immune response. Ubiquitin has seven lysine residues (Lys 6, 11, 27, 29, 33, 48 and 63) onto which other ubiquitin moieties can be conjugated, forming ubiquitin chains of different types. These different ubiquitin chains can have different functions and the relationship between both is often referred to as “the ubiquitin code”. Although the enzymatic cascade leading to ubiquitination of proteins is well described, the ubiquitin code remains largely unresolved. Accumulation of insoluble ubiquitinated proteins is a hallmark of neurodegenerative disorders such as Parkinson’s and Alzheimer’s disease, making the study of these cellular processes relevant to human health. Specific proteins can also impair the function of the proteasome such as progerin in the Hutchinson-Gilford Progeria Syndrome (HGPS). The importance of ubiquitination in many cellular processes and its involvement in many human pathologies inspired us to develop an inducible ubiquitination system that could be used as a tool to better understand the ubiquitin code and its role in different cellular compartments. In paper I, we have engineered a ubiquitin ligase, ProxE3, which assembles specific ubiquitin chain (lysine 63) onto a fluorescent substrate. We have used this tool to generate ubiquitin chains on the surface of mitochondria and investigate mitophagy, more precisely if lysine 63 (K63) ubiquitin chains are sufficient to trigger aggregation of mitochondria or mitophagy. Upon successful ubiquitination of the surface of mitochondria by ProxE3 and depolarization of mitochondria by carbonyl cyanide mchlorophenyl hydrazone (CCCP), peri-nuclear clustering of mitochondria was observed but not mitophagy. The lack of mitophagy indicates that either the amount of K63 ubiquitination is insufficient in our system, that other types of ubiquitin chains are required, that a specific substrate need to be ubiquitinated or that the PINK1 feedforward loop is essential for mitophagy. Nonetheless, this work presents a valid tool for studies of ubiquitination in living cells, while reaffirming the complexity of the regulation of mitophagy. In paper II, we were interested in ubiquitination in a different cellular compartment: the nucleus. In this paper, we used a human cell-line expressing a fluorescent proteasomal substrate to investigate if the ubiquitin-proteasome system was impaired upon overexpression of progerin. Progerin is a mutated form of lamin A which is the cause of HGPS. It has been suspected that progerin might inhibit the catalytic activity of the proteasome, which could lead to neuronal dysfunction. However, we did not detect proteasomal impairment in human cells overexpressing progerin compared to wild-type lamin A. This observation is further supported by the lack of progerin/lamin A inclusions in hippocampal neurons of HGPS mice, implying that the ubiquitin/proteasome system is not sensitive to the expression of progerin in neurons of mice

Biophotonic Tools in Cell and Tissue Diagnostics.

In order to maintain the rapid advance of biophotonics in the U.S. and enhance our competitiveness worldwide, key measurement tools must be in place. As part of a wide-reaching effort to improve the U.S. technology base, the National Institute of Standards and Technology sponsored a workshop titled "Biophotonic tools for cell and tissue diagnostics." The workshop focused on diagnostic techniques involving the interaction between biological systems and photons. Through invited presentations by industry representatives and panel discussion, near- and far-term measurement needs were evaluated. As a result of this workshop, this document has been prepared on the measurement tools needed for biophotonic cell and tissue diagnostics. This will become a part of the larger measurement road-mapping effort to be presented to the Nation as an assessment of the U.S. Measurement System. The information will be used to highlight measurement needs to the community and to facilitate solutions

Origin Gaps and the Eternal Sunshine of the Second-Order Pendulum

The rich experiences of an intentional, goal-oriented life emerge, in an unpredictable fashion, from the basic laws of physics. Here I argue that this unpredictability is no mirage: there are true gaps between life and non-life, mind and mindlessness, and even between functional societies and groups of Hobbesian individuals. These gaps, I suggest, emerge from the mathematics of self-reference, and the logical barriers to prediction that self-referring systems present. Still, a mathematical truth does not imply a physical one: the universe need not have made self-reference possible. It did, and the question then is how. In the second half of this essay, I show how a basic move in physics, known as renormalization, transforms the "forgetful" second-order equations of fundamental physics into a rich, self-referential world that makes possible the major transitions we care so much about. While the universe runs in assembly code, the coarse-grained version runs in LISP, and it is from that the world of aim and intention grows.Comment: FQXI Prize Essay 2017. 18 pages, including afterword on Ostrogradsky's Theorem and an exchange with John Bova, Dresden Craig, and Paul Livingsto

Experimental investigation of the impact of elastic turbulence on heat transfer in a serpentine channel

