Expression of human cation-independent mannose 6-phosphate receptor cDNA in receptor-negative mouse P388D1 cells following gene transfer

We recently reported the cDNA cloning, sequence, and expression of the human cation-independent mannose 6-phosphate receptor (hCI-MPR) (Oshima, A., Nolan, C. M., Kyle, J. W., Grubb, J. H., and Sly, W. S. (1988) J. Biol. Chem. 263, 2553-2562). The sequence of the hCI-MPR was virtually identical to that of the human insulin-like growth factor II receptor cDNA (Morgan, D. O., Edman, J. C., Standring, D. N., Fried, V. A., Smith, M. C., Roth, R. A., and Rutter, W. J. (1987) Nature 329, 301-307). To test the role of the putative bifunctional receptor in intracellular sorting of acid hydrolases, we studied its effect on lysosomal enzyme transport following gene transfer to receptor-negative cells. Receptor-negative mouse P388D1 cells were transfected with a cDNA construct containing the entire coding sequence of hCI-MPR under the control of the mouse metallothionine I promoter. Stable transformants were isolated and characterized. The expressed hCI-MPR was localized in membranes including the plasma membrane, bound mannose 6-phosphate containing ligands, and mediated endocytosis which could be specifically blocked by mannose 6-phosphate. We next measured the effect of the expressed hCI-MPR on intracellular and secreted acid hydrolases. The intracellular activity of the lysosomal marker enzymes beta-glucuronidase and beta-hexosaminidase increased up to 2-fold following transformation. In addition, expression of the receptor greatly reduced the fraction of acid hydrolases secreted. These phenotypic changes in the transformed cell lines support the proposed role of the cation-independent mannose 6-phosphate receptor in intracellular sorting and targeting of lysosomal enzymes.     

No evidence for mycoviruses in Entomophthora egressa

Summary Two strains of Entomophthora egressa which differ in their pathogenicity towards the spruce budworm were surveyed for the presence of double-stranded RNA mycoviruses. There was no evidence for the occurrence of any mycovirus in either strain. This indicates that virulence in E. egressa is not associated with a mycovirus.     

Prevalence of asthma in Melbourne schoolchildren: changes over 26 years.

OBJECTIVES--To determine the prevalence of asthma in the past 12 months in Melbourne schoolchildren aged 7, 12, and 15 years and to compare the prevalence of a history of asthma with that of 26 years ago. DESIGN--A questionnaire on respiratory symptoms was distributed to children for completion by parents and return to the school. Subjects were selected by a stratified cluster design. SETTING--Government and non-government schools in the greater Melbourne area, Australia. SUBJECTS--10,981 children. Parents completed questionnaires for 3324 children aged 7, 2899 aged 12, and 2968 aged 15. The overall response rate was 90%. MAIN OUTCOME MEASURES--History of wheeze or asthma in the past 12 months and in lifetime. RESULTS--The prevalences of wheeze in the past 12 months were 23.1%, 21.7%, and 18.6% for 7, 12, and 15 year olds respectively. A history of wheeze was more common in boys than in girls at age 7 (443/1711 v 324/1614) and 12 (418/1767 v 322/1718) but not at age 15. Overall, 78% (1548) of those reporting wheeze also reported a history of asthma and 83% (1611) had used a bronchodilator. The prevalence of a history of asthma among 7 year olds was 46% compared with 19.1% in the 1964 survey, an increase of 141%. CONCLUSIONS--The current prevalence of asthma in Melbourne schoolchildren is high and has risen substantially over the past 26 years.     

Testosterone affects reproductive success by influencing extra-pair fertilizations in male dark-eyed juncos (Aves: Junco hyemalis)

Monogamous male birds typically allocate less effort to courtship and more to parental behaviour than males of polygynous species. The seasonal pattern of testosterone (T) secretion varies accordingly. Monogamous males exhibit a spring peak in plasma T followed by lower levels during the parental phase, while males of polygynous species continue to court females and maintain T at higher levels. To determine whether testosterone underlies the trade-off between mating and parental effort, we treated male dark-eyed juncos (Junco hyemalis) with exogenous T and compared the reproductive success (RS) of T-treated males (T-males) to that of controls. T-males had lower apparent annual RS than controls, probably because elevated T reduced parental care. Nevertheless, annual genetic RS of the treatment groups was similar because (i) T-males suffered fewer losses in genetic RS due to extra-pair fertilizations (EPFs), and (ii) T-males gained more genetic RS through their own EPFs. This is the first hormonal manipulation of an avian phenotype shown to have influenced male RS through EPFs. Together with other studies, it suggests that testosterone may have mediated the evolution of inter- and intraspecific differences in allocation of reproductive effort to mate attraction and parental care.     

Impact of vaginal microbiome communities on HIV antiretroviral-based pre-exposure prophylaxis (PrEP) drug metabolism

Despite the efficacy of antiretroviral-based pre-exposure prophylactics (PrEP) in men who have sex with men, studies in women have produced widely varying outcomes. Recent evidence demonstrates that vaginal microbial communities are associated with increased HIV acquisition risk and may impact PrEP efficacy. Here, we investigate the mechanisms underlying how vaginal bacteria alter PrEP drug levels and impact HIV infection rates ex vivo. Using cervicovaginal lavages (CVLs) from women with or without bacterial vaginosis (BV), we identified microbial metabolism of PrEP drugs in BV samples through LC-MS/MS analysis of soluble drug levels and metabolite formation in dual T-cell cultures. CVL samples were assessed for microbiome analysis using sequencing of bacterial 16S rRNA genes. We also observed non-Lactobacillus bacteria that are associated with BV may potentially impact PrEP efficacy through increased HIV infection rates in co-cultures containing Lactobacillus or BV bacteria, PrEP drugs, CEM-GFP cells, and HIV-1_LAI virus. Finally, we used these data to develop a novel predictive mathematical simulation modeling system to predict these drug interactions for future trials. These studies demonstrate how dysbiotic vaginal microbiota may impact PrEP drugs and provides evidence linking vaginal bacteria to PrEP efficacy in women.