Functions and Mechanisms of SAC Phosphoinositide Phosphatases in Plants

Phosphatidylinositol (PtdIns) is one type of phospholipid comprising an inositol head group and two fatty acid chains covalently linked to the diacylglycerol group. In addition to their roles as compositions of cell membranes, phosphorylated PtdIns derivatives, termed phosphoinositides, execute a wide range of regulatory functions. PtdIns can be phosphorylated by various lipid kinases at 3-, 4- and/or 5- hydroxyls of the inositol ring, and the phosphorylated forms, including PtdIns3P, PtdIns4P, PtdIns5P, PtdIns(3,5)P2, PtdIns(4,5)P2, can be reversibly dephosphorylated by distinct lipid phosphatases. Amongst many other types, the SUPPRESSOR OF ACTIN (SAC) family of phosphoinositide phosphatases recently emerged as important regulators in multiple growth and developmental processes in plants. Here, we review recent advances on the biological functions, cellular activities, and molecular mechanisms of SAC domain-containing phosphoinositide phosphatases in plants. With a focus on those studies in the model plant Arabidopsis thaliana together with progresses in other plants, we highlight the important roles of subcellular localizations and substrate preferences of various SAC isoforms in their functions.

Organismal roles for the PI3Kα and β isoforms: their specificity, redundancy or cooperation is context-dependent

PI3Ks are important lipid kinases that produce phosphoinositides phosphorylated in position 3 of the inositol ring. There are three classes of PI3Ks: class I PI3Ks produce PIP3 at plasma membrane level. Although D. melanogaster and C. elegans have only one form of class I PI3K, vertebrates have four class I PI3Ks called isoforms despite being encoded by four different genes. Hence, duplication of these genes coincides with the acquisition of coordinated multi-organ development. Of the class I PI3Ks, PI3Kα and PI3Kβ, encoded by PIK3CA and PIK3CB, are ubiquitously expressed. They present similar putative protein domains and share PI(4,5)P2 lipid substrate specificity. Fifteen years after publication of their first isoform-selective pharmacological inhibitors and genetically engineered mouse models (GEMMs) that mimic their complete and specific pharmacological inhibition, we review the knowledge gathered in relation to the redundant and selective roles of PI3Kα and PI3Kβ. Recent data suggest that, further to their redundancy, they cooperate for the integration of organ-specific and context-specific signal cues, to orchestrate organ development, physiology, and disease. This knowledge reinforces the importance of isoform-selective inhibitors in clinical settings.

Function of AtPGAP1 in GPI anchor lipid remodeling and transport to the cell surface of GPI-anchored proteins

ABSTRACTGPI-anchored proteins (GPI-APs) play an important role in a variety of plant biological processes including growth, stress response, morphogenesis, signalling and cell wall biosynthesis. The GPI-anchor contains a lipid-linked glycan backbone that is synthesized in the endoplasmic reticulum (ER) where it is subsequently transferred to the C-terminus of proteins containing a GPI signal peptide by a GPI transamidase. Once the GPI anchor is attached to the protein, the glycan and lipid moieties are remodelled. In mammals and yeast, this remodelling is required for GPI-APs to be included in Coat Protein II (COPII) coated vesicles for their ER export and subsequent transport to the cell surface. The first reaction of lipid remodelling is the removal of the acyl chain from the inositol group by Bst1p (yeast) and PGAP1 (mammals). In this work, we have used a loss-of-function approach to study the role of PGAP1/Bst1 like genes in plants. We have found that Arabidopsis PGAP1 localizes to the ER and probably functions as the GPI inositol-deacylase which cleaves the acyl chain from the inositol ring of the GPI anchor. In addition, we show that PGAP1 function is required for efficient ER export and transport to the cell surface of GPI-APs.One sentence summaryGPI anchor lipid remodeling in GPI-anchored proteins is required for their transport to the cell surface in Arabidopsis.

Can Inositol Pyrophosphates Inform Strategies for Developing Low Phytate Crops?