AbstractThe characteristics of convective heat transfer and fluid flow within a square cross-section serpentine channel are experimentally studied for two groups of polymeric viscoelastic fluids, shear-thinning and constant-viscosity Boger solutions. The elastic turbulence can be created by the non-linear interaction between elastic stresses generated within the flowing high-molecular-weight polymer solutions and the streamline curvature. In order to confirm elastic turbulence in this geometry, pressure drop across the serpentine channel was measured. The findings indicate that the measurements of non-dimensional pressure-drop increase approximately from 1.48 to 4.82 for viscoelastic solutions compared with the Newtonian fluid over a range of Weissenberg number from 4 to 211. The convective heat transfer enhances due to elastic turbulence by up to 200% for low polymer concentration (dilute) solutions and reaches up to 380% for higher polymer concentration (semi-dilute) solutions under creeping-flow conditions in comparison to that achieved by the equivalent Newtonian fluid flow at low Graetz number (up to 14.6). We propose a modified Weissenberg number which is able to approximately collapse the mean Nusselt number data for each solution group

Quantification of mesoscale variability and geometrical reconstruction of a textile

Automated image analysis of textile surfaces allowed determination and quantification of intrinsic yarn path variabilities in a 2/2 twill weave during the lay-up process. The yarn paths were described in terms of waves and it was found that the frequencies are similar in warp and weft directions and hardly affected by introduced yarn path deformations. The most significant source of fabric variability was introduced during handling before cutting. These resulting systematic deformations will need to be considered when designing or analysing a composite component. An automated method for three dimensional reconstruction of the analysed lay-up was implemented in TexGen which will allow virtual testing of components in the future

Off-axis digital flash photography: a common cause of artefact leukocoria in children.

Leukocoria, a presenting sign in several significant pediatric ocular conditions, can be artefactually produced by off-axis flash photography in healthy eyes. The authors demonstrate the conditions needed to produce this phenomenon, which is more commonly seen in children due to their larger-sized pupils, photogenicity, and frequent off-axis shots

Longitudinal study of trachomatous trichiasis in The Gambia: barriers to acceptance of surgery.

PURPOSE: Investigation of compliance with surgery for trachomatous trichiasis has become a priority of the World Health Organization. This study was conducted to investigate attitudes toward trichiasis and its treatment and to determine the rate of surgical uptake in The Gambia. METHODS: A 1-year longitudinal study was performed in 190 subjects with trichiasis. Persons with major trichiasis (involving five lashes or more) were referred for surgery, and those with minor trichiasis were advised to epilate. Outcome measures included attitudes toward trichiasis and its treatment, reported barriers to surgical uptake, acceptance rates for surgery, and factors affecting acceptance. RESULTS: Twenty-three percent (95% confidence interval [CI] 16.5%-30.6%) of subjects with major trichiasis attended for surgery during the year. Degree of ignorance about surgery, symptoms impeding work, and a multiple income source for the head of household predicted attendance. Reported lack of time predicted nonattendance. Sixty-eight percent of patients who had undergone surgery were trichiasis free at last follow-up. CONCLUSIONS: Poor attendance for surgery remains a problem in The Gambia. Barriers include ignorance and lack of time and money. Health education and surgical delivery strategies are needed to overcome these barriers. Regular audit of surgical results is necessary, with retraining where indicated

Wake Development behind Paired Wings with Tip and Root Trailing Vortices: Consequences for Animal Flight Force Estimates

Recent experiments on flapping flight in animals have shown that a variety of unrelated species shed a wake behind left and right wings consisting of both tip and root vortices. Here we present an investigation using Particle Image Velocimetry (PIV) of the behaviour and interaction of trailing vortices shed by paired, fixed wings that simplify and mimic the wake of a flying animal with a non-lifting body. We measured flow velocities at five positions downstream of two adjacent NACA 0012 aerofoils and systematically varied aspect ratio, the gap between the wings (corresponding to the width of a non-lifting body), angle of attack, and the Reynolds number. The range of aspect ratios and Reynolds number where chosen to be relevant to natural fliers and swimmers, and insect flight in particular. We show that the wake behind the paired wings deformed as a consequence of the induced flow distribution such that the wingtip vortices convected downwards while the root vortices twist around each other. Vortex interaction and wake deformation became more pronounced further downstream of the wing, so the positioning of PIV measurement planes in experiments on flying animals has an important effect on subsequent force estimates due to rotating induced flow vectors. Wake deformation was most severe behind wings with lower aspect ratios and when the distance between the wings was small, suggesting that animals that match this description constitute high-risk groups in terms of measurement error. Our results, therefore, have significant implications for experimental design where wake measurements are used to estimate forces generated in animal flight. In particular, the downstream distance of the measurement plane should be minimised, notwithstanding the animal welfare constraints when measuring the wake behind flying animals