Roselyne Rousseaux-Prévost

Y-autosome translocation and infertility: usefulness of molecular, cytogenetic and meiotic studies

Abstract An apparently balanced reciprocal translocation 46,X,t(Y;6) (q11.23 ∼ q12;p11.1) was observed in an infertile man with severe oligozooteratozoospermia. Different mitotic chromosome banding patterns were performed and fluorescence in situ hybridization indicated a breakpoint in the fluorescent Yq heterochromatin. Molecular genetic deletion experiments for the azoospermia factor region in distal Yq11 showed the retention of the DAZ gene and meiotic pairing configurations suggested that the man’s infertility could be due to the pairing behaviour of the Y;6 translocation chromosome with the X chromosome visualised by synaptonemal complex analysis at the electron microscopy level. The morphological appearance of the normal chromosome 6 and the Y;6 translocated chromosome included in the compartment of the sex vesicle may allow an explanation of the degeneration of most spermatocytes after the pachytene stage.

Actin-Binding Properties and Colocalization with Actin During Spermiogenesis of Mammalian Sperm Calicin

Abstract The nucleus of mammalian spermatozoa is surrounded by a rigid layer, the perinuclear theca, which is divided into a subacrosomal layer and a postacrosomal calyx. Among the proteins characterized in the perinuclear theca, calicin is one of the main components of the calyx. Its sequence contains three kelch repeats and a BTB/POZ domain. We have studied the association of boar calicin with F-actin and the distribution of boar and human calicin during spermiogenesis compared with the distribution of actin. Calicin was purified from boar sperm heads under nondenaturating conditions. The molecule bound actin with high affinity (Kd = ∼5 nM), and a stoichiometry of approximately one calicin per 12 actin monomers was observed. Gel filtration studies showed that calicin forms homomultimers (tetramers and higher polymers). According to immunocytochemical results, calicin is present (together with actin) in the acrosomal region of round spermatids and is mainly localized in the postacrosomal region of late spermatids and spermatozoa. Taken together, the results suggest that the affinity of calicin to F-actin allows targeting of calicin at the subacrosomal space of round spermatids, and that its ability to form homomultimers contributes to the formation of a rigid calyx.

Influence of vitamin A on the immune response of Schistosoma mansoni-infected rats

Nutritional (vitamin A levels, weights), parasitological (adult worm burden, count of eggs in liver, stool examination) and immunological (IgE serum levels, anti-Schistosoma mansoni antibodies, lymphocyte stimulation by concanavalin A and S. mansoni antigenic extract) parameters were studied in three groups of rats, a non-infected and normally fed control group, a S. mansoni-infected but normally fed group, and a S. mansoni-infected group with experimentally induced vitamin A deficiency. The number of worms was found significantly higher in the third (53 +/- 19) than in the second group (2 +/- 2) (p less than 0.001). There were many eggs in the liver surrounded by granulomatous reactions in the third group (399 +/- 73 epg liver). All stool examinations were negative. IgE levels and anti-S. mansoni antibody titres were significantly lower (p less than 0.001) in the third than in the second group. The concanavalin A lymphocyte stimulation indexes did not differ significantly between groups 2 and 3; the S. mansoni lymphocyte stimulation index was only significantly positive in group 3 (p less than 0.001). These results indicate a decrease in the humoral immune response without alteration of cellular immune response in vitamin A-deficient rats infected with S. mansoni.

Proteolysis of Rat Igg Subclasses By Staphylococcus-aureus V8 Proteinase

Monoclonal IgG belonging to the four rat IgG subclasses (IgG1, IgG2a, IgG2b, IgG2c) and some IgG subclasses from normal rat serum were subjected to enzymatic degradation with Staphylococcus aureus V8 proteinase. The results show that only one subclass, IgG2b, is significantly cleaved by the enzyme, with the release of two main products identified as F(ab)2 and Fc-like fragments. This unique susceptibility of the IgG2b subclass represents therefore an easy means of identification and also offers a simple procedure for a preparation of F(ab)2 fragments from monoclonal IgG2b antibodies.