Inositol pyrophosphates (PP-InsPs) are an emerging class of “high-energy” intracellular signaling molecules, containing one or two diphosphate groups attached to an inositol ring, that are connected with phosphate sensing, jasmonate signaling, and inositol hexakisphosphate (InsP6) storage in plants. While information regarding this new class of signaling molecules in plants is scarce, the enzymes responsible for their synthesis have recently been elucidated. This review focuses on InsP6 synthesis and its conversion into PP-InsPs, containing seven and eight phosphate groups (InsP7 and InsP8). These steps involve two types of enzymes: the ITPKs that phosphorylate InsP6 to InsP7, and the PPIP5Ks that phosphorylate InsP7 to InsP8. This review also considers the potential roles of PP-InsPs in plant hormone and inorganic phosphate (Pi) signaling, along with an emerging role in bioenergetic homeostasis. PP-InsP synthesis and signaling are important for plant breeders to consider when developing strategies that reduce InsP6 in plants, as this will likely also reduce PP-InsPs. Thus, this review is primarily intended to bridge the gap between the basic science aspects of PP-InsP synthesis/signaling and breeding/engineering strategies to fortify foods by reducing InsP6.

The Function of Inositol Phosphatases in Plant Tolerance to Abiotic Stress

Inositol signaling is believed to play a crucial role in various aspects of plant growth and adaptation. As an important component in biosynthesis and degradation of myo-inositol and its derivatives, inositol phosphatases could hydrolyze the phosphate of the inositol ring, thus affecting inositol signaling. Until now, more than 30 members of inositol phosphatases have been identified in plants, which are classified intofive families, including inositol polyphosphate 5-phosphatases (5PTases), suppressor of actin (SAC) phosphatases, SAL1 phosphatases, inositol monophosphatase (IMP), and phosphatase and tensin homologue deleted on chromosome 10 (PTEN)-related phosphatases. The current knowledge was revised here in relation to their substrates and function in response to abiotic stress. The potential mechanisms were also concluded with the focus on their activities of inositol phosphatases. The general working model might be that inositol phosphatases would degrade the Ins(1,4,5)P3 or phosphoinositides, subsequently resulting in altering Ca2+ release, abscisic acid (ABA) signaling, vesicle trafficking or other cellular processes.

Inositol Trisphosphate Kinase and Diphosphoinositol Pentakisphosphate Kinase Enzymes Constitute the Inositol Pyrophosphate Synthesis Pathway in Plants

ABSTRACTInositol pyrophosphates (PP-InsPs) are an emerging class of “high-energy” intracellular signaling molecules containing one or two diphosphate groups attached to an inositol ring, with suggested roles in bioenergetic homeostasis and inorganic phosphate (Pi) sensing. Information regarding the biosynthesis of these unique class of signaling molecules in plants is scarce, however the enzymes responsible for their biosynthesis in other eukaryotes have been well described. Here we report the characterization of the two Arabidopsis VIP kinase domains, a newly discovered activity of the Arabidopsis ITPK1 and ITPK2 enzymes, and the subcellular localization of the enzymes involved in the synthesis of InsP6and PP-InsPs. Our data indicate that AtVIP1-KD and AtVIP2-KD act primarily as 1PP-specific Diphosphoinositol Pentakisphosphate Kinases (PPIP5) Kinases. The AtITPK enzymes, in contrast, can function as InsP6kinases, and thus are the missing enzyme in the plant PP-InsP synthesis pathway. Together, these enzyme classes can function in plants to produce PP-InsPs, which have been implicated in signal transduction and Pisensing pathways. We measured a higher InsP7level (increased InsP7/InsP8ratio) invip1/vip2double loss-of-function mutants, and an accumulation of InsP8(decreased InsP7/InsP8ratio) in the 35S:VIP2overexpression line relative to wild-type plants. We also report that enzymes involved in the synthesis of InsPs and PP-InsPs accumulate within the nucleus and cytoplasm of plant cells. Our work defines a molecular basis for understanding how plants synthesize PP-InsPs which is crucial for determining the roles of these signaling molecules in processes such as Pisensing.SIGNIFICANCE STATEMENTInositol pyrophosphate signaling molecules are of agronomic importance as they can control complex responses to the limited nutrient, phosphate. This work fills in the missing steps in the inositol pyrophosphate synthesis pathway and points to a role for these molecules in the plant cell nucleus. This is an important advance that can help us design future strategies to increase phosphate efficiency in plants.

