Characterisation of leucine-rich repeat kinase-2 regulation and kinase function

Mutations in Leucine-Rich Repeat Kinase 2 (LRRK2) are one of the most common causes of genetic Parkinson’s disease (PD), with mutations thought to account for around 5% of all familial cases. LRRK2 is a large protein with a kinase and GTPase domain and multiple protein-­‐protein interaction domains. Regulation of this protein is complex, with GTPase activity known to regulate kinase activity. Similarly, LRRK2 can autophosphorylate and is thought to form a dimer when active. Mutations in LRRK2 are numerous, with the most prevalent mutations occurring in the enzymatic core of this protein. This thesis describes work done to characterise the regulation and functioning of LRRK2, in order to further contribute towards understanding how mutations in this protein can lead to the pathogenesis of PD. Using BlueNative PAGE and glycerol gradient centrifugation, the quaternary structure of LRRK2 was assessed. In vitro kinase assays were used to characterise kinase activity of recombinant LRRK2 and a number of putative kinase substrates were also investigated. Identification of new kinase substrates was attempted and immunoprecipitation of LRRK2 to identify novel binding partners was also performed. Results of these experiments showed that familial mutations do not affect the ability of LRRK2 to form complexes. Instead, some mutations are affecting the enzymatic activity of LRRK2. Dephosphorylating LRRK2 showed that dimer formation is dependent on phosphorylation. Dephosphorylated forms of LRRK2 were more likely to be monomeric and displayed lower kinase activity than higher molecular weight forms. In vitro kinase assays to evaluate LRRK2 kinase substrates showed that α-synuclein is phosphorylated at low levels by G2019S but not wild-type LRRK2. Attempts to identify novel kinase substrates and binding partners of LRRK2 were unsuccessful, however evaluation of putative kinase substrates in vitro showed that DVL3 and TUBB5 may be good candidates for further investigation, as they were robustly phosphorylated by LRRK2. These results contribute towards our understanding of how LRRK2 functions and future studies based on these results may prove useful in aiding our understanding of how LRRK2 can cause PD pathogenesis

Direct detection of blood nitric oxide reveals a burn-dependent decrease of nitric oxide in response to Pseudomonas aeruginosa infection

Burn injury is associated with severe immune dysfunction, including an anti-inflammatory state that occurs late after burn injury. While increased nitric oxide (NO) production is associated with severe infection and sepsis, the effect of burn trauma on these levels during a non-lethal infection remains unknown. We hypothesized that in a mouse model, 1) NO levels would be increased after infection without trauma and 2) burn injury would lead to decreased NO production even during infection

Riparian buffer strips influence nitrogen losses as nitrous oxide and leached N from upslope permanent pasture

