Purification and immunolocalization of an annexin-like protein in pea seedlings

Abstract

As part of a study to identify potential targets of calcium action in plant cells, a 35-kDa, annexin-like protein was purified from pea (Pisum sativum L.) plumules by a method used to purify animal annexins. This protein, called p35, binds to a phosphatidylserine affinity column in a calcium-dependent manner and binds 45Ca2+ in a dot-blot assay. Preliminary sequence data confirm a relationship for p35 with the annexin family of proteins. Polyclonal antibodies have been raised which recognize p35 in Western and dot blots. Immunofluorescence and immunogold techniques were used to study the distribution and subcellular localization of p35 in pea plumules and roots. The highest levels of immunostain were found in young developing vascular cells producing wall thickenings and in peripheral root-cap cells releasing slime. This localization in cells which are actively involved in secretion is of interest because one function suggested for the animal annexins is involvement in the mediation of exocytosis.

Annexins of Plant Cells

“…Vol. 109, 1995 In pea plumules a single 35-kD annexin band on SDS-PAGE yields three isoforms with pls of 6.5, 6.8, and 7.0 when subjected to two-dimensional gel analysis, and in cotton fibers the 34-kD annexin band is made up of three isoforms ranging from pI of 6.1 to 6.5 (Clark et al, 1992;Andrawis et al, 1993). 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).…”

“…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.…”

Immunolocalization of a novel annexin‐like protein encoded by a stress and abscisic acid responsive gene in alfalfa

“…Some of the plant annexins have been shown to possess intrinsic phosphodiesterase activity (Calvert et al, 1996;McClung et al, 1994). Annexins were also suggested to play various roles in secretion and ripening processes (Blackbourn et al, 1992;Clark et al, 1992;Clark and Roux, 1995). Recently, an annexin-like protein has been identified from Arabidopsis thaliana which may be involved in the response to oxidative stress (Gidrol et al, 1996).…”

Imaging of the Yellow Cameleon 3.6 Indicator Reveals That Elevations in Cytosolic Ca2+ Follow Oscillating Increases in Growth in Root Hairs of Arabidopsis

“…Indeed, the structures of the apical actin cytoskeleton and its regulatory proteins (villins, gelsolins, and actin-depolymerizing factors; Smertenko et al, 1998;Tominaga et al, 2000;Allwood et al, 2001;Ketelaar et al, 2003;Fan et al, 2004) are thought to be regulated by the tip-high Ca 21 (Yokota et al, 2005). Similarly, there is evidence that annexins (Blackbourn et al, 1992;Clark et al, 1992;Carroll et al, 1998), phosphoinositide metabolism (Preuss et al, 2006), and calmodulin and protein kinases (Moutinho et al, 1998;Yoon et al, 2006) also play roles in sustaining tip growth and are regulated via the cytosolic Ca 21 gradient.…”