A Functional Screen Provides Evidence for a Conserved, Regulatory, Juxtamembrane Phosphorylation Site in Guanylyl Cyclase A and B

Kinase homology domain (KHD) phosphorylation is required for activation of guanylyl cyclase (GC)-A and -B. Phosphopeptide mapping identified multiple phosphorylation sites in GC-A and GC-B, but these approaches have difficulty identifying sites in poorly detected peptides. Here, a functional screen was conducted to identify novel sites. Conserved serines or threonines in the KHDs of phosphorylated receptor GCs were mutated to alanine and tested for reduced hormone to detergent activity ratios. Mutation of Ser-489 in GC-B to alanine but not glutamate reduced the activity ratio to 60% of wild type (WT) levels. Similar results were observed with Ser-473, the homologous site in GC-A. Receptors containing glutamates for previously identified phosphorylation sites (GC-A-6E and GC-B-6E) were activated to ∼20% of WT levels but the additional glutamate substitution for S473 or S489 increased activity to near WT levels. Substrate-velocity assays indicated that GC-B-WT-S489E and GC-B-6E-S489E had lower Km values and that WT-GC-B-S489A, GC-B-6E and GC-B-6E-S489A had higher Km values than WT-GC-B. Homologous desensitization was enhanced when GC-A contained the S473E substitution, and GC-B-6E-S489E was resistant to inhibition by a calcium elevating treatment or protein kinase C activation – processes that dephosphorylate GC-B. Mass spectrometric detection of a synthetic phospho-Ser-473 containing peptide was 200–1300-fold less sensitive than other phosphorylated peptides and neither mass spectrometric nor 32PO4 co-migration studies detected phospho-Ser-473 or phospho-Ser-489 in cells. We conclude that Ser-473 and Ser-489 are Km-regulating phosphorylation sites that are difficult to detect using current methods

Style, Function and Cultural Transmission

Recent evolutionary approaches to the understanding of lithic variability take us back to long-standing issues in lithic studies to do with the claimed contrast between style and function and the Binford-Bordes debate of the 1960s concerning the factors that affect inter-assemblage variation. In fact, the style and function contrast is an unhelpful one, not least when considering the question of convergence. Taking the definition of style as ‘a way of doing’, all functions are carried out in locally specific ways that have a transmission history, although the extent to which the history of the attributes relevant to the function have been subject to random drift and innovation patterns, as opposed to selection, will vary. Moreover, in a subtractive technology like lithics the extent to which a transmission signal will be visible in an attribute like the angle of a cutting edge is unclear. The contrasting view is that, in the case of lithics, functional requirements will always call into existence the technical innovations to satisfy them, which in any case are not that difficult to find. The paper addresses these and related issues with reference to previous work by Shennan and colleagues on the use of material culture to identify within and between group variation, the extent to which isolation-by-distance in space and time can account for the similarities and differences between assemblages, and the role of phylogenetic methods

Pneumococcal protein antigen serology varies with age and may predict antigenic profile of colonizing isolates

BACKGROUND: Several Streptococcus pneumoniae proteins play a role in pathogenesis and are being investigated as vaccine targets. It is largely unknown whether naturallyacquired antibodies reduce the risk of colonization with strains expressing a particular antigenic variant. METHODS: Serum IgG titers to 28 pneumococcal protein antigens were measured among 242 individuals, aged < 6 months - 78 years in Native American communities between 2007-2009. Nasopharyngeal swabs were collected at least 30 days after serum collection, and the protein antigen variant in each pneumococcal isolate was determined using genomic data. We assessed the association between preexisting variant-specific antibody titers and subsequent carriage of pneumococcus expressing a particular antigen variant. RESULTS: Antibody titers often increased across pediatric groups before decreasing among adults. PspA and StkP IgG titers decreased from <6 months to 6-12 months (p<0.01). Individuals with low titers against Group 3 PspC variants were more likely to be colonized with pneumococci expressing those variants. For other antigens, variantspecific IgG titers do not predict colonization with pneumococci expressing particular variants CONCLUSION: We observed an inverse association between variant-specific antibody concentration and homologous pneumococcal colonization for only one protein. Further assessment of antibody repertoires may elucidate the nature of anti-pneumococcal antibody-mediated mucosal immunity while informing future vaccine development

The context of HIV risk behaviours among HIV-positive injection drug users in Viet Nam: Moving toward effective harm reduction

<p>Abstract</p> <p>Background</p> <p>Injection drug users represent the largest proportion of all HIV reported cases in Viet Nam. This study aimed to explore the perceptions of risk and risk behaviours among HIV-positive injection drug users, and their experiences related to safe injection and safe sex practices.</p> <p>Methods</p> <p>This study used multiple qualitative methods in data collection including in-depth interviews, focus group discussions and participant observation with HIV-positive injection drug users.</p> <p>Results</p> <p>The informants described a change in the sharing practices among injection drug users towards more precautions and what was considered 'low risk sharing', like sharing among seroconcordant partners and borrowing rather than lending. However risky practices like re-use of injection equipment and 'syringe pulling' i.e. the use of left-over drugs in particular, were frequently described and observed. Needle and syringe distribution programmes were in place but carrying needles and syringes and particularly drugs could result in being arrested and fined. Fear of rejection and of loss of intimacy made disclosure difficult and was perceived as a major obstacle for condom use among recently diagnosed HIV infected individuals.</p> <p>Conclusion</p> <p>HIV-positive injection drug users continue to practice HIV risk behaviours. The anti-drug law and the police crack-down policy appeared as critical factors hampering ongoing prevention efforts with needle and syringe distribution programmes in Viet Nam. Drastic policy measures are needed to reduce the very high HIV prevalence among injection drug users.</p

