Identification of two cross-neutralizing linear epitopes within the L1 major capsid protein of human papillomaviruses

The neutralizing activities of polyclonal antibodies and monoclonal antibodies (MAbs) obtained by immunization of mice with L1 virus-like particles (VLPs) were investigated by using pseudovirion infectivity assays for human papillomavirus type 16 (HPV-16), HPV-31, HPV-33, HPV-45, HPV-58, and HPV-59 to obtain a better definition of cross-neutralization between high-risk HPVs. In this study, we confirmed and extended previous studies indicating that most genital HPV genotypes represent separate serotypes, and the results suggest that the classification of serotypes is similar to that of genotypes. In addition, three cross-neutralizing MAbs were identified (HPV-16.J4, HPV-16.I23, and HPV-33.E12). MAb HPV-16.J4 recognized a conserved linear epitope located within the FG loop of the L1 protein, and HPV-16.I23 recognized another located within the DE loop. The results suggested that reactivity of MAb HPV-16.I23 to L1 protein is lost when leucine 152 of the HPV-16 L1 protein is replaced by phenylalanine. This confirmed the existence of linear epitopes within the L1 protein that induce neutralizing antibodies, and this is the first evidence that such linear epitopes induce cross-neutralization. However, the cross-neutralization induced by L1 VLPs represents less than 1% of the neutralizing activity induced by the dominant conformational epitopes, and it is questionable whether this is sufficient to offer cross-protection in vivo.     

Genetic diversity within human erythroviruses: identification of three genotypes

B19 virus is a human virus belonging to the genus Erythrovirus: The genetic diversity among B19 virus isolates has been reported to be very low, with less than 2% nucleotide divergence in the whole genome sequence. We have previously reported the isolation of a human erythrovirus isolate, termed V9, whose sequence was markedly distinct (>11% nucleotide divergence) from that of B19 virus. To date, the V9 isolate remains the unique representative of a new variant in the genus Erythrovirus, and its taxonomic position is unclear. We report here the isolation of 11 V9-related viruses. A prospective study conducted in France between 1999 and 2001 indicates that V9-related viruses actually circulate at a significant frequency (11.4%) along with B19 viruses. Analysis of the nearly full-length genome sequence of one V9-related isolate (D91.1) indicates that the D91.1 sequence clusters together with but is notably distant from the V9 sequence (5.3% divergence) and is distantly related to B19 virus sequences (13.8 to 14.2% divergence). Additional phylogenetic analysis of partial sequences from the V9-related isolates combined with erythrovirus sequences available in GenBank indicates that the erythrovirus group is more diverse than thought previously and can be divided into three well-individualized genotypes, with B19 viruses corresponding to genotype 1 and V9-related viruses being distributed into genotypes 2 and 3.     

A germline-specific gap junction protein required for survival of differentiating early germ cells

Germ cells require intimate associations and signals from the surrounding somatic cells throughout gametogenesis. The zero population growth (zpg) locus of Drosophila encodes a germline-specific gap junction protein, Innexin 4, that is required for survival of differentiating early germ cells during gametogenesis in both sexes. Animals with a null mutation in zpg are viable but sterile and have tiny gonads. Adult zpg-null gonads contain small numbers of early germ cells, resembling stem cells or early spermatogonia or oogonia, but lack later stages of germ cell differentiation. In the male, Zpg protein localizes to the surface of spermatogonia, primarily on the sides adjacent to the somatic cyst cells. In the female, Zpg protein localizes to germ cell surfaces, both those adjacent to surrounding somatic cells and those adjacent to other germ cells. We propose that Zpg-containing gap junctional hemichannels in the germ cell plasma membrane may connect with hemichannels made of other innexin isoforms on adjacent somatic cells. Gap junctional intercellular communication via these channels may mediate passage of crucial small molecules or signals between germline and somatic support cells required for survival and differentiation of early germ cells in both sexes.     

