Glycoprotein M and ESCRT in herpes simplex virus type 1 assembly

Herpes simplex virus type 1 (HSV-1) has a large linear double-stranded DNA genome in an icosahedral capsid shell, a cell-derived lipid envelope and a proteinaceous tegument layer. There are over fifty viral proteins and many host proteins identified in HSV-1 virions. The final formation of mature virus particles requires the membrane wrapping of tegumented capsids in the cytoplasm, a process termed secondary envelopment. This process involves the coordination of numerous viral and cellular proteins and results in double-membrane structures with enveloped virions contained within cellular vesicles. Mature viruses are then released through the fusion of these virion-containing vesicles and plasma membranes. This thesis describes investigation into the functions of viral glycoprotein M (gM) and the cellular Endosomal Sorting Complexes Required for Transport (ESCRT) in secondary envelopment. Firstly, it has been reported that gH/L can be efficiently internalised and targeted to the TGN by the co-expression of gM in transfection assays. In order to examine the role of gM in guiding the localisation of viral proteins in infected cells, a HSV-1 gM deletion virus (∆gM), and its revertant virus were constructed. The major phenotype demonstrated was that the absence of gM caused the internalisation of cell surface gH/L to be inhibited and higher levels of gH/L to be observed on the cell surface. Further, lower levels of gH/L were detected in purified ∆gM virions, which was in agreement with the delayed entry kinetics, smaller plaque sizes and greater replication deficits at low multiplicity of infection observed in ∆gM infected cells. Over all the results presented in this thesis demonstrate that in infected cells the efficient incorporation of gH/L into virions relies on the function of gM in HSV-1. Secondly, during HSV-1 secondary envelopment the budding and scission of the viral envelope from the host membrane share topological similarities with the formation of intraluminal vesicle in multivesicular bodies, retrovirus budding, and abscission at the end of cytokinesis, processes that require the cellular ESCRT machinery. There are four multiprotein ESCRT complexes and many associated proteins involved in their regulation. It has been previously shown that the ESCRT-III complex and a functional ATPase VPS4 are required for HSV-1 secondary envelopment, but different from the strategy utilised by HIV-1, the recruitment of ESCRT during HSV-1 infection is independent of TSG101 and/or ALIX. Data presented in this thesis demonstrate that CHMP4A/B/C proteins of the ESCRT-III complex are specifically crucial for HSV-1 secondary envelopment. Simultaneous depletion of CHMP4A/B/C proteins significantly inhibited HSV-1 replication. Ultrastructure analysis revealed that there were virtually no extracellular virions in CHMP4A/B/C depleted samples while more free capsids were observed in the cytoplasm, although the nuclear capsids and primary envelopment events appeared to be normal. In order to identify interactions between HSV-1 and ESCRT proteins, 22 HSV-1 tegument proteins were cloned and tested against a panel of ESCRT and ESCRT-associated proteins in yeast two-hydrid assays. Analysis of positive hits from yeast two-hybrid interaction screens using GST pull-down, co-immunoprecipitation and protein co-localisation assays have validated interactions of pUL47 with CC2D1A/1B, CIN85, CHMP6 and ALIX, pUL46 and pUL49 with CC2D1A/1B and CIN85, and pUL16 with CC2D1A/1B. Furthermore, the newly identified ESCRT associated proteins CC2D1A and CC2D1B have been detected in purified virions. The role of the identified ESCRT proteins in HSV-1 replication has been investigated using siRNA depletion. Unfortunately siRNA depletions of the various ESCRT candidates individually or in combinations did not show any significant effect on HSV-1 replication. Overall these data suggest that unlike HIV and other retroviruses, HSV-1 has evolved multiple parallel pathways to hijack the ESCRT machinery to facilitate its replication, particularly, through the interactions that lead directly to the recruitment of CHMP4A/B/C proteins. Disruption of some of these pathways did not prevent HSV-1 replication in tissue culture, suggesting any one potential pathway is sufficient for ESCRT recruitment to sites of HSV-1 assembly.This work was supported by the Royal Society (UF090010) and the MRC Grant (G0700129). PhD studentship funding for Yudan Ren was offered jointly by Cambridge Overseas Trust, Department of Pathology, Robinson College, the Board of Graduate Studies and Mr. Xuefeng He

