Dominique Treton

IL4 and IL13 receptors share the γc chain and activate STAT6, STAT3 and STAT5 proteins in normal human B cells

IL13 induces the same biological effects as IL4 in normal human B cells. We show that as in the IL4R complex, both IL4Rα and IL2Rγc are components of the IL13R and that both cytokines induced STAT6, STAT3 and STAT5 activation in B cells. In spite of this similar downstream signalling, IL4 and IL13 used a different set of Janus kinases: IL13 is unable to activate JAK1 and JAK3.

Expression of Interleukin 13 Receptor in Glioma and Renal Cell Carcinoma: IL13Rα2 as a Decoy Receptor for IL13

Glioma and renal cell carcinoma (RCC) cells express high affinity interleukin 13 (IL13) binding sites, but only RCC cell proliferation was inhibited by IL13. Both of these two cell types are IL2-receptor γc chain-negative. We thus used these cell models to investigate the patterns of expression of IL13Rα1, IL13Rα2, and IL4Rα chains and the role of IL13Rα2 in the response to IL13. Using new specific antibodies and flow cytometry, we observed a similar surface expression of IL4Rα and IL13Rα1 chains in most RCC and glioma cells, whereas IL13Rα2 was only present on five of six glioma cell lines. In all glioma cell lines, the amount of IL13Rα2 expression was 10 to 30 times higher than that of the two other chains. Although there was no surface or intracellular expression of IL13Rα2, its mRNA was detected in three of seven RCC cell lines. The expression on RCC cells of IL13Rα2 mRNA and/or that of high-affinity IL13 binding sites is not sufficient to predict IL13Rα2 protein expression. Blocking experiments showed that IL4 and IL13 strongly inhibited RCC cell proliferation through a unique receptor composed of IL4Rα and IL13Rα1 chains. Using RCC cells stably transfected with IL13Rα2 cDNA, we showed that the overexpression of IL13Rα2 decreased the response to IL13 but not that to IL4. Our results demonstrate that IL13Rα2 acts as a decoy receptor for IL13 and that it may exert a tight regulation of IL13 activity without impairing the IL4 response of the same cell target.

HIV Type 1 Glycoprotein 120 Inhibits Human B Cell Chemotaxis to CXC Chemokine Ligand (CXCL) 12, CC Chemokine Ligand (CCL)20, and CCL21

We analyzed the modulation of human B cell chemotaxis by the gp120 proteins of various HIV-1 strains. X4 and X4/R5 gp120 inhibited B cell chemotaxis toward CXCL12, CCL20, and CCL21 by 40–50%, whereas R5 gp120 decreased inhibition by 20%. This gp120-induced inhibition was strictly dependent on CXCR4 or CCR5 and lipid rafts but not on CD4 or VH3-expressing BCR. Inhibition did not impair the expression or ligand-induced internalization of CCR6 and CCR7. Our data suggest that gp120/CXCR4 and gp120/CCR5 interactions lead to the cross-desensitization of CCR6 and CCR7 because gp120 does not bind CCR6 and CCR7. Unlike CXCL12, gp120 did not induce the activation of phospholipase Cβ3 and PI3K downstream from CXCR4, whereas p38 MAPK activation was observed. Similar results were obtained if gp120-treated cells were triggered by CCL21 and CCL20. Our results are consistent with a blockade restricted to signaling pathways using phosphatidylinositol-4,5-bisphosphate as a substrate. X4 and X4/R5 gp120 induced the cleavage of CD62 ligand by a mechanism dependent on matrix metalloproteinase 1 and 3, CD4, CXCR4, Gαi, and p38 MAPK, whereas R5 gp120 did not. X4 and X4/R5 gp120 also induced the relocalization of cytoplasmic CD95 to the membrane and a 23% increase in CD95-mediated apoptosis. No such effects were observed with R5 gp120. The gp120-induced decrease in B cell chemotaxis and CD62 ligand expression, and increase in CD95-mediated B cell apoptosis probably have major deleterious effects on B cell responsiveness during HIV infection and in vaccination trials.

Lead Reduces Messenger RNA and Protein Levels of Cytochrome P450 Aromatase and Estrogen Receptor β in Human Ovarian Granulosa Cells

Abstract Exposure to lead causes decreased fertility in women. In the present study, we examined the in vitro effects of lead on cytochrome P450 aromatase (P450 arom) and on estrogen receptor β (ERβ), two key proteins for the human ovary. Aromatase is required for the bioconversion of androgen to estradiol; ERβ mediates estrogen effects in granulosa cells. Granulosa cells were collected from women undergoing in vitro fertilization and then cultured with 10 µM lead acetate. Using atomic absorption spectrometry, we showed that lead accumulated in cells. Aromatase activity as measured by a tritiated water production assay was significantly reduced. Using semiquantitative reverse transcription-polymerase chain reaction and Western blotting procedures, we showed that P450 arom and ERβ mRNA and protein content were both significantly reduced. Adding 10 µg/ml of cycloheximide, a protein inhibitor, did not eliminate the effects of lead. The present results support the hypothesis that the action of lead on fertility in women may result, in part, from the down-regulation of P450 arom and ERβ gene transcription in ovarian granulosa.

