Sarah Wadd

Herpes simplex virus ICP27 protein provides viral mRNAs with access to the cellular mRNA export pathway.

The role of herpes simplex virus ICP27 protein in mRNA export is investigated by microinjection into Xenopus laevis oocytes. ICP27 dramatically stimulates the export of intronless viral mRNAs, but has no effect on the export of cellular mRNAs, U snRNAs or tRNA. Use of inhibitors shows, in contrast to previous suggestions, that ICP27 neither shuttles nor exports viral mRNA via the CRM1 pathway. Instead, ICP27‐mediated viral RNA export requires REF and TAP/NXF1, factors involved in cellular mRNA export. ICP27 binds directly to REF and complexes containing ICP27, REF and TAP are found in vitro and in virally infected cells. A mutant ICP27 that does not interact with REF is inactive in viral mRNA export. We propose that ICP27 associates with viral mRNAs and recruits TAP/NXF1 via its interaction with REF proteins, allowing the otherwise inefficiently exported viral mRNAs to access the TAP‐mediated export pathway. This represents a novel mechanism for export of viral mRNAs.

Herpes Simplex Virus IE63 (ICP27) Protein Interacts with Spliceosome-Associated Protein 145 and Inhibits Splicing prior to the First Catalytic Step

ABSTRACT The multifunctional herpes simplex virus type 1 (HSV-1) protein IE63 (ICP27) interacts with the essential pre-mRNA splicing factor, spliceosome-associated protein 145 (SAP145), and in infected cells IE63 and SAP145 colocalize. This interaction was reduced or abrogated completely using extracts from cells infected with IE63 viral mutants, with mutations in IE63 KH and Sm homology domains, which do not exhibit host shutoff or inhibit splicing. In the presence of IE63, splicing in vitro was inhibited prior to the first catalytic step and the B/C complex formed during splicing was shifted up in mobility and reduced in intensity. With the use of splicing extracts, IE63 and SAP145 both comigrated with the B/C complex, suggesting that they interact within this complex to inhibit B/C complex formation or conversion. The inhibition of splicing may facilitate the export of viral or cellular transcripts, possibly via other protein partners of IE63. These data provide important new insights into how IE63 influences pre-mRNA processing during HSV-1 infection.

The Multifunctional Herpes Simplex Virus IE63 Protein Interacts with Heterogeneous Ribonucleoprotein K and with Casein Kinase 2

Herpes simplex virus type 1 (HSV-1), the prototype α-herpesvirus, causes several prominent diseases. The HSV-1 immediate early (IE) protein IE63 (ICP27) is the only regulatory gene with a homologue in every mammalian and avian herpesvirus sequenced so far. IE63 is a multifunctional protein affecting transcriptional and post-transcriptional processes, and it can shuttle from the nucleus to the cytoplasm. To identify interacting cellular proteins, a HeLa cDNA library was screened in the yeast two-hybrid system using IE63 as bait. Several interacting proteins were identified including heterogeneous nuclear ribonucleoprotein K (hnRNP K), a multifunctional protein like IE63, and the β subunit of casein kinase 2 (CK2), a protein kinase, and interacting regions were mapped. Confirmation of interactions was provided by fusion protein binding assays, co-immunoprecipitation from infected cells, and CK2 activity assays. hnRNP K co-immunoprecipitated from infected cells with anti-IE63 serum was a more rapidly migrating subfraction than hnRNP K immunoprecipitated by anti-hnRNP K serum. Using anti-IE63 serum, both IE63 and hnRNP K were phosphorylated in vitro by CK2, while in immunoprecipitates using anti-hnRNP K serum, IE63 but not hnRNP K was phosphorylated by CK2. These data provide important new insights into how this key viral regulatory protein exerts its functions.

Prevalence of, and risk factors for, hepatitis C virus infection among recent initiates to injecting in London and Glasgow: cross sectional analysis.