Photo inactivation of virus particles in microfluidic capillary systems

It has long been established that UVC light is a very effective method for inactivating pathogens in a fluid, yet the application of UVC irradiation to modern biotechnological processes is limited by the intrinsic short penetration distance of UVC light in optically dense protein solutions. This experimental and numerical study establishes that irradiating a fluid flowing continuously in a microfluidic capillary system, in which the diameter of the capillary is turned to the depth of penetration of UVC light, uniquely treats the whole volume of the fluid to UVC light resulting in fast and effective inactivation of pathogens, with particular focus to virus particles. This was demonstrated by inactivating human herpes simplex virus type-1 (HSV-1, a large enveloped virus) on a dense 10% fetal calf serum solution in a range of fluoropolymer capillary systems, including a 0.75 mm and 1.50 mm internal diameter capillaries and a high-throughput MicroCapillary Film with mean hydraulic diameter of 206 μm. Up to 99.96% of HSV-1 virus particles were effectively inactivated with a mean exposure time of up to 10s, with undetectable collateral damage to proteins. The kinetics of virus inactivation matched well the results from a new mathematical model that considers the parabolic flow profile in the capillaries, and showed the methodology is fully predictable and scalable and avoids both the side effect of UVC light to proteins and the dilution of the fluid in current tubular UVC inactivation systems. This is expected to speed up the industrial adoption of non-invasive UVC virus inactivation in clinical biotechnology and biomanufacturing of therapeutic molecules

The Natural Environmental Factors Influencing the Spatial Distribution of Marathon Events：A Case Study from China

Purpose: The purpose of this study was to investigate the influence of natural environmental factors on the spatial distribution of marathon events in China, and to identify the suitable natural environmental factors for the marathon events. Methods: Geographic information system (GIS) spatial analysis tools were used to perform coupling analysis, e.g. overlap, neighborhood, intersection and buffer for terrain, climate, air quality, mountains and water resources with 342 marathon events held in China in 2018. Results: The results indicate that the spatial distribution of marathon events in China is negatively correlated with the elevation of the terrain (plain > hill > plateau > mountain > basin); climate (subtropical monsoon climate > temperate monsoon climate > temperate continental climate > tropical monsoon climate > plateau alpine climate), air quality (level 3 > level 2 > level 4 > level 1). Results indicate that buffer zones can protect water resources: there are 24 items in the buffer zone of river 0.5 km and lake 1 km, 131 items in the buffer zone of river 3 km and lake 5 km, 191 items in the buffer zone of river 5 km and lake 10 km, 298 items in the buffer zone of river 10 km and lake 20 km. Results indicate for mountain range buffer: 13 items in the 20 km buffer and 39 items in the 50 km buffer. Conclusions: Marathon events are more likely to be held on the third rung of China’s topography where a city has a typical landform (plains, basins, hills, or mountain) with good climate and air quality. Meanwhile a city with water and mountain resources for recreational events such as cross-country or obstacle course are essential. The contribution of this study is to systematically and intuitively reflect the influence of natural environment factors on the distribution of marathon events in China, and to provide evidence for the medium and long-term planning of marathon events in China, the selection of venues for different types of marathon events and how to attract participants

The natural environmental factors influencing the spatial distribution of marathon event: A case study from China

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. Purpose: The purpose of this study was to investigate the influence of natural environmental factors on the spatial distribution of marathon events in China, and to identify the suitable natural environmental factors for the marathon events. Methods: Geographic information system (GIS) spatial analysis tools were used to perform coupling analysis, e.g. overlap, neighborhood, intersection and buffer for terrain, climate, air quality, mountains and water resources with 342 marathon events held in China in 2018. Results: The results indicate that the spatial distribution of marathon events in China is negatively correlated with the elevation of the terrain (plain \u3e hill \u3e plateau \u3e mountain \u3e basin); climate (subtropical monsoon climate \u3e temperate monsoon climate \u3e temperate continental climate \u3e tropical monsoon climate \u3e plateau alpine climate), air quality (level 3 \u3e level 2 \u3e level 4 \u3e level 1). Results indicate that buffer zones can protect water resources: there are 24 items in the buffer zone of river 0.5 km and lake 1 km, 131 items in the buffer zone of river 3 km and lake 5 km, 191 items in the buffer zone of river 5 km and lake 10 km, 298 items in the buffer zone of river 10 km and lake 20 km. Results indicate for mountain range buffer: 13 items in the 20 km buffer and 39 items in the 50 km buffer. Conclusions: Marathon events are more likely to be held on the third rung of China’s topography where a city has a typical landform (plains, basins, hills, or mountain) with good climate and air quality. Meanwhile a city with water and mountain resources for recreational events such as cross-country or obstacle course are essential. The contribution of this study is to systematically and intuitively reflect the influence of natural environment factors on the distribution of marathon events in China, and to provide evidence for the medium and long-term planning of marathon events in China, the selection of venues for different types of marathon events and how to attract participants