Natural Mutations of the anti-Müllerian Hormone type II Receptor Found in Persistent Müllerian Duct Syndrome affect Ligand Binding, Signal Transduction and Cellular Transport

The anti-Müllerian hormone type II (AMHRII) receptor is the primary receptor for anti-Müllerian hormone (AMH), a protein produced by Sertoli cells and responsible for the regression of the Müllerian duct in males. AMHRII is a membrane protein containing an N-terminal extracellular domain (ECD) that binds AMH, a transmembrane domain, and an intracellular domain with serine/threonine kinase activity. Mutations in the AMHRII gene lead to persistent Müllerian duct syndrome in human males. In this paper, we have investigated the effects of 10 AMHRII mutations, namely 4 mutations in the ECD and 6 in the intracellular domain. Molecular models of the extra- and intracellular domains are presented and provide insight into how the structure and function of eight of the mutant receptors, which are still expressed at the cell surface, are affected by their mutations. Interestingly, two soluble receptors truncated upstream of the transmembrane domain are not secreted, unless the transforming growth factor beta type II receptor signal sequence is substituted for the endogenous one. This shows that the AMHRII signal sequence is defective and suggests that AMHRII uses its transmembrane domain instead of its signal sequence to translocate to the endoplasmic reticulum, a characteristic of type III membrane proteins.

Immunoregulatory functions of paf-acether. VI. Dual effect on human B cell proliferation

The role of paf-acether (paf), a phospholipid cytokine, in the modulation of human B cell function was investigated. Paf, from 1×10−5 M to 10−6 M, decreased B cell proliferation induced by both phorbol myristate acetate (PMA) and anti-IgM antibodies (anti-IgM Ab). By contrast, 1×10−7 M to 1×10−9 M paf enhanced PMA triggered, but not anti-IgM triggered B cell proliferation. B cell proliferation was modulated between 24 and 72 hr of culture indicating that the effect of paf did not merely reflect a shift in proliferation kinetics. Interestingly, paf also enhanced the spontaneous proliferation of a Burkitt lymphoma-derived B cell line, Raji, which suggests that paf can directly act on B cells. The modulatory effect of paf on peripheral blood B cells was independent of PMA concentration, yet the effect on Raji cells was dependent upon cell density. The data suggest that paf is a potent modulator of B cell function, and may be involved in the control of humoral immune response.

SOB3, a human sperm protein involved in zona pellucida binding: Physiological and biochemical analysis, purification

LB5 antibody was selected from a monoclonal antibody (mAb) library directed against human sperm proteins. LB5 mAb detected the corresponding protein SOB3 in the neck region and the flagellum of most live ejaculated sperm while it labelled, in addition, the acrosome of about 10–20% of spermatozoa. The percentage of LB5 acrosome‐stained sperm was significantly correlated with the percentages of either spontaneous or A23187‐induced acrosome‐reacted sperm. While SOB3 could not be detected in the testis, it appeared in spermatozoa from the corpus epididymis segment. LB5 mAb impaired neither sperm motion parameters, acrosomal reaction triggering, nor sperm binding to zona‐free hamster oocytes. By contrast, LB5 Fab fragments (200 μg/ml) inhibited sperm binding to human zonae pellicidae by 35.7%. If sperm were induced to acrosome react with A23187 prior to LB5 treatment, the inhibitory effect shifted to 59.9%, while no significant effect was observed following A23187 incubation alone. Western blotting of human sperm and cauda epididymis extracts revealed two bands of 18 and 19 kDa. While no cross‐reaction was observed with other tested organs, a similar 18‐kDa band was revealed in erythocytes and one of 19 kDa in B‐lymphocytes. No cross‐reactivity could be evidenced in any animal sperm analyzed. SOB3 was first separated in a 17‐ to 20‐kDa preparative electrophoresis fraction and finally purified by isoelectrofocusing according to its pI of 9.8. These results suggest that SOB3 is localized under the outer acrosomal membrane, that it participates in secondary sperm binding to the zona pellucida, and that it shares homologies with the immune system. Mol. Reprod. Dev. 49:286–297, 1998.

Regulation of platelet-activating factor receptor expression in human B cells and B cell lines

We extended our previous data regarding the modulation of human platelet-activating factor receptor (hPAF-R) expression on human B cell lines as well as normal B cells. First, we showed that hPAF-R, but was absent from cell lines devoid of hPAF-R. Second, enhanced hPAF-R membrane expression induced in IM9 line by IL9 line by IL4 was preceeded by hPAF-R mRNA accumulation that was detectable by 8 h and which peaked at 24 h. Similar results were observed for 10 nM platelet-activating factor treatment, which increased hPAF-R mRNA content up to 120% at 48 h, whereas hPAF-R membrane expression was up-regulated by 130%. Third, our data indicate that functional hPAF-R are expressed on resting, as well as on activated, B cells and that B cell activation is required for maintaining hPAF-R membrane and mRNA expression. Thus, in normal B cells, as well as in B cell lines, transcriptional regulation and/or messager stability control hPAF-R expression.