Summary.  Our aim was to compare the prevalence of antibody to hepatitis C virus (anti‐HCV) among recently initiated injecting drug users (IDUs) in London and Glasgow, and to identify risk factors which could explain differences in prevalence between the cities. Complementary studies of community recruited IDUs who had initiated injection drug use since 1996 were conducted during 2001–2002. Data on HCV risk behaviours were gathered using structured questionnaires with identical core questions and respondents were asked to provide an oral fluid specimen which was tested anonymously for anti‐HCV but was linked to the questionnaire. Sensitivities of the anti‐HCV assays for oral fluid were 92–96%. Prevalence of anti‐HCV was 35% (122/354) in London and 57% (207/366) in Glasgow (P < 0.001). Multifactorially, factors significantly associated with raised odds of anti‐HCV positivity were increasing length of injecting career, daily injection, polydrug use, having had a needlestick injury, and having served a prison sentence. In addition lower odds of anti‐HCV positivity were associated with non‐injection use of crack cocaine and recruitment from drug agencies. After adjustment for these factors, the increased odds of anti‐HCV associated with being a Glasgow IDU were diminished but remained significant. HCV continues to be transmitted among the IDU population of both cities at high rates despite the availability of syringe exchange and methadone maintenance. Effectiveness of harm reduction interventions may be compromised by inadequate coverage and failure to reduce sufficiently the frequency of sharing different types of injecting equipment, as well as the high background prevalence of HCV, and its high infectivity. Comprehensive action is urgently required to reduce the incidence of HCV among injectors.

Interaction between Herpes Simplex Virus Type 1 IE63 Protein and Cellular Protein p32

ABSTRACT The herpes simplex virus type 1 (HSV-1) immediate-early gene IE63 (ICP27), the only HSV-1 regulatory gene with a homologue in every mammalian and avian herpesvirus sequenced so far, is a multifunctional protein which regulates transcriptional and posttranscriptional processes. One of its posttranscriptional effects is the inhibition of splicing of viral and cellular transcripts. We previously identified heterogeneous nuclear ribonucleoprotein (hnRNP) K and casein kinase 2 (CK2) as two protein partners of IE63 (H. Bryant et al., J. Biol. Chem. 274:28991–28998, 1999). Here, using a yeast two-hybrid assay, we identify another partner of IE63, the cellular protein p32. Confirmation of this interaction was provided by coimmunoprecipitation from virus-infected cells and recombinant p32 binding assays. A p32-hnRNP K-CK2 complex, which required IE63 to form, was isolated from HSV-1-infected cells, and coimmunoprecipitating p32 was phosphorylated by CK2. Expression of IE63 altered the cytoplasmic distribution of p32, with some now colocalizing with IE63 in the nuclei of infected and transfected cells. As p32 copurifies with splicing factors and can inhibit splicing, we propose that IE63 together with p32, possibly with other IE63 partner proteins, acts to disrupt or regulate pre-mRNA splicing. As well as contributing to host cell shutoff, this effect could facilitate splicing-independent nuclear export of viral transcripts.

Hepatitis C virus infection among injecting drug users in Scotland: a review of prevalence and incidence data and the methods used to generate them

It is estimated that of 50,000 persons in Scotland (1% of the countys population), infected with the hepatitis C virus (HCV), around 90% injected drugs. This paper reviews data on the prevalence and incidence of HCV, and the methods used to generate such information, among injecting drug users (IDUs), in Scotland. The prevalence estimate for HCV among IDUs in Scotland as a whole (44% in 2000), is comparable with those observed in many European countries. Incidence rates ranged from 11.9 to 28.4/100 person-years. The data have shaped policy to prevent infection among IDUs and have informed predictions of the number of HCV-infected IDUs who will likely progress to, and require treatment and care for, severe HCV-related liver disease. Although harm reduction interventions, in particular needle and syringe exchanges and methadone maintenance therapy, reduced the transmission of HCV among IDUs during the early to mid-1990s, incidence in many parts of the country remains high. The prevention of HCV among IDUs continues to be one of Scotlands major public health challenges.

Drinking behaviour and alcohol-related harm amongst older adults: analysis of existing UK datasets

BackgroundOlder adults experience age-related physiological changes that increase sensitivity and decrease tolerance to alcohol and there are a number of age-related harms such as falls, social isolation and elder abuse, which are compounded by alcohol misuse. Despite this unique vulnerability and the fact that the number of older adults is increasing, the literature on drinking behaviour and alcohol-related harm in older adults is sparse. This article describes a secondary analysis of UK data to address this knowledge gap.MethodSecondary analysis of national statistics on alcohol-related hospital admissions and alcohol-related deaths, and data on drinking behaviour from the General Lifestyle Survey. Trends were identified by calculating percentage changes between time periods. The association between drinking behaviour and selected age groups was investigated using one way analysis of variance or chi-square tests.ResultsOlder adults (aged 65 and over) drink less and are less likely to exceed the recommended drink limits than younger adults. However, they are more likely to be admitted to hospital for an alcohol-related condition than younger adults and the most significant increases in alcohol-related hospital admission rates in recent years have occurred in older age groups. Alcohol-related death rates are highest amongst those aged 55–74 years old. Alcohol consumption and the prevalence of exceeding the recommended drink limits has fluctuated but not significantly increased in older adults in recent decades.ConclusionOlder adults experience high and increasing levels of alcohol-related harm and as the population ages, this is likely to put increasing pressure on health and social services. Careful monitoring and age-appropriate strategies to detect and treat older adults at risk of alcohol-related harm are required.