Mapping the time-varying functional brain networks in response to naturalistic movie stimuli

One of human brain’s remarkable traits lies in its capacity to dynamically coordinate the activities of multiple brain regions or networks, adapting to an externally changing environment. Studying the dynamic functional brain networks (DFNs) and their role in perception, assessment, and action can significantly advance our comprehension of how the brain responds to patterns of sensory input. Movies provide a valuable tool for studying DFNs, as they offer a naturalistic paradigm that can evoke complex cognitive and emotional experiences through rich multimodal and dynamic stimuli. However, most previous research on DFNs have predominantly concentrated on the resting-state paradigm, investigating the topological structure of temporal dynamic brain networks generated via chosen templates. The dynamic spatial configurations of the functional networks elicited by naturalistic stimuli demand further exploration. In this study, we employed an unsupervised dictionary learning and sparse coding method combing with a sliding window strategy to map and quantify the dynamic spatial patterns of functional brain networks (FBNs) present in naturalistic functional magnetic resonance imaging (NfMRI) data, and further evaluated whether the temporal dynamics of distinct FBNs are aligned to the sensory, cognitive, and affective processes involved in the subjective perception of the movie. The results revealed that movie viewing can evoke complex FBNs, and these FBNs were time-varying with the movie storylines and were correlated with the movie annotations and the subjective ratings of viewing experience. The reliability of DFNs was also validated by assessing the Intra-class coefficient (ICC) among two scanning sessions under the same naturalistic paradigm with a three-month interval. Our findings offer novel insight into comprehending the dynamic properties of FBNs in response to naturalistic stimuli, which could potentially deepen our understanding of the neural mechanisms underlying the brain’s dynamic changes during the processing of visual and auditory stimuli

Buli dan Hubungannya Dengan Prestasi Akademik di Sekolah Berasrama Penuh di Kedah

The purpose of this study is to describe gangterism in relation to bullying behavior and determine its relationship with students' academic achievement in three residential schools (SBP) in Kedah. The dimensions of bullying studied are types of bullying behavior, types of bullies and victims, places where bullying frequently happened either direct or indirect bullying and bullies and victims emotions and feelings. It is also attempted to identify the relationship between dimensions of bullying behavior and students' academic achievement. The samples of study are students from Form One to Form Five of the three selected SBPs in Kedah. Descriptive analysis and Pearson Correlation (r) are used for data analysis. Results showed that bullying incidents occured mainly in the hostel, cafeteria and bathroom or toilet more frequently compared to other places. More boys and groups of boys were involved in bullying compared to girls and the occurrence is more of indirect bullying. However, there are responses indicating that direct physical bully do exist though at a low rate but this call for appropriate action before the incident is beyond the control of the authority. Result of correlation analysis showed academic achievement was not significantly related with bullying and victims of bullying. Nevertheless, a total of 77 percent of 396 respondents belong to the hogh achievers group compared to 23 percent who belong to the low achievers group

Growing like China revisit : a quantitative analysis during 1998 -- 2015

The growth pattern of the Chinese economy since 1990s is very controversial: the high output growth is accompanied by sustained increases in returns on capital, investment rates, and saving rates. Song et al. (2011) have proposed a theory of economic transition which can comprehensively explain Chinese experience from 1992 to 2007. But from 2008 onwards, due to the global recession and financial crisis, the Chinese economy has experienced tremendous changes including government’s four-trillion-Yuan stimulus package, the reform of state-owned firms and the revolution in the capital market. Therefore, we modify Song’s model to mimic China’s economic development until 2015. The private firms operated by entrepreneurs are more productive, but the financial imperfections force them to rely on internal savings to raise capital. State-owned firms with low-productivity depend on their superiority in the financial market to remain in the game. As the financial market in China becomes more developed, financial frictions gradually decrease; meanwhile, the continuous growth of the saving rate increases entrepreneurial firms’ capital stock and relaxes the limits of the high-productivity firms’ growth. While the calibrated model, in general, captures the features of the Chinese economy, it no longer matches as perfectly as before. This is likely due to the government intervention, housing bubble, foreign direct investments and overseas investments.Bachelor of Art