Post-transcriptional regulation controlled by E-cadherin is important for c-Jun activity in melanoma.

A central event in the development of malignant melanoma is the loss of the tumor-suppressor protein E-cadherin. Here, we report that this loss is linked to the activation of the proto-oncogene c-Jun, a key player in tumorigenesis. In vivo, malignant melanomas show strong expression of the c-Jun protein in contrast to melanocytes. Interestingly, c-Jun mRNA levels did not differ in the melanoma cell lines when compared to melanocytes, suggesting that c-Jun could be regulated at the post-transcriptional level. To uncover the link between E-cadherin and c-Jun, we re-expressed E-cadherin in melanoma cells and detected decreased protein expression and activity of c-Jun. Furthermore, c-Jun accumulation is dependent on active E-cadherin-mediated cell-cell adhesion and regulated via the cytoskeleton. Additionally, we determined that, with respect to c-Jun regulation, there are two melanoma subgroups. One subgroup regulates c-Jun expression via the newly discovered E-cadherin-dependent signaling pathway, whereas the other subgroup uses the MAPKinases to regulate its expression. In summary, our data provide novel insights into the tumor-suppressor function of E-cadherin, which contributes to the suppression of c-Jun protein translation and transcriptional activity independent of MAPKinases.

Hydrophobic binding of pertussis toxin is enhanced by oligosaccharide receptors.

Pertussis toxin is one of several virulence factors produced by Bordetella pertussis, the etiologic agent of whooping cough. Pertussis toxin is an oligomeric A-B class toxin composed of an ADP-ribosyltransferase S1 (A) subunit and a B oligomer containing lectin-like binding domains. The carbohydrate binding specificity of the B oligomer is for sialooligosaccharide sequences expressed on target cell receptors and asparagine-linked glycans found in many serum glycoproteins. Pertussis toxin also has the ability to bind to the inert surfaces of culture tubes. In this report we present data showing that pertussis toxin binding to polypropylene microcentrifuge tubes was enhanced in a time- and concentration-dependent manner by the addition of soluble glycoprotein or oligosaccharide receptor analogs. Evidence obtained using the hydrophilic and hydrophobic surfaces of Gel Bond electrophoresis casting film indicated that receptor-enhanced binding was likely due to hydrophobic interactions. Hydrophobic binding of the isolated B oligomer of pertussis toxin was enhanced only in the presence of high concentrations of glycoproteins. Therefore, the S1 (A) subunit of pertussis holotoxin appears to play a role in receptor-enhanced hydrophobic binding. We propose, therefore, that pertussis toxin binding to its receptors may expose or preferentially orient hydrophobic residues that may contribute to the functional association of the toxin with host cell plasma membranes and delivery of the S1 subunit to its intracellular target.

Binding to native proteins by antipeptide monoclonal antibodies.

mAb raised against synthetic peptides derived from cholera toxin, myohemerythrin, and sickle hemoglobin were analyzed by both solid-phase and solution-phase methods. Antipeptide mAb against cholera toxin (mAb TE32 and TE33), against myohemerythrin (mAb B13I2, B13C2, and B13F2), and against sickle hemoglobin (mAb HuS-1 and HuS-2), had been previously described and used for vaccine development, structural characterization, or identification of a specific antigenic determinant, and each was apparently capable of binding both peptide and native Ag. In this study, all were found to bind whole protein when tested against immobilized Ag in a standard solid-phase assay (ELISA), yet none of the antibodies recognized the Ag in its true native form, failing to bind when tested in several solution-phase assay systems, including size exclusion HPLC. This discrepancy may be the result of modifications of the epitope created by interaction and possible denaturation of the protein on the solid-phase matrix. As a consequence, binding of these antibodies to peptides, either immobilized or in solution, or to immobilized protein, cannot be used to infer that the peptide has assumed a conformation that corresponds to that of the cognate sequence in the native protein. A re-evaluation of binding data that relates antipeptide mAb to native structural characteristics may be necessary.