Sec14-like phosphatidylinositol transfer proteins and the biological landscape of phosphoinositide signaling in plants.

Phosphoinositides and soluble inositol phosphates are essential components of a complex intracellular chemical code that regulates major aspects of lipid signaling in eukaryotes. These involvements span a broad array of biological outcomes and activities, and cells are faced with the problem of how to compartmentalize and organize these various signaling events into a coherent scheme. It is in the arena of how phosphoinositide signaling circuits are integrated and, and how phosphoinositide pools are functionally defined and channeled to privileged effectors, that phosphatidylinositol (PtdIns) transfer proteins (PITPs) are emerging as critical players. As plant systems offer some unique advantages and opportunities for study of these proteins, we discuss herein our perspectives regarding the progress made in plant systems regarding PITP function. We also suggest interesting prospects that plant systems hold for interrogating how PITPs work, particularly in multi-domain contexts, to diversify the biological outcomes for phosphoinositide signaling. This article is part of a Special Issue entitled: Plant Lipid Biology edited by Kent D. Chapman and Ivo Feussner.

A national survey of assertive community treatment services in England.

BACKGROUND: Assertive community treatment (ACT) teams have been implemented across England since 1999. Although successful at engaging clients, the model has failed to show the same clinical effectiveness in trials in Europe as in the US and Australia. AIM: To investigate current ACT provision in England, aims of treatment and team managers' views of the effectiveness of ACT and the most important interventions. METHOD: Postal survey of managers of all services in England identifying as ACT teams. RESULTS: A total of 104/187 (56%) completed questionnaires were received. The majority of teams were in urban or mixed urban/rural areas. One third (36%) of teams had no psychiatrist, one half (48%) had no psychologist and less than a fifth (18%) had designated inpatient admission beds. The areas of intervention rated as most important by team managers could be delivered by non-professionally trained staff (engagement, accommodation, finances). The majority of managers reported positive clinical outcomes but only one third had collected data to support this. One third of teams were undergoing review or being reconfigured or closed. CONCLUSION: Successful client engagement is not being used as a vehicle to deliver evidence-based interventions. Many ACTs in England are not adequately staffed to deliver these.

Leveraging Wettability Engineering to Develop Three-Layer DIY Face Masks From Low-Cost Materials.

With the rapid spread of COVID-19 worldwide, the demand for appropriate face masks in the market has also skyrocketed. To ease strain on the supply of masks to the essential healthcare sector, it has become imperative that ordinary people rely more on home-made masks that can be easily put together using commonly available materials, while at the same time performing reasonably at arresting the ingress or egress of airborne droplets. Here, we propose a simple do-it-yourself (DIY) method for preparing a three-layered face mask that deploys two hydrophobic polypropylene nonwoven layers interspaced with a hydrophilic cellulosic cloth. The first hydrophobic layer, facing the user, allows high-momentum droplets (e.g., expelled by a sneeze or cough) to pass through and get absorbed in the next hydrophilic layer, thereby keeping the skin in contact with the mask dry and comfortable. The third (outermost) hydrophobic layer prevents penetration of the liquids from the middle layer to the outside, and also arrests any airborne droplets on its exterior. Simple tests show that our masks perform better in arresting the droplet transmission as compared to surgical masks available in the market.

Two-ligand priming mechanism for potentiated phosphoinositide synthesis is an evolutionarily conserved feature of Sec14-like phosphatidylinositol and phosphatidylcholine exchange proteins.

Lipid signaling, particularly phosphoinositide signaling, plays a key role in regulating the extreme polarized membrane growth that drives root hair development in plants. The Arabidopsis AtSFH1 gene encodes a two-domain protein with an amino-terminal Sec14-like phosphatidylinositol transfer protein (PITP) domain linked to a carboxy-terminal nodulin domain. AtSfh1 is critical for promoting the spatially highly organized phosphatidylinositol-4,5-bisphosphate signaling program required for establishment and maintenance of polarized root hair growth. Here we demonstrate that, like the yeast Sec14, the AtSfh1 PITP domain requires both its phosphatidylinositol (PtdIns)- and phosphatidylcholine (PtdCho)-binding properties to stimulate PtdIns-4-phosphate [PtdIns(4)P] synthesis. Moreover, we show that both phospholipid-binding activities are essential for AtSfh1 activity in supporting polarized root hair growth. Finally, we report genetic and biochemical evidence that the two-ligand mechanism for potentiation of PtdIns 4-OH kinase activity is a broadly conserved feature of plant Sec14-nodulin proteins, and that this strategy appeared only late in plant evolution. Taken together, the data indicate that the PtdIns/PtdCho-exchange mechanism for stimulated PtdIns(4)P synthesis either arose independently during evolution in yeast and in higher plants, or a suitable genetic module was introduced to higher plants from a fungal source and subsequently exploited by them.