Exploring associations between perceived HCV status and injecting risk behaviors among recent initiates to injecting drug use in Glasgow

The aim of this study was to explore the influence of testing for hepatitis C virus (HCV) and perceived HCV status on injecting risk behavior. A cross-sectional, community-wide survey was undertaken at multiple sites throughout Greater Glasgow during 2001–2002. Four hundred ninety-seven injecting drug users (IDUs) consented to participate and were interviewed using a structured questionnaire to ascertain HCV test history and injecting risk behavior. The average age of participants was 27 years and the majority of the sample were male (70.4%). Participants had been injecting for an average duration of 2.5 years. Logistic regression analysis revealed no significant associations between having been tested and injecting risk behavior. After adjustment for potential confounding variables, HCV-negatives were significantly less likely to borrow needles/syringes and spoons or filters as compared with unawares and were significantly less likely to borrow spoons or filters as compared with HCV-positives. Due to the cross-sectional design of the study, it is uncertain whether this reduction in risk behavior could be attributed to perception of HCV status. Further research is recommended to consolidate the evidence for this relationship.

Working with Older People with Alcohol Problems: Insight from Specialist Substance Misuse Professionals and their Service Users

Significant numbers of older people worldwide have a drinking level or pattern which places them at risk of harm. In England, older people are more likely to be admitted to hospital for an alcohol-related condition than younger people and levels of alcohol-related harm are increasing fastest in this population. Whilst alcohol problems in older people are highly treatable, they frequently go undetected or ignored. The aim of this study was to develop guidelines for health and social care workers on what intervention strategies are likely to work best with older drinkers. Insight from alcohol practitioners who specialise in working with older people and the perspectives of older people receiving alcohol treatment were gained through focus groups and individual interviews. This paper reports some of the key findings including a perception that health and social care workers often did not intervene when alcohol misuse was suspected because of ageist attitudes and false beliefs about older peoples drinking. Participants however acknowledged that social workers faced difficult choices in relation to the ‘right’ of older people with alcohol problems to continue to drink and the ‘risk’ associated with them doing so. The implications for social work education and training are discussed.

Alcohol Screening in People With Cognitive Impairment: An Exploratory Study

Objective: Alcohol misuse can coexist with and/or contribute to the development of cognitive impairment in the older adult population but continues to be underestimated and undetected in older people. This study aimed to examine the feasibility and acceptability of routine screening for alcohol misuse in a small sample of older people with cognitive impairment receiving services in memory clinics. Methods: This study employed a qualitative and exploratory design, using a convenience sample of individuals attending a memory clinic in England. Ten service users older than 65 with a diagnosis of cognitive impairment (i.e., mild cognitive impairment or dementia) took part in the study. Individuals who met inclusion criteria were invited to take part in an hour-long interview, which included the interviewer administering the alcohol screening tools. Interview transcripts were analyzed using thematic analysis. Results: Participants were able to engage with the screening tools and could, with assistance, complete them in a collaborative and timely manner without distress. All participants reported that these tools were acceptable as part of the clinic assessment. Administering the screening tools was not time-consuming or difficult, making their use feasible within the memory clinic setting. While there were some challenges (e.g., arithmetic, recall, language problems), these challenges could be overcome with the aid of the person administering the screening tool using standardized techniques for assessment administration. Conclusions: Routine screening for alcohol misuse in older people with cognitive impairment receiving services in memory clinics is feasible and acceptable. The process of completing alcohol screening tools with older adults receiving services at memory clinics may increase awareness of the potential impact of alcohol on cognitive functioning and provide practitioners with an opportunity to educate service users about the ways that their drinking is affecting their memory. Several techniques to facilitate completion of screening tools were identified. Future research should evaluate the reliability and validity of alcohol screening tools with older people through corroborating screening results with other assessment methods.