The effects of copper on actin and fibronectin organization in Mytilus galloprovincialis haemocytes

The effects of copper on actin and fibronectin organization in Mytilus galloprovincialis haemocytes were studied. The Cu2+ exposure of mussels caused severe perturbations in haemocyte actin and fibronectin organization with respect to non-exposed organisms. Cytoskeletal actin was analysed by indirect immunofluorescence, using an antitotal actin monoclonal antibody, and by rhodamine-conjugated phalloidin. The majority of haemocytes from Cu(2+)-exposed mussels displayed a round morphology, with short and blunt filopodia; they lacked the polarized phenotype which was typical in control samples. The cytoskeleton alteration, more evident after phalloidin staining, resulted in the disappearance of filamentous actin. The actin cortical meshwork also appeared disorganized. The cytoskeletal morphology studied by transmission electron microscopy after negative staining of Triton X-100-treated haemocytes confirmed these observations. The structural organization of actin when analysed by Western blotting showed a larger number of Triton-soluble actin pools in treated mussel haemocytes. Fibronectin was studied by indirect immunofluorescence using a polyclonal antiserum directed against mussel fibronectin. In treated mussels, fibronectin appeared to be strongly disorganized and its levels decreased in both haemocytes and haemolymph. The mechanism(s) of the copper-induced alterations on actin and fibronectin organization in mussel immunocytes is discussed.