Marlène Bouillon

CD40 Ligand Binds to α5β1 Integrin and Triggers Cell Signaling

It was originally thought that the critical role of the CD40 ligand (CD40L) in normal and inflammatory immune responses was mainly mediated through its interaction with the classic receptor, CD40. However, data from CD40L–/– and CD40–/– mice suggest that the CD40L-induced inflammatory immune response involves at least one other receptor. This hypothesis is supported by the fact that CD40L stabilizes arterial thrombi through an αIIbβ3-dependent mechanism. Here we provide evidence that soluble CD40L (sCD40L) binds to cells of the undifferentiated human monocytic U937 cell line in a CD40- and αIIbβ3-independent manner. Binding of sCD40L to U937 cells was inhibited by anti-CD40L monoclonal antibody 5C8, anti-α5β1 monoclonal antibody P1D6, and soluble α5β1. The direct binding of sCD40L to purified α5β1 was confirmed in a solid phase binding assay. Binding of sCD40L to α5β1 was modulated by the form of α5β1 expressed on the cell surface as the activation of α5β1 by Mn2+ or dithiothreitol resulted in the loss of sCD40L binding. Moreover, sCD40L induced the translocation of α5β1 to the Triton X-100-insoluble fraction of U937 cells, the rapid activation of the MAPK pathways ERK1/2, and interleukin-8 gene expression. The binding of sCD40L to CD40 on BJAB cells, an α5β1-negative B cell line, and the resulting activation of ERK1/2 was not inhibited by soluble α5β1, suggesting that sCD40L can bind concomitantly to both receptors. These results document the existence of novel CD40L-dependent pathways of physiological relevance for cells expressing multiple receptors (CD40, α5β1, and αIIbβ3) for CD40L.

The two isoforms of the cAMP-dependent protein kinase catalytic subunit are involved in the control of dimorphism in the human fungal pathogen Candida albicans

We have cloned the Candida albicans TPK2 gene encoding a cAMP-dependent protein kinase (PKA) catalytic subunit and generated a tpk2 homozygous null mutant to assess its ability to germinate in liquid media. N-acetylglucosamine (GlcNAc)-induced germ-tube formation was attenuated in the tpk2 strain and enhanced by compounds that are known to increase the PKA activity in situ. Germination was completely blocked in the presence of the myristoylated derivative of the heat-stable PKA inhibitor (MyrPKI). These results indicate that TPK1 acts positively in regulating the morphogenetic transition in C. albicans in the absence of the TPK2 gene. We were able to identify an mRNA from this second form of PKA in both wild-type and tpk2 null mutant cells. We found that PKA activity measured in the mutant lacking the TPK2 gene was about 10% of that displayed by the wild-type. The finding that the germinative response of tpk2 null mutant to serum was severely diminished at low serum concentrations indicates that the level of PKA is an important determinant of filamentous growth at low serum concentrations. The extent of germination attained at higher serum concentrations (5%) was similar in the wild-type and in the tpk2 null mutant strains suggesting that under these conditions germination was triggered through a PKA-independent pathway.

Regulation by retinoic acid of ICAM-1 expression on human tumor cell lines

In a group of four human tumor cell lines comprising one melanoma, one glioma, one teratocarcinoma and one neuroblastoma, the expression of the intercellular adhesion molecule-1 (ICAM-1) was found to be significantly increased following treatment with 10 microM of all-trans retinoic acid. In the melanoma and glioma cell lines HS 294T and HS 683, greater than 90% of the cells reacted with the anti-ICAM-1 monoclonal antibody (mAb) CL203.4 in the absence of treatment. Retinoic acid increased the cell surface expression of the molecule by 2-fold. In the teratocarcinoma and neuroblastoma cell lines, TERA-2 and SK-N-SH, the constitutive expression of ICAM-1 was weak, the percentage of cells stained above the background being less than 25%. Retinoic acid induced ICAM-1 expression in greater than 80% of the cells and increased the levels of expression by 2.5 to 3-fold. Immunoprecipitation studies in biosynthetically labeled cells as well as RNase protection analysis confirmed that retinoic acid treatment increased the amount of ICAM-1 at both the protein and mRNA level. The induction or stimulation occurred within 24 h, was maximal after 4 days and reversible.

Reduced frequency of blood donors with false‐positive HIV‐1 and ‐2 antibody EIA reactivity after elution of low‐affinity nonspecific natural antibodies

BACKGROUND : The detection by EIA of antibodies (Abs) specific to HIV antigens in the serum of blood donors is important for transfusion safety. A small but significant number of donor sera (0.1‐0.3%) yield false‐positive results in EIA, and these donors must be permanently deferred from the blood donor list, causing operational and public relations problems.