IN THIS ISSUE

Plant PtdIns 3-Kinase Goes Nuclear

Phosphoinositides are dynamic and vital members of the cell's repertoire of signaling molecules. They are involved in regulating diverse processes, such as cytoskeletal organization, membrane trafficking, and ion transport (for review see Drobak 1992 ; Munnik et al. 1998 ; Stevenson et al. 2000 ). The functional diversity of the polyphosphorylated inositol lipids is in part predi-cated by the stereospecificity of the phosphate groups on the inositol ring and by the subcellular localization of the phospholipids. In eukaryotic cells, the multiple phosphorylated isomers and the specific lipid kinases involved in their synthesis, (phosphatidylinositol [PtdIns] 3-kinases, PtdIns 4-kinases and PtdInsP kinases), are located in various intracellular compartments, including the plasma membrane, endomembranes, the cytoskeleton, or the nucleus.

The PtdIns 3-kinase family comprises three classes of enzymes that phosphorylate inositol phospholipids specifically at the D-3 position of the inositol ring (Wyman and Pirola, 1998). Both the class I and class II PtdIns 3-kinases are an integral part of receptor-mediated signaling pathways prevalent in animal cells. In contrast, plants and yeast contain only class III PtdIns 3-kinases. The class III PtdIns 3-kinases are typified by the yeast Vps34 protein (Stack and Emr 1994 ). In temperature-sensitive mutants of Vps34p, proteins normally targeted to the vacuole are misrouted (Stack et al. 1995 ), implying a role for PtdIns 3P in vesicle trafficking. Class III-type PtdIns 3-kinase genes have been isolated and characterized from soybean (Hong and Verma 1994 ) and from Arabidopsis (Welters et al. 1994 ). Previous reports in plants have supported a potential function for the PtdIns 3-kinases in vesicle trafficking and vacuolar sorting similar to yeast (Matsuoka et al. 1995 ).

In this issue of THE PLANT CELL, Bunney et al. (pages 1679–1688) provide a new perspective on the role of PtdIns 3-kinase in plant cells: their results show that PtdIns 3P is formed in plant nuclei and that a class III PtdIns 3-kinase is localized at active nuclear transcription sites. The authors first demonstrate that both PtdIns 3P and PtdIns 4P are present in isolated soybean nuclei, and the identity of the lipids is confirmed by high performance liquid chromatography of deacylated lipid headgroups. When nuclei were isolated in the presence of detergent, the amount of PtdIns 4P in the nuclei was reduced, whereas the amount of PtdIns 3P was unchanged. From these data, the authors conclude that PtdIns 4-kinase activity is associated with the nuclear envelope, whereas PtdIns 3-kinase activity and PtdIns 3P reside within the nuclear matrix. In order to study the nuclear localization of PtdIns 3-kinase in more detail, a monoclonal antibody was generated against soybean PtdIns 3-kinase and used for immunolocalization studies. Immunolabeling of PtdIns 3-kinase was observed mainly in the nucleolus. Root sections were probed with both anti-PtdIns 3-kinase and anti-Br-UTP antibodies, after in vitro transcription in the presence of Br-UTP. Colocalization of the PtdIns 3-kinase and Br-UTP signals suggests that PtdIns 3-kinase is associated with active nuclear or nucleolar transcription sites. From these data, a new function is implied for plant PtdIns 3-kinase and/or PtdIns 3P in transcriptional regulation, in addition to previously established functions in vesicle trafficking.

Although the functional significance of a nuclear phosphoinositide cycle is not well understood, it is becoming increasingly clear that in animals and yeast many of the phosphoinositides and the corresponding lipid kinases are present in the nucleus (Divecha et al. 2000 ). Both type I and type II PtdInsP kinases have been shown to be associated with distinct subnuclear domains, known as nuclear speckles, which are functionally linked to mRNA metabolism and, therefore, may play a role in mRNA splicing (Boronenkov et al. 1998 ). A role for phosphoinositides in regulating nuclear transcription is also implicated by the specific binding of PtdIns(4,5)P₂ to histones H1 and H3 (Yu et al. 1998 ). York et al. 1999 have shown that inositol hexakisphosphate (InsP₆) plays an essential role in the export of mRNA from the nucleus and that a kinase with dual specificity for Ins(1,4,5)P₃ and Ins(1,4,5,6)P₄ is a vital part of a complex of transcription factors directing the expression of genes involved in arginine metabolism (Odom et al. 2000 ).

Some evidence exists from the animal literature that the class I, and potentially class II, PtdIns 3-kinases are present in the nucleus (Maraldi et al. 1999 ) and that they may translocate to the nucleus in response to stimulation (Bavelloni et al. 1999 ). However, no data have been reported so far in any eukaryotic system of a class III PtdIns 3-kinase in the nucleus. The information provided by Bunney et al. adds another piece to the puzzle that is the complex regulatory network of phosphoinositides and thereby opens new avenues for investigation. For example, it will be interesting to see whether translocation of a class III PtdIns 3-kinase to the nucleus plays a role in regulating transcription in response to a stimulus in plants. Because of the relative simplicity of plant PtdIns-3P metabolism, studies of the plant nuclear PtdIns 3-kinases should continue to provide insights that will help to delineate the functional roles of nuclear phosphoinositides.

The results presented by Bunney et al. are intriguing also from an evolutionary standpoint because the class III PtdIns 3-kinases are found throughout the eukaryotic realm in mammals, insects, slime molds, yeast, algae, and plants and may represent the primordial form of PtdIns 3-kinases. The ubiquitous nature of the class III PtdIns 3-kinases and their nuclear localization may reflect functional evolution of PtdIns 3P from the nucleus to membrane vesicles and ultimately the plasma membrane. In this regard, it will be important to determine whether a nuclear role for the enigmatic PtdIns 3P is unique to plants or whether the findings by Bunney et al. will be corroborated in other eukaryotic systems. In any event, in a field in which plant science is frequently modeled after animal research, it is inspiring when a new paradigm is proposed based on novel plant phosphoinositide research.

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