Proteomic analysis of the cellular responses induced in uninfected immune cells by cell-expressed X4 HIV-1 envelope

HIV-1 envelope gp120 and gp41 glycoproteins (Env), expressed at the cell surface, induce uninfected CD4 T-cell death, but the molecular mechanisms leading to this demise are still largely unknown. To better understand these events, we analyzed by a proteomic approach the differential protein expression profile of two types of uninfected immune cells after their coculture for 1-3 days with cells that express, or not, Env. First, umbilical cord blood mononuclear cells (UCBMCs) were used to approach the in vivo situation, i.e., blood uninfected naive cells that encounter infected cells. Second, we used the A2.01/CD4.403 T-cell line expressing wild type CXCR4 and a truncated form of CD4 that still undergoes Env-mediated apoptosis, independently of CD4 signaling. After coculture with cells expressing Env, 35 and 39 proteins presenting an altered expression in UCBMCs and the A2.01/CD4.403 T-cell line, respectively, were identified by mass-spectrometry. Whatever the cell type analyzed, the majority of these proteins are involved in degradation processes, redox homeostasis, metabolism and cytoskeleton dynamics, and linked to mitochondrial functions. This study provides new insights into the events that sequentially occur in bystander T lymphocytes after contact with HIV-infected cells and leading, finally, to apoptotic cell death.     

Isolation of four HSP70 genes in the pig and localization on Chromosomes 7 and 14

For insight into the general organization of the swine leukocyte antigen (SLA) complex, the swine major histocompatibility complex (MHC), four sequences related to the heat-shock proteins HSP70 were characterized by screening of a pig genomic cosmid library with a swine cDNA HSP70 2.6-kb probe. This yielded three positive clones: HC2.2, HC3.2, and HC4.2. Restriction site maps revealed a large overlap of HC2.2 with HC3.2, whereas HC4.2 was independent. Southern blot hybridization with the 5 section, the central section, and the 3 section of the 2.6-kb probe and also with a swine 4.5-kb HSP70 genomic probe suggested the existence, within the overlapping clones, of three distinct HSP70 sequences encompassing a segment no longer than 22 kb. The HC4.2 clone, which hybridized with the same probes, displayed a single band of 7.3 kb, probably corresponding to one gene only. Fluorescent in situ hybridization on swine chromosome metaphases with the whole HC2.2 or HC4.2 cosmids allowed the assignment of HC2.2 to MHC region on Chromosome (Chr) 7 (Cen-p1.1), and of HC4.2 to Chr 14 (q2.4–2.5). Thus, as in humans, the swine MHC comprises three closely linked HSP70 loci. The presence of additional genes belonging to the same inducible HSP70 gene family can be expected from what is known in humans. The HSP70 gene found here on the pig Chr 14 may be one of these putative unidentified genes.     

Using the International Classification of Functioning,Disability and Health (ICF) to Describe Children Referred to Special Care or Paediatric Dental Services

Children in dentistry are traditionally described in terms of medical diagnosis and prevalence of oral disease. This approach gives little information regarding a child’s capacity to maintain oral health or regarding the social determinants of oral health. The biopsychosocial approach, embodied in the International Classification of Functioning, Disability and Health - Child and Youth version (ICF-CY) (WHO), provides a wider picture of a child’s real-life experience, but practical tools for the application of this model are lacking. This article describes the preliminary empirical study necessary for development of such a tool - an ICF-CY Core Set for Oral Health. An ICF-CY questionnaire was used to identify the medical, functional, social and environmental context of 218 children and adolescents referred to special care or paediatric dental services in France, Sweden, Argentina and Ireland (mean age 8 years ±3.6yrs). International Classification of Disease (ICD-10) diagnoses included disorders of the nervous system (26.1%), Down syndrome (22.0%), mental retardation (17.0%), autistic disorders (16.1%), and dental anxiety alone (11.0%). The most frequently impaired items in the ICF Body functions domain were ‘Intellectual functions’, ‘High-level cognitive functions’, and ‘Attention functions’. In the Activities and Participation domain, participation restriction was frequently reported for 25 items including ‘Handling stress’, ‘Caring for body parts’, ‘Looking after one’s health’ and ‘Speaking’. In the Environment domain, facilitating items included ‘Support of friends’, ‘Attitude of friends’ and ‘Support of immediate family’. One item was reported as an environmental barrier – ‘Societal attitudes’. The ICF-CY can be used to highlight common profiles of functioning, activities, participation and environment shared by children in relation to oral health, despite widely differing medical, social and geographical contexts. The results of this empirical study might be used to develop an ICF-CY Core Set for Oral Health - a holistic but practical tool for clinical and epidemiological use.     

Temporal and sexual variation of leaf-produced pollinator-attracting odours in the dwarf palm

Information on intra-specific variation in pollinator-attracting floral traits provides clues to selective pressures imposed by pollinators. However, these traits also reflect constraints related to floral phenology or morphology. The specific weevil pollinator Derelomus chamaeropsis of the dioecious Mediterranean dwarf palm Chamaerops humilis is attracted by volatile compounds that leaves, and not flowers, release during anthesis. Production of these olfactory cues is thus probably not constrained by any other floral function. This provides the opportunity to study variation of a floral trait that is not produced by a floral organ. We studied volatile compounds emitted by leaves of 12 individual C. humilis over the whole flowering season. The quantity of volatile compounds emitted by leaves reached a maximum when plants required pollinator visits. The relationship between odour emission and floral phenology was slightly different between male and female plants, probably reflecting differences in the exact time at which females and males benefit from pollinator visitation. Male plants produced higher quantities of volatile compounds than females. Odour composition was highly variable among individuals but did not differ between male and female plants. In this system, female C. humilis are pollinated by deceit and pollinators should be selected to avoid visiting them. The absence of sexual difference in blend composition may thus prevent pollinators from discriminating between male and female plants.An erratum to this article can be found at     

