CYTOSOLIC CATABOLITES OF H-ATPASE IN SENESCING CARNATION PETALS()

The localisation of H+-ATPase catabolites in subcellular fractions iso lated from carnation petals has been determined immunologically using two antibodies, one raised against a mid-section of the protein (amino acids 340-650) and the other corresponding to the C-terminus. The nat ive M(r) 100000 polypeptide of the H+-ATPase was clearly discernible i n Western blots of microsomal membranes probed with either antibody; e ach antibody also depicted a distinguishable pattern of lower abundanc e catabolites of the protein in microsomal membranes. Some of these ca tabolites were present in the cytosolic fraction as well, in higher ab undance than in microsomal membranes. The antibodies also depicted dis tinguishable patterns of other catabolites of the H+-ATPase in the cyt osol that were not present in microsomal membranes and that changed wi th advancing senescence of the petals. Immunocytochemical localization of the Hf-ATPase and its catabolites using the antibody specific to t he C-terminus showed staining of the cytoplasm as well as the plasma m embrane. Moreover, the cytosolic H+-ATPase catabolites contain membran e-spanning domains of the protein and coeluted with phospholipid durin g Sepadex G-25 gel filtration. These observations indicate collectivel y that catabolites of the H+-ATPase formed within the plasma membrane are subsequently released into the Cytosol, possibly in association wi th lipid. Copyright (C) 1997 Elsevier Science Ltd