Association of Phosphatidylinositol 4-Kinase with the Plant Cytoskeleton

Abstract

In eukaryotic cells, phosphatidylinositol 4-hydroxy kinase and phosphatidylinositol-4-phosphate 5-hydroxy kinase are responsible for the formation of the two second messenger precursors phosphatidylinositol-4-phosphate (Ptdlns(4)P) and phosphatidylinositol-4,5-bisphosphate (Ptdlns(4,5)P2). In plant cells, these kinases have been considered to be exclusively membrane associated, with the majority of activity residing in the inner leaflet of the plasmalemma. By sequentially extracting carrot protoplasts with the detergent Nonidet P-40 then more rigorously with Triton X-100, we were able to remove the activity of three separate plasma membrane marker enzymes and to demonstrate that a significant proportion of cellular Ptdlns 4-kinase is associated with the cytoskeleton. When only endogenous substrates were present, Nonidet P-40-permeabilized protoplasts and Nonidet P-40-extracted cytoskeletons displayed a pattern of lipid phosphorylation similar to that obtained with isolated plant membranes or permeabilized cells, whereas the Triton X-100-extracted cytoskeletons showed little or no activity. In contrast, when exogenous substrates were added, a major proportion of PtdlnsP formed was due to kinase activity associated with the cytoskeleton as well as nuclei. However, by subtracting the activity of isolated nuclei, it could be demonstrated that a significant proportion of the detergent-resistant Ptdlns kinase activity resides with the cytoskeletal fraction. These findings suggest that the pathways of polyphosphoinositide biosynthesis in plant cells should be reevaluated to take account of the cytoskeleton and that Ptdlns(4)P itself may play a unique role in modulation of plant cytoskeletal integrity and cellular signal transduction.

Association of Phosphatidylinositol Kinase, Phosphatidylinositol Monophosphate Kinase, and Diacylglycerol Kinase with the Cytoskeleton and F-Actin Fractions of Carrot (Daucus carota L.) Cells Grown in Suspension Culture

“…This is in agreement with the data from animal cells (20). While this paper was being revised, Xu et al (24) reported that PI kinase activity was associated with cytoskeleton isolated from carrot protoplasts; however, they could not detect DAG or PIP kinase activity. The fact that these authors could not detect DAG and PIP kinase activity and that we could may be due to differences in the starting material, the methods for preparing cytoskeleton, the assay conditions, or, in the case of DAG kinase, the substrate used.…”

Actin Purified from Maize Pollen Functions in Living Plant Cells

“…Protein immunoblotting was performed as described previously (Xu et al, 1992). Actin was detected using a mouse monoclonal antibody raised against chicken gizzard actin (Lessard, 1988; a kind gift of J.…”

Phosphatidylinositol 4-Phosphate Negatively Regulates Chloroplast Division in Arabidopsis

“…In addition to being important for signaling as a direct substrate for phospholipase C (Gonorazky et al, 2010) or as a precursor for phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ] (Berridge and Irvine, 1989), PI4P itself acts as a signaling lipid (Jung et al, 2002;Preuss et al, 2006;Chapman and Goring, 2011). PI4P is synthesized by phosphatidylinositol 4-kinase (PI4K), and PI4K activities are present in many cellular compartments, including the plasma membrane (Sommarin and Sandelius, 1988), cytosol (Okpodu et al, 1995), cytoskeleton (Xu et al, 1992), and nucleus (Hendrix et al, 1989). The existence of PI kinase activity was reported in chloroplast envelope membranes, and phosphatidylinositol 3-phosphate (PI3P) and PI4P were identified as its products (Bovet et al, 2001).…”