Inositol phosphates and phosphoinositides activate insulin-degrading enzyme, while phosphoinositides also mediate binding to endosomes

Insulin-degrading enzyme (IDE) hydrolyzes bioactive peptides, including insulin, amylin, and the amyloid β peptides. Polyanions activate IDE toward some substrates, yet an endogenous polyanion activator has not yet been identified. Here we report that inositol phosphates (InsPs) and phosphatdidylinositol phosphates (PtdInsPs) serve as activators of IDE. InsPs and PtdInsPs interact with the polyanion-binding site located on an inner chamber wall of the enzyme. InsPs activate IDE by up to ∼95-fold, affecting primarily Vmax. The extent of activation and binding affinity correlate with the number of phosphate groups on the inositol ring, with phosphate positional effects observed. IDE binds PtdInsPs from solution, immobilized on membranes, or presented in liposomes. Interaction with PtdInsPs, likely PtdIns(3)P, plays a role in localizing IDE to endosomes, where the enzyme reportedly encounters physiological substrates. Thus, InsPs and PtdInsPs can serve as endogenous modulators of IDE activity, as well as regulators of its intracellular spatial distribution.

Phosphate, inositol and polyphosphates

Eukaryotic cells have ubiquitously utilized the myo-inositol backbone to generate a diverse array of signalling molecules. This is achieved by arranging phosphate groups around the six-carbon inositol ring. There is virtually no biological process that does not take advantage of the uniquely variable architecture of phosphorylated inositol. In inositol biology, phosphates are able to form three distinct covalent bonds: phosphoester, phosphodiester and phosphoanhydride bonds, with each providing different properties. The phosphoester bond links phosphate groups to the inositol ring, the variable arrangement of which forms the basis of the signalling capacity of the inositol phosphates. Phosphate groups can also form the structural bridge between myo-inositol and diacylglycerol through the phosphodiester bond. The resulting lipid-bound inositol phosphates, or phosphoinositides, further expand the signalling potential of this family of molecules. Finally, inositol is also notable for its ability to host more phosphates than it has carbons. These unusual organic molecules are commonly referred to as the inositol pyrophosphates (PP-IPs), due to the presence of high-energy phosphoanhydride bonds (pyro- or diphospho-). PP-IPs themselves constitute a varied family of molecules with one or more pyrophosphate moiety/ies located around the inositol. Considering the relationship between phosphate and inositol, it is no surprise that members of the inositol phosphate family also regulate cellular phosphate homoeostasis. Notably, the PP-IPs play a fundamental role in controlling the metabolism of the ancient polymeric form of phosphate, inorganic polyphosphate (polyP). Here we explore the intimate links between phosphate, inositol phosphates and polyP, speculating on the evolution of these relationships.

Ice nucleation of an insect lipoprotein ice nucleator (LPIN) correlates with retardation of the hydrogen bond dynamics at the myo-inositol ring

Remarkably little is known about the mechanism of action of ice nucleation proteins (INPs), although their ability to trigger ice nucleation could be used in a broad variety of applications.

Trypanosoma brucei Bloodstream Forms Depend upon Uptake ofmyo-Inositol for Golgi Complex Phosphatidylinositol Synthesis and Normal Cell Growth

ABSTRACTmyo-Inositol is a building block for all inositol-containing phospholipids in eukaryotes. It can be synthesizedde novofrom glucose-6-phosphate in the cytosol and endoplasmic reticulum. Alternatively, it can be taken up from the environment via Na+- or H+-linkedmyo-inositol transporters. While Na+-coupledmyo-inositol transporters are found exclusively in the plasma membrane, H+-linkedmyo-inositol transporters are detected in intracellular organelles. InTrypanosoma brucei, the causative agent of human African sleeping sickness,myo-inositol metabolism is compartmentalized.De novo-synthesizedmyo-inositol is used for glycosylphosphatidylinositol production in the endoplasmic reticulum, whereas themyo-inositol taken up from the environment is used for bulk phosphatidylinositol synthesis in the Golgi complex. We now provide evidence that the Golgi complex-localizedT. bruceiH+-linkedmyo-inositol transporter (TbHMIT) is essential in bloodstream-formT. brucei. Downregulation of TbHMIT expression by RNA interference blocked phosphatidylinositol production and inhibited growth of parasites in culture. Characterization of the transporter in a heterologous expression system demonstrated a remarkable selectivity of TbHMIT formyo-inositol. It tolerates only a single modification on the inositol ring, such as the removal of a hydroxyl group or the inversion of stereochemistry at a single hydroxyl group relative tomyo-inositol.