Genome-wide analysis of mRNAs targeted to yeast mitochondria

It is agreed that nuclear-encoded mitochondrial proteins are post-translationally targeted to mitochondria, even if, in some cases, a co-translational phase can assist the import of precursor proteins. We used yeast DNA microarrays to analyse the mRNA populations associated with free and mitochondrion-bound polysomes. As expected, many mRNAs, known to encode mitochondrial proteins, are localized to free cytoplasmic polysomes, but many are localized to mitochondrion-bound polysomes. Furthermore, the 3′-UTR of six randomly chosen mitochondrion-bound mRNAs contains sufficient information to target, in vivo, non-translatable RNA to the vicinity of mitochondria. Interestingly, genes producing mRNAs that are targeted to mitochondria are mainly of ancient bacterial origin, whereas those producing mRNAs that are translated in the cytoplasm are mainly of eukaryotic origin. These observations, which support the recent hypotheses concerning the dual origin of the mitochondrial proteome, provide new insights into the biogenesis of mitochondria.     

DNA immunisation. New histochemical and morphometric data

Splenic germinal center reactions were measured during primary response to a plasmidic DNA intramuscular injection. Cardiotoxin-pretreated Balb/c mice were immunized with DNA plasmids encodmg or not the SAG1 protein, a membrane antigen of Toxoplasma gondii. Specific anti-SAG1 antibodies were detected on days 16 and 36 after injection of coding plasmids. The results of ELISAs showed that the SAG1-specific antibodies are of the IgG2a class. Morphometric analyses were done on serial immunostained cryosections of spleen and draining or non-draining lymph nodes. This new approach made it possible to evaluate the chronological changes induced by DNA immunisation in the germinal centres (in number and in size). Significant increases in the number of germinal centres were measured in the spleen and only in draining lymph nodes after plasmid injection, the measured changes of the germinal centers appeared to result from the adjuvant stimulatory effect of the plasmidic DNA since both the coding and the noncoding plasmid DNA induced them. No measurable changes were recorded in the T-dependent zone of lymph organs.     

2-alkyl-3-Alkylamino-2H-Benzo- and pyridothiadiazine 1,1-dioxides: from K+ATP channel openers to Ca++ channel blockers?

A series of 2-alkyl-3-alkylamino-2H-benzo- and 2-alkyl-3-alkylamino-2H-pyrido[4,3-e]-1,2,4-thiadiazine 1,1-dioxides, structurally related to BPDZ 44 and BPDZ 73, two potent pancreatic B-cells K+ATP channel openers, were synthesized and tested on rat pancreatic islets (endocrine tissue) as well as on rat aorta rings (vascular smooth muscle tissue). Alkylation of the 2-position led to double bond tautomerization and formation of compounds with a 2H-conformation. In contrast to the previously described pyridothiadiazine dioxides, such as BPDZ 44, and 7-chlorobenzothiadiazine dioxides, such as BPDZ 73, the 2-alkyl-substituted analogs were found to be poorly active on the insulin releasing process although most drugs exhibited a vasorelaxant activity. As a result, the new 2-alkyl-substituted pyridinic compounds expressed a selectivity profile (vascular smooth muscle tissue vs pancreatic tissue) opposite to that of their non-alkyl-substituted counterparts, i.e. BPDZ 44. Additional investigations revealed that, in contrast to their non 2-alkyl-substituted analogs, the most interesting 2-methyl-substituted derivatives did not express the pharmacological profile of classical K+ATP channel openers. The pharmacological results rather suggest that alkylation of the 2-position of the thiadiazine ring led to drugs that could act as Ca2+ channel blockers rather than as potassium channel openers.     

Peptide fragments released from Phe-caseinomacropeptide in vivo in the rat

The aim of this study was to investigate the pharmacokinetics of bovine Phe-caseinomacropeptide (Phe-CMP) in the rat after oral administration. This polypeptide was monophosphorylated and mainly nonglycosylated: Phe-CMP-1P. During gastrointestinal digestion and absorption, Phe-CMP-1P was degraded. Intact Phe-CMP-1P and CMP-1P were rapidly released from the stomach. In contrast, partial hydrolysis by pancreatic enzymes was observed. In vitro hydrolysis by brush-border membrane vesicles also indicated that the peptide was degraded. In the blood, "CMP-immunoreactive material" appeared rapidly, reaching a maximum level of 5.5 microg/ml at 60 min.