Characterization of a Highly Replicative Intergroup M/O Human Immunodeficiency Virus Type 1 Recombinant Isolated from a Cameroonian Patient

A Cameroonian patient with antibodies reacting simultaneously to human immunodeficiency virus type 1 (HIV-1) group O- and group M-specific V3-loop peptides was identified. In order to confirm that this patient was coinfected with both viruses, PCRs with O- and M-specific discriminating primers corresponding to different regions of the genome were carried out with both primary lymphocyte DNA and the corresponding viral strains isolated from three consecutive patient samples. The PCR data suggested that this patient is coinfected with a group M virus and a recombinant M/O virus. Indeed, only type M gag sequences could be amplified, while for the env region, both type M and O sequences were amplified, from plasma or from DNA extracted from primary lymphocytes. Sequence analysis of a complete recombinant genome isolated from the second sample (97CA-MP645 virus isolate) revealed two intergroup breakpoints, one in the vpr gene and the second in the long terminal repeat region around the TATA box. Comparison of the type M sequences shared by the group M and the recombinant M/O viruses showed that these sequences were closely related, with only 3% genetic distance, suggesting that the M virus was one of the parental viruses. In this report we describe for the first time a recombination event in vivo between viruses belonging to two different groups, leading to a replicative virus. Recombination between strains with such distant lineages (65% overall homology) may contribute substantially to the emergence of new HIV-1 variants. We documented that this virus replicates well and became predominant in vitro. At this time, group O viruses represent a minority of the strains responsible for the HIV-1 pandemic. If such recombinant intergroup viruses gained better fitness, inducing changes in their biological properties compared to the parental group O virus, the prevalences of group O sequences could increase rapidly. This will have important implications for diagnosis of HIV-1 infections by serological and molecular tests, as well as for antiviral treatment.     

The influence of correlated protein-water volume fluctuations on the apparent compressibility of proteins determined by ultrasonic velocimetry

The elasticity of proteins, expressed by the compressibility, is potentially one of the most important properties of proteins because of the close relationship with its functionality. The compressibility of solutions can be determined by measurements of sound velocity and density. These quantities are related by the Newton-Laplace equation. In order to interpret the apparent compressibility of solutes in highly dilute solutions, it is required to consider the relation between compressibility and sound velocity of the solution using an appropriate reference system. The classical approach usually gives too small values for the apparent compressibility when compared with other methods. We show that the difference can partially be explained if the correlated volume fluctuations of the solvent are taken into consideration. A special attention is given to the compressibility of proteins. Finally, the present paper is not intended to replace established approaches, but it wants to create awareness that the classical mixing rules refer to ideal gasses assuming uncorrelated volume fluctuations and that a considerable part of the hydration effects could be explained by correlated volume fluctuations.     

Cell autonomy of the mouse claw paw mutation

Mice homozygous for the autosomal recessive mutation claw paw (clp) are characterized by limb posture abnormalities and congenital hypomyelination, with delayed onset of myelination of the peripheral nervous system but not the central nervous system. Although this combination of limb and peripheral nerve abnormalities in clp/clp mice might suggest a common neurogenic origin of the syndrome, it is not clear whether the clp gene acts primarily in the neurone, the Schwann cell or both. In the work described here, we address this question of cell autonomy of the clp mutation through reciprocal nerve grafting experiments between wild-type and clp/clp animals. Our results demonstrate that the clp mutation affects the Schwann cell compartment and possibly also the neuronal compartment. These data suggest that the clp gene product is expressed in Schwann cells as well as neurones and is likely to be involved in direct axon--Schwann cell interactions. Within the Schwann cell, clp affects a myelin-related signaling pathway that regulates periaxin and Krox-20 expression, but not Oct-6.     

Fitness of Toxoplasma gondii is not related to DHFR single-nucleotide polymorphism during congenital toxoplasmosis

Factors that regulate the pathogenesis of Toxoplasma gondii in humans are poorly understood. When acquired during pregnancy, toxoplasmosis can be disastrous, leading to fetal loss or conversely to subclinical disease. In congenitally infected infants, evolution is highly unpredictable. Genotype based virulence patterns have been described in mice, but in humans this classification does not correlate with the gravity of the disease. Mutations on DHFR-TS loci have recently been reported to confer T. gondii fitness cost. In this study, we investigated the relationship between the virulence of the parasite, as measured by clinical outcome in the fetus or newborn, fitness, as measured by parasitic load in amniotic fluid, and allelic polymorphism in DHFR. Six cases of severe congenital toxoplasmosis and 23 cases of mild congenital infections were included in the study. Quantitative PCR was performed to evaluate total T. gondii DNA load in amniotic fluid and detection of mutations was carried out with a LightCycler using hybridisation probes. Parasitic load was significantly higher in severe infections than in mild diseases. Among isolates from severe or non-severe cases of congenital toxoplasmosis, no polymorphism could be detected at loci 36, 83 or 245 of the DHFR gene. The virulent RH strain presented the same melting temperature as the non-virulent PRU strain for codons 36, 83 and 245. Only mutated clones, M2M3 and M2M4 with allelic replacement at these positions, displayed different profiles allowing a clear distinction between wild and mutant types. We concluded that the DHFR gene mutations we investigated do not regulate T. gondii fitness in humans.