PYRENOID FORMATION ASSOCIATED WITH THE CELL CYCLE IN THE BROWN ALGA,SCYTOSIPHON LOMENTARIA(SCYTOSIPHONALES, PHAEOPHYCEAE)¹

Abstract

Vegetative cells of the brown alga Scytosiphon lomentaria (Lyngbye) Link characteristically have only one chloroplast with a prominent protruding pyrenoid, whereas zygotes have both paternal and maternal chloroplasts. In zygotes, before cell and chloroplast division, each chloroplast has an old and a new pyrenoid. In this study, we raised a polyclonal antibody to RUBISCO and examined the distribution of RUBISCO by immunofluorescence microscopy, focusing on new pyrenoid formation in vegetative cells of gametophytes and zygotes in Scytosiphon. In interphase, only one old pyrenoid was positively indicated by anti-RUBISCO antibody in vegetative cells of gametophytes. From mid-S phase, small fluorescence aggregates reflecting RUBISCO localization started to appear at stroma positions other than adjacent to the old protruding pyrenoid. The fluorescent spots eventually coalesced into a protrusion into the adjacent cytoplasm. We also used inhibitors to clarify the relationship between the cell cycle and new pyrenoid formation, using zygotes after fertilization. When DNA replication was blocked by aphidicolin, new pyrenoid formation was also inhibited. Washing out aphidicolin permitted new pyrenoid formation with the progression of the cell cycle. When mitosis was prolonged by nocodazole, which disrupted the spindle microtubules, the fluorescent masses indicating RUBISCO localization continued to increase when compared with pyrenoid formation in untreated zygotes. During treatment with chloramphenicol, mitosis and cytokinesis were completed. However, there was no occurrence of new RUBISCO localization within the chloroplast stroma beyond the old pyrenoid. From these observations, it seems clear that new pyrenoid formation in the brown alga Scytosiphon depends on the cell cycle.

Identification and characterization of a fluorescent flagellar protein from the brown algaScytosiphon lomentaria(Scytosiphonales, Phaeophyceae): A flavoprotein homologous to Old Yellow Enzyme

“…A broad band around 52 kDa in the intact zoospores was mainly ribulose-bisphosphate carboxylase/oxygenase, as evaluated by Western blotting analysis (data not shown, cf. Nagasato et al, 2003). The fluorescence spectra confirmed that flavin-like substances were present in the posterior flagella, as previously reported (Mu¨ller et al, 1987;Kawai, 1988;Kawai et al, 1996).…”

Re‐examination of ultrastructures of the stellate chloroplast organization in brown algae: Structure and development of pyrenoids

“…The samples were incubated for 30 min at 37°C in blocking solution (2.5% skim milk, 5% normal goat serum, and 0.05% NaN 3 in PBS). Afterward, they were incubated overnight at 22°C with the anti‐RUBISCO antibody (diluted 1:1000 in PBS; Nagasato et al . 2003).…”

“…Dynamic changes in de novo formation of the pyrenoid and distribution of RUBISCO molecules in the chloroplast were observed during mitosis in Scytosiphon lomentaria (Nagasato & Motomura 2002a; Nagasato et al . 2003).…”

“…2003). New pyrenoid formation is associated with the cell cycle, and appearance of the new pyrenoid, independent of the old one, occurs before chloroplast division (Nagasato & Motomura 2002a; Nagasato et al . 2003).…”

Characterization of centrin genes from Ochromonas danica (Chrysophyceae) and Scytosiphon lomentaria (Phaeophyceae)

“…The unialgal strain of the gametophyte of Scytosiphon lomentaria (Lyngbye) Link was described previously (Nagasato & Motomura 2003). It was cultured in PESI medium (Tatewaki 1966) at 18 ° C long day condition; namely, 14:10 h LD, 30-40 µ mol photons m -2 s -1 ).…”