Sec14-nodulin proteins and the patterning of phosphoinositide landmarks for developmental control of membrane morphogenesis.

Polarized membrane morphogenesis is a fundamental activity of eukaryotic cells. This process is essential for the biology of cells and tissues, and its execution demands exquisite temporal coordination of functionally diverse membrane signaling reactions with high spatial resolution. Moreover, mechanisms must exist to establish and preserve such organization in the face of randomizing forces that would diffuse it. Here we identify the conserved AtSfh1 Sec14-nodulin protein as a novel effector of phosphoinositide signaling in the extreme polarized membrane growth program exhibited by growing Arabidopsis root hairs. The data are consistent with Sec14-nodulin proteins controlling the lateral organization of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) landmarks for polarized membrane morphogenesis in plants. This patterning activity requires both the PtdIns(4,5)P2 binding and homo-oligomerization activities of the AtSfh1 nodulin domain and is an essential aspect of the polarity signaling program in root hairs. Finally, the data suggest a general principle for how the phosphoinositide signaling landscape is physically bit mapped so that eukaryotic cells are able to convert a membrane surface into a high-definition lipid-signaling screen.

Devising powerful genetics, biochemical and structural tools in the functional analysis of phosphatidylinositol transfer proteins (PITPs) across diverse species.

The minor cellular lipid phosphoinositides represents key regulators of diverse intracellular processes such as signal transduction at membrane-cytosol interface, regulation of membrane trafficking, cytoskeleton organization, nuclear events and the permeability, and transport functions of the membrane. The heterogeneous subcellular localization of phosphoinositides and their multiple and co-operative membrane-protein recognition mechanisms contribute to a "coincidence detection code" for the membrane-cytosol interactions in eukaryotic signaling networks. Such a "coincidence detection code" relies on the fine coordination of the broader lipid metabolism and organization, and their coupling to dedicated physiological processes. The phosphatidylinositol transfer proteins (PITPs) play a key regulatory role, essentially as "coincidence detectors" or "nanoreactors" in this "signal detection code" that spatially and temporally coordinate the diverse aspects of lipid metabolome with phosphoinositide signaling to effect various cellular functions. The integral role of PITPs in the highly conserved eukaryotic signal transduction strategy is amply demonstrated by the mammalian diseases associated with the derangements in the function of these proteins, to stress response and developmental regulation in plants, to fungal dimorphism and pathogenicity, to membrane trafficking in yeast and higher eukaryotes. The study of PITPs is fundamental to understanding of how the phosphoinositide signal transduction network is regulated and integrated to the larger lipid metabolome in diverse cellular processes. To comprehend how the PITPs integrate phosphoinositide signaling to broader lipid metabolome in diverse cellular processes, it is necessary to devise methods that can correlate the biochemical properties of these non-enzymatic proteins to biologically relevant functional insights. In this chapter, we present combinatorial approaches that primarily employ genetics and structural tools to assess the functional role of PITPs in yeast, plant and mammalian systems. An elaborate discussion on the various genetic models devised for interpreting the functional role of PITPs in relation to their operational assays has been included. We also describe the structural and biophysical methods that have advanced our understanding of how these proteins operate as "nanoreactor" molecules.

Phosphatidylinositol transfer proteins: negotiating the regulatory interface between lipid metabolism and lipid signaling in diverse cellular processes.