Riparian buffer strips can have a significant role in reducing nitrogen (N) transfers from agricultural land to freshwater primarily via denitrification and plant uptake processes, but an unintended trade-off can be elevated nitrous oxide (N2O) production rates. Against this context, our replicated bounded plot scale study investigated N2O emissions from un-grazed ryegrass pasture served by three types of riparian buffer strips with different vegetation, comprising: (i) grass riparian buffer with novel deep-rooting species, (ii) willow (young trees at establishment phase) riparian buffer, and (iii) deciduous woodland (also young trees at establishment phase) riparian buffer. The experimental control was ryegrass pasture with no buffer strip. N2O emissions were measured at the same time as total oxidized N in run-off, and soil and environmental characteristics in the ri parian buffer strips and upslope pasture between 2018 and 2019. During most of the sampling days, the no-buffer control treatment showed significantly (P < 0.05) greater N2O fluxes and cumulative N2O emissions compared to the remainder of the treatments. Our results also showed that the grass riparian buffer strip is a sink of N2O equivalent to − 2310.2 g N2O-N ha− 1 day− 1 (95% confidence interval:− 535.5 to 492). Event-based water quality results obtained during storms (12 November 2018 and 11 February 2019) showed that the willow riparian buffer treatment had the highest flow-weighted mean N concentrations (N-FWMC) of 0.041 ± 0.022 and 0.031 ± 0.015 mg N L− 1, when compared to the other treatments. Our 9-month experiment therefore, shows that ri parian buffer strips with novel deep-rooting grass can therefore potentially address emissions to both water and air. The results imply that over a shorter timeline similar to the current study, the grass riparian buffer strip can potentially address N emission to both air and water, particularly when serving a permanent pasture in similar settings as the current experiment.Fil: Dlamini, J.C. Crop and Climate Sciences. Departament of Soil; Sudáfrica. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido. University of Pretoria. Department of Plant and Soil Sciences; SudáfricaFil: Cardenas, L.M. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Tesfamarian, E.H. University of Pretoria. Department of Plant and Soil Sciences; SudáfricaFil: Dunn, R.M. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Loick, N. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Charteris, A.F. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Cocciaglia, L. Università degli Studi di Perugia. Dipartimento di Scienze Agrarie, Alimentari e Ambientali; ItaliaFil: Vangeli, Sebastián. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Clima y Agua; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Ingeniería Agrícola y Uso de la Tierra. Cátedra de Manejo y Conservación de Suelo; ArgentinaFil: Blackwell, M.S.A. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Upadhayay, H.R. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Hawkins, J.M.B. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido.Fil: Evans, J. Rothamsted Research. Computational and Analytical Sciences; Reino UnidoFil: Collins, A.L. Rothamsted Research. Sustainable Agriculture Sciences; Reino Unido

Vulnerability to bullying in children with a history of specific speech and language difficulties

This is an electronic version of an article published in Lindsay, Geoff and Dockrell, Julie and Mackie, Clare (2008) Vulnerability to bullying in children with a history of specific speech and language difficulties. European Journal of Special Needs Education, 23 (1). pp. 1-16. European Journal of Special Needs Education is available online at: http://www.informaworld.com/10.1080/0885625070179120

Validated TRNSYS model for forced circulation solar water heating systems with flat plate and heat pipe evacuated tube collectors

This paper presents a validated TRNSYS model for forced circulation solar water heating systems used in temperate climates. The systems consist of two flat plate collectors (FPC) and a heat pipe evacuated tube collector (ETC) as well as identical auxiliary components. The systems were fitted with an automated unit that controlled the immersion heaters and hot water demand profile to mimic hot water usage in a typical European domestic dwelling. The main component of the TRNSYS model was the Type 73 FPC or Type 538 ETC. A comparison of modelled and measured data resulted in percentage mean absolute errors for collector outlet temperature, heat collected by the collectors and heat delivered to the load of 16.9%, 14.1% and 6.9% for the FPC system and 18.4%, 16.8% and 7.6% for the ETC system respectively. The model underestimated the collector outlet fluid temperature by -9.6% and overestimated the heat collected and heat delivered to load by 7.6% and 6.9% for the FPC system. The model overestimated all three parameters by 13.7%, 12.4% and 7.6% for the ETC system

Do NO, N2O, N2 and CO2 fluxes differ in soils sourced from cropland and varying riparian buffer vegetation? An incubation study

DATA AVAILABILITY STATEMENT : Data available from authors upon reasonable request.Please read abstract in the article.Biotechnology and Biological Sciences Research Council and National Research Foundation South Africa.http://www.wileyonlinelibrary.com/journal/sumhj2024Plant Production and Soil ScienceSDG-06:Clean water and sanitatio

Operationalising and measuring language dominance

The paper offers a new way to measure language ability in bilinguals, based on measures of lexical richness. The validity of proposed approach is tested in a variety of ways

One Loop Multiphoton Helicity Amplitudes

We use the solutions to the recursion relations for double-off-shell fermion currents to compute helicity amplitudes for $n$-photon scattering and electron-positron annihilation to photons in the massless limit of QED. The form of these solutions is simple enough to allow {\it all}\ of the integrations to be performed explicitly. For $n$-photon scattering, we find that unless $n=4$, the amplitudes for the helicity configurations (+++...+) and (-++...+) vanish to one-loop order.Comment: 27 pages + 4 uuencoded figures (included), Fermilab-Pub-93/327-T, RevTe