Polar distribution of annexin-like proteins during phytochrome-mediated initiation and growth of rhizoids in the ferns Dryopteris and Anemia

Abstract

Although the calcium requirement of phytochrome-mediated fern spore germination and early rhizoid growth is well established, the calcium-binding proteins that serve as transducers for these responses are not known. Here we report the presence of annexin-like proteins in germinating spores of Dryopteris filix-mas (L.) Schott and Anemia phyllitidis (L.) Sw. and evidence that they may be important participants in early photomorphogenic changes in gametophytes. Immunolocalization and immunoblot assays of these proteins were carried out using polyclonal antibodies raised either against a 35-kDa annexin-like protein from pea or against anchorin CII from chicken. Western-blot analysis showed that crude protein extracts obtained from both species after red-light treatment contained two cross-reactive protein bands with molecular weights around 70 kDa. These proteins were annexin-like in that they bound to a phosphatidylserine affinity column in a calcium-dependent fashion. Using this column, two protein bands around 70 kDa, i.e. 67 and 73 kDa, were partially purified together with proteins at 36 kDa and a doublet at 54 kDa. Proteins of these latter molecular weights are suggested to be members of the annexin family, but no cross-reactivity could be found between these and the two antibodies used in our investigations. Immunodetectable levels of these proteins were observed only after light-mediated induction of spore germination. Imaging of the immuno-localization patterns observed with both antibodies showed that the annexin-like proteins are concentrated at the extreme tips of the rhizoids in D. filix-mas and A. phyllitidis during rhizoid initiation and all stages of elongation. We suggest that these proteins may play a major role in the tip-oriented exocytosis events that are critical for the initiation and growth of fern rhizoids.

Characterization and gene expression of an annexin during fruit development in Capsicum annuum

“…These 2 proteins might be encoded by divergent mRNAs, not detected by the E511 cDNA probe. The 2 other proteins are not recognized by the maize antiserum, but the use of antibodies from mammals can provide an alternative assay to detect other annexins (immunoblottings were carried out using polyclonal antibodies raised against anchorin CII from chicken to identify annexins in the fern rhizoids) [18].…”

Annexins of Plant Cells

“…These different isoforms may represent a single gene product that has undergone posttranslational modification, such as phosphorylation, or different gene products, such as seen in rat liver, where three different annexin gene products co-migrate at 35 kD on SDS-PAGE (Kaetzel et al, 1989). High molecular mass (67-70 kD) annexins have been observed in Tradescantia, maize, peas, and ferns (Blackbourn et al, 1991;Clark et al, 1995). cDNA sequence data for these higher molecular mass annexin-like proteins will resolve whether these proteins are homologous with annexin VI, which has eight repeats, or represent a stable dimer caused by transglutamination, as observed in the case of annexin I (Ando et al, 1989(Ando et al, , 1991.…”

“…The staining pattern in pea and corn seedlings reveals that annexins are highly concentrated in secretory cell types, such as the outer cells of the root cap, epidermal cells, and developing xylem and phloem cells (Clark et al, 1992(Clark et al, , 1994. Immunolocalization results also indicate that there are high levels of annexins in the tip region of polarly growing cells such as pollen tubes and fern rhizoids, cell types that are actively involved in the Golgi-mediated secretion of new wall materials and plasma membrane (Blackbourn et al, 1992;Clark et al, 1995). In the case of pollen tubes it has been shown that there are higher levels of Ca2+ at the tip (Miller et al, 19921, and it has been suggested that annexins may be one target for this Ca2+ signal.…”

“…The evidence that Ca2+ is involved in certain types of secretion in plant cells has been reviewed (Steer, 1988), and the potential role of annexins in exocytosis in plants has been discussed Clark et al, 1995). It is still unclear what the leve1 of free Ca2' is at the site of vesicle fusion with the plasma membrane during secretion in plant cells.…”

“…Immunolocalization studies of plant annexins have been carried out in pea plumules and roots (Clark et al, 1992), Lilium pollen tubes (Blackbourn et al, 1992), corn roots (Clark et al, 1994), and fern rhizoids (Clark et al, 1995). The staining pattern in pea and corn seedlings reveals that annexins are highly concentrated in secretory cell types, such as the outer cells of the root cap, epidermal cells, and developing xylem and phloem cells (Clark et al, 1992(Clark et al, , 1994.…”