Phosphoinositides represent only a small percentage of the total cellular lipid pool. Yet, these molecules play crucial roles in diverse intracellular processes such as signal transduction at membrane-cytosol interface, regulation of membrane trafficking, cytoskeleton organization, nuclear events, and the permeability and transport functions of the membrane. A central principle in such lipid-mediated signaling is the appropriate coordination of these events. Such an intricate coordination demands fine spatial and temporal control of lipid metabolism and organization, and consistent mechanisms for specifically coupling these parameters to dedicated physiological processes. In that regard, recent studies have identified Sec14-like phosphatidylcholine transfer protein (PITPs) as "coincidence detectors," which spatially and temporally link the diverse aspects of the cellular lipid metabolome with phosphoinositide signaling. The integral role of PITPs in eukaryotic signal transduction design is amply demonstrated by the mammalian diseases associated with the derangements in the function of these proteins, to stress response and developmental regulation in plants, to fungal dimorphism and pathogenicity, to membrane trafficking in yeast, and higher eukaryotes. This review updates the recent advances made in the understanding of how these proteins, specifically PITPs of the Sec14-protein superfamily, operate at the molecular level and further describes how this knowledge has advanced our perception on the diverse biological functions of PITPs.

Two functionally distinctive phosphopantetheinyl transferases from amoeba Dictyostelium discoideum.

The life cycle of Dictyostelium discoideum is proposed to be regulated by expression of small metabolites. Genome sequencing studies have revealed a remarkable array of genes homologous to polyketide synthases (PKSs) that are known to synthesize secondary metabolites in bacteria and fungi. A crucial step in functional activation of PKSs involves their post-translational modification catalyzed by phosphopantetheinyl transferases (PPTases). PPTases have been recently characterized from several bacteria; however, their relevance in complex life cycle of protozoa remains largely unexplored. Here we have identified and characterized two phosphopantetheinyl transferases from D. discoideum that exhibit distinct functional specificity. DiAcpS specifically modifies a stand-alone acyl carrier protein (ACP) that possesses a mitochondrial import signal. DiSfp in contrast is specific to Type I multifunctional PKS/fatty acid synthase proteins and cannot modify the stand-alone ACP. The mRNA of two PPTases can be detected during the vegetative as well as starvation-induced developmental pathway and the disruption of either of these genes results in non-viable amoebae. Our studies show that both PPTases play an important role in Dictyostelium biology and provide insight into the importance of PPTases in lower eukaryotes.

Fractional order phase shaper design with Bode's integral for iso-damped control system.

The phase curve of an open loop system is flat in nature if the derivative of its phase with respect to frequency is zero. With a flat-phase curve, the corresponding closed loop system exhibits an iso-damped property i.e. maintains constant overshoot with the change of gain. This implies enhanced parametric robustness e.g. to variation in system gain. In the recent past, fractional order (FO) phase shapers have been proposed by contemporary researchers to achieve enhanced parametric robustness. In this paper, a simple methodology is proposed to design an appropriate FO phase shaper to achieve phase flattening in a control loop, comprising a plant controlled by a classical Proportional Integral Derivative (PID) controller. The methodology is demonstrated with MATLAB simulation of representative plants and accompanying PID controllers.

Dissecting the functional role of polyketide synthases in Dictyostelium discoideum: biosynthesis of the differentiation regulating factor 4-methyl-5-pentylbenzene-1,3-diol.

Dictyostelium discoideum exhibits the largest repository of polyketide synthase (PKS) proteins of all known genomes. However, the functional relevance of these proteins in the biology of this organism remains largely obscure. On the basis of computational, biochemical, and gene expression studies, we propose that the multifunctional Dictyostelium PKS (DiPKS) protein DiPKS1 could be involved in the biosynthesis of the differentiation regulating factor 4-methyl-5-pentylbenzene-1,3-diol (MPBD). Our cell-free reconstitution studies of a novel acyl carrier protein Type III PKS didomain from DiPKS1 revealed a crucial role of protein-protein interactions in determining the final biosynthetic product. Whereas the Type III PKS domain by itself primarily produces acyl pyrones, the presence of the interacting acyl carrier protein domain modulates the catalytic activity to produce the alkyl resorcinol scaffold of MPBD. Furthermore, we have characterized an O-methyltransferase (OMT12) from Dictyostelium with the capability to modify this resorcinol ring to synthesize a variant of MPBD. We propose that such a modification in vivo could in fact provide subtle variations in biological function and specificity. In addition, we have performed systematic computational analysis of 45 multidomain PKSs, which revealed several unique features in DiPKS proteins. Our studies provide a new perspective in understanding mechanisms by which metabolic diversity could be generated by combining existing functional scaffolds.