Outcome measurement in clinical trials for Ulcerative Colitis: towards standardisation

Clinical trials on novel drug therapies require clear criteria for patient selection and agreed definitions of disease remission. This principle has been successfully applied in the field of rheumatology where agreed disease scoring systems have allowed multi-centre collaborations and facilitated audit across treatment centres. Unfortunately in ulcerative colitis this consensus is lacking. Thirteen scoring systems have been developed but none have been properly validated. Most trials choose different endpoints and activity indices, making comparison of results from different trials extremely difficult. International consensus on endoscopic, clinical and histological scoring systems is essential as these are the key components used to determine entry criteria and outcome measurements in clinical trials on ulcerative colitis. With multiple new therapies under development, there is a pressing need for consensus to be reached

Downregulation of the anaphase-promoting complex (APC)7 in invasive ductal carcinomas of the breast and its clinicopathologic relationships

INTRODUCTION: The anaphase-promoting complex (APC) is a multiprotein complex with E3 ubiquitin ligase activity, which is required for the ubiquitination of securin and cyclin-B. Moreover, the mitotic spindle checkpoint is activated if APC activation is prevented. In addition, several APC-targeting molecules such as securin, polo-like kinase, aurora kinase, and SnoN have been reported to be oncogenes. Therefore, dysregulation of APC may be associated with tumorigenesis. However, the clinical significance and the involvement of APC in tumorigenesis have not been investigated. METHODS: The expression of APC7 was immunohistochemically investigated in 108 invasive ductal carcinomas of the breast and its relationship with clinicopathologic parameters was examined. The expression of APC7 was defined as positive when the summed scores of staining intensities (0 to 3+) and stained proportions (0 to 3+) exceeded 3+. RESULTS: Positive APC7 expression was less frequent than its negative expression when histologic (P = 0.009) or nuclear grade (P = 0.009), or mitotic number (P = 0.0016) was elevated. The frequency of APC7 negative expression was higher in high Ki-67 or aneuploid groups than in low Ki-67 or diploid groups. CONCLUSION: These data show that loss of APC7 expression is more common in breast carcinoma cases with poor prognostic parameters or malignant characteristics. They therefore suggest that dysregulation of APC activity, possibly through downregulation of APC7, may be associated with tumorigenesis in breast cancer

The correlates of HIV testing and impacts on sexual behavior: evidence from a life history study of young people in Kisumu, Kenya

<p>Abstract</p> <p>Background</p> <p>HIV counseling and testing is considered an important component of HIV prevention and treatment. This paper examines the characteristics of young males and females at the time of first reported HIV test, including the influence of recent sexual partnerships, and investigates how HIV testing and the cumulative number of tests are associated with sexual behaviors within six months of testing.</p> <p>Methods</p> <p>The study uses data from a random sample of youth aged 18-24 years living in Kisumu, Kenya, who were interviewed using a 10-year retrospective life history calendar. Cox regression models were used to examine the correlates of the timing of first HIV test. Variance-correction models for unordered repeated events were employed to examine whether having an HIV test in the previous six months and the cumulative number of tests predict unsafe sexual practices in a given month.</p> <p>Results</p> <p>Sixty-four percent of females and 55% of males reported at least one HIV test in the last 10 years and 40% of females were pregnant the month of first test. Significant correlates of first HIV test included marital aspirations among non-pregnant females, unprotected sex in the previous six months among pregnant females, and concurrency in the previous six months among males. Having a recent HIV test was associated with a decreased likelihood of unprotected sex among ever-pregnant females, an increased likelihood of unprotected sex and "risky" sexual partnerships among never-pregnant females, and an increased likelihood of concurrency among males. Repeated HIV testing was associated with a lower likelihood of concurrency among males and involvement in "risky" sexual partnerships among males and never-pregnant females.</p> <p>Conclusions</p> <p>The high rate of pregnancy at first test suggests that promotion of HIV testing as part of prevention of mother-to-child transmission is gaining success. Further research is warranted to examine how and why behavior change is influenced by client- versus provider-initiated testing. The influence of different sexual partnership variables for males and females suggests that interventions to assess risk and promote testing should be gender- and relationship-specific. The findings also suggest that encouraging repeat or routine testing could potentially increase the uptake of safer sexual behaviors.</p

Wolbachia-Mediated Male Killing Is Associated with Defective Chromatin Remodeling

Male killing, induced by different bacterial taxa of maternally inherited microorganisms, resulting in highly distorted female-biased sex-ratios, is a common phenomenon among arthropods. Some strains of the endosymbiont bacteria Wolbachia have been shown to induce this phenotype in particular insect hosts. High altitude populations of Drosophila bifasciata infected with Wolbachia show selective male killing during embryonic development. However, since this was first reported, circa 60 years ago, the interaction between Wolbachia and its host has remained unclear. Herein we show that D. bifasciata male embryos display defective chromatin remodeling, improper chromatid segregation and chromosome bridging, as well as abnormal mitotic spindles and gradual loss of their centrosomes. These defects occur at different times in the early development of male embryos leading to death during early nuclear division cycles or large defective areas of the cellular blastoderm, culminating in abnormal embryos that die before eclosion. We propose that Wolbachia affects the development of male embryos by specifically targeting male chromatin remodeling and thus disturbing mitotic spindle assembly and chromosome behavior. These are the first observations that demonstrate fundamental aspects of the cytological mechanism of male killing and represent a solid base for further molecular studies of this phenomenon