Anchoring mechanism of the intracellulan organization of actin cytoskeleton by the cell wall.

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In mesophyll cells of an aquatic angiosperm Vallisneria gigantea, bundles of actin filaments serve as the tracks for rotational streaming of the cytoplasm. From the previous studies, we hypothesized that the cell wall plays a crucial role to maintain the intracellular organization of the bundles of actin filaments through the plasma membrane as seen in the structure of focal adhesion in animal cells. Exogenously applied RGD or RYD hexapeptide caused a drastic disorder in the pattern of cytoplasmic streaming as well as in the intracellular array of actin filaments. Polyclonal antibodies raised against the RGD motif detected two polypeptides, 54 and 27 kDa, in the whole extract from the leaves, which were localized in the cell wall by immunofluorescence microscopy. Putative protein factors for these polypeptides were searched in highly purified plasma membrane fraction by affinity chromatography using the RGD peptide as a ligand. The eluted fraction with EDTA solution contained protein factors, whereas the amount of those was too small to be resolved after SDS PAGE.To know intracellular factors involved, the mode of disturbance of the tracks of cytoplasmic streaming was observed in a living mesophyll cell by video microscopy during plasmolysis and deplasmolysis. The bundle structure of the tracks seemed to be ramified only transiently. Even after recovery of the structure of bundles, the pattern of cytoplasmic streaming was unfixed and continued to change dynamically. These observations suggested the involvement of intracellular factors that bundle the actin filaments and fix the bundles to the plasma membrane and/or the cell wall.