Regeneration of actin bundles in Chara: polarized growth and orientation by endoplasmic flow.

Strong irradiation of localized areas of the alga Chara produces chloroplast damage and extensive loss of the actin bundles responsible for cytoplasmic streaming. Immunofluorescence using a monoclonal antibody binding to the actin bundles has been used to follow their regrowth. Bundle regeneration is polarized so that new bundles develop from the ends of the actin bundles delivering endoplasm to the damaged area and not from bundles removing endoplasm. According to the previously established polarity of the actin filaments this growth is occurring from the "barbed" but not the "pointed" ends of the component filaments. The frequently irregular orientation of the regenerated bundles contrasts with the straight, parallel arrangement of the bundles before destruction. The arrangement of the regenerated bundles is suggested to depend on orientation by passive endoplasmic flow rather than a cortical template. As a result, bundles follow sweeping curves and can form a U-turn connecting oppositely polarized bundles normally separated by the neutral line. In addition to development in continuity with the free ends of pre-existing bundles, visualization of small, discrete fluorescent structures suggests that bundles can begin to form in isolation within the damaged areas. The results are discussed in terms of the polarized actin polymerisation seen in vitro, additional controls which may operate on bundle growth in vivo, and the ability of flow to orient F-actin. The relevance of the findings to normal cell ontogeny is assessed.