Differential Reduction of the Inter-chain Disulfide Bonds of Rat Immunoglobulin-e - Relation To Biological-activity

Monoclonal rat IgE was reduced over a range of dithiothreitol (DTT) concns. The number of disulfide bonds reduced and their location in the IgE molecule were studied. One millimolar DTT was found to split the two inter-heavy-chain disulfide bonds of the C epsilon 2 domain while increasing DTT concn to 10 mM split the two inter-heavy-light-chain disulfide bridges. Therefore, the sensitivities to reduction of disulfide bonds in rat IgE were found to be the opposite of those in human IgE. In addition, the results indicated the absence, in rat IgE, of the intra-epsilon-chain labile disulfide bond of the C epsilon 1 domain, which is reduced by 2 mM DTT in human IgE. Circular dichroism studies showed significant modifications, mainly of tertiary structure, for rat IgE reduced with 10 mM DTT, but not for IgE reduced with 1 mM DTT. The ability to block passive sensitization with reaginic antibody was not modified when IgE was reduced with 1 mM DTT (which split the two inter-heavy-chain disulfide bonds), but was lost when inter-heavy-light-chain bridges were reduced with 10 mM DTT. In addition, a non-covalent epsilon-chain dimer was found to have the same blocking activity as native IgE (or IgE reduced with 1 mM DTT). Therefore, the results suggest that reduction of most or all the inter-chain disulfide bonds, in rat as in human IgE, induces changes in quaternary structure, more especially in the relationship between the Fab and Fc parts of the molecule, leading to steric blockade, by Fab, of the binding sites for mast cells present on Fc.

Distribution of gelsolin in human testis

Gelsolin, an actin‐binding and severing protein present in many mammalian cells, was characterized in human testis. Although abundant in testicular extracts, gelsolin was not detected in purified spermatogenic cells by immunoblot analysis. Immunofluorescence studies of testis sections showed that gelsolin has two main localizations: peritubular cells and the seminiferous epithelium. In peritubular cells, gelsolin was present together with α‐SM actin, in agreement with the myoid cell characteristics of these cells. In a large proportion of the tubules, gelsolin was found mainly, together with actin, in the apical part of the seminiferous epithelium. This localization of gelsolin also was observed in seminiferous tubules with a partial or complete absence of germinal cells, which evokes a presence of gelsolin at the apex of Sertoli cells. However, in normal testis, a complex pattern of gelsolin labeling was also present, mostly in the apical third of the epithelium, around cells or groups of cells, mainly spermatids, and, less frequently, in various other localizations from the apical to the basal part of the seminiferous epithelium. Taken together, these observations suggest that gelsolin may play different functions in the seminiferous epithelium: (1) regulation of the dynamic alterations of the actin cytoskeleton in the apical cytoplasm of Sertoli cells, and (2) modification of actin filaments assemblies in specific structures at germ cell‐Sertoli cell contacts. Thereby, the actin‐modulating properties of gelsolin are probably involved in reorganization of the seminiferous epithelium related to germ cell differentiation. Mol. Reprod. Dev. 48:63–70, 1997.

Molecular mapping of a Yq deletion in a patient with normal stature

Abstract The proximal long arm of the Y chromosome probably contains a gene (GCY) involved in stature determination. Recent reports have proposed the critical region extends from interval 4B to interval 5G (or 5E). In the present study, the deletion breakpoint in a male adult patient of normal height with a 46,X,del(Yq) karyotype was defined by the use of sequence-tagged site markers. The breakpoint was found between sY78 (interval 4B) and sY79 (interval 5A). The existence of a normal stature in this patient suggests that the growth determinant is proximal to sY79, therefore probably located in interval 4B or in proximal interval 5A of the Y chromosome.

Conformational Studies of Rat Monoclonal Immunoglobulin-e

Two monoclonal IgEs (IR 2 and IR 162) were studied in terms of their conformational features by circular dichroism and differential u.v. absorption. The studies were performed both on the native proteins and under different conditions known to affect the biological properties of IgE (heating at 56 degrees C, acid pH, and presence of reducing agents). Whereas IgE IR 162 appeared to undergo significant conformational changes upon heating and at acid pH, IgE IR 2 was found to be unaffected or less affected by these treatments. The two monoclonal IgEs were found to be equally effective in blocking passive sensitization by IgE antibodies. However, the blocking capacity of IgE IR 162 was affected much more by heating at 56 degrees C than that of IgE IR 2.

Subclass restriction of the antiidiotypic antibody response in the rat

Antiidiotypic antibodies were induced in LOU/M rats by immunization with two myeloma proteins of LOU origin: IR-162 (IgE) and IR-418 (IgG2a). Antibodies to IR-162 were easily obtained after a limited number of immunizations with protein in soluble form; polymerization with glutaraldehyde did not enhance immunogenicity. Antibodies to IR-418 appeared only after a large number of immunizations with protein in polymerized form or with protein copolymerized with rabbit IgG. All of the antibodies, either to IR-162 or to IR-418, were found to be idiotype-specific. In every case for which significant levels of antiidiotypic antibodies were produced, most or all of the antibodies belonged to rat IgG1 subclass. Since, in mice, antiidiotypic antibodies are restricted to the IgG1 subclass, our results indicate a functional analogy between rat and mouse IgG1. Our studies also suggest that the rat IgG1 subclass may be predominantly expressed in T-cell-dependent antibody responses, such as production of antiidiotypic antibodies.