Animal heterotrimeric G proteins are activated by guanine nucleotide exchange factors (GEF), typically seven transmembrane receptors that trigger GDP release and subsequent GTP binding. In contrast, ...the Arabidopsis thaliana G protein (AtGPA1) rapidly activates itself without a GEF and is instead regulated by a seven transmembrane Regulator of G protein Signaling (7TM-RGS) protein that promotes GTP hydrolysis to reset the inactive (GDP-bound) state. It is not known if this unusual activation is a major and constraining part of the evolutionary history of G signaling in eukaryotes. In particular, it is not known if this is an ancestral form or if this mechanism is maintained, and therefore constrained, within the plant kingdom. To determine if this mode of signal regulation is conserved throughout the plant kingdom, we analyzed available plant genomes for G protein signaling components, and we purified individually the plant components encoded in an informative set of plant genomes in order to determine their activation properties in vitro. While the subunits of the heterotrimeric G protein complex are encoded in vascular plant genomes, the 7TM-RGS genes were lost in all investigated grasses. Despite the absence of a Gα-inactivating protein in grasses, all vascular plant Gα proteins examined rapidly released GDP without a receptor and slowly hydrolyzed GTP, indicating that these Gα are self-activating. We showed further that a single amino acid substitution found naturally in grass Gα proteins reduced the Gα-RGS interaction, and this amino acid substitution occurred before the loss of the RGS gene in the grass lineage. Like grasses, non-vascular plants also appear to lack RGS proteins. However, unlike grasses, one representative non-vascular plant Gα showed rapid GTP hydrolysis, likely compensating for the loss of the RGS gene. Our findings, the loss of a regulatory gene and the retention of the "self-activating" trait, indicate the existence of divergent Gα regulatory mechanisms in the plant kingdom. In the grasses, purifying selection on the regulatory gene was lost after the physical decoupling of the RGS protein and its cognate Gα partner. More broadly these findings show extreme divergence in Gα activation and regulation that played a critical role in the evolution of G protein signaling pathways.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Arabidopsis, 7-transmembrane Regulator of G signaling protein 1 (AtRGS1) modulates canonical G protein signaling by promoting the inactive state of heterotrimeric G protein complex on the plasma ...membrane. It is known that plant leucine-rich repeat receptor-like kinases (LRR RLKs) phosphorylate AtRGS1 in vitro but little is known about the in vivo interaction, molecular dynamics, or the cellular consequences of this interaction.
Therefore, a subset of the known RLKs that phosphorylate AtRGS1 were selected for elucidation, namely, BAK1, BIR1, FLS2. Several microscopies for both static and dynamic protein-protein interactions were used to follow in vivo interactions between the RLKs and AtRGS1 after the presentation of the Pathogen-associated Molecular Pattern, Flagellin 22 (Flg22). These microscopies included Förster Resonance Energy Transfer, Bimolecular Fluoresence Complementation, and Cross Number and Brightness Fluorescence Correlation Spectroscopy. In addition, reactive oxygen species and calcium changes in living cells were quantitated using luminometry and R-GECO1 microscopy.
The LRR RLKs BAK1 and BIR1, interact with AtRGS1 at the plasma membrane. The RLK ligand flg22 sets BAK1 in motion toward AtRGS1 and BIR1 away, both returning to the baseline orientations by 10 minutes. The C-terminal tail of AtRGS1 is important for the interaction with BAK1 and for the tempo of the AtRGS1/BIR1 dynamics. This window of time corresponds to the flg22-induced transient production of reactive oxygen species and calcium release which are both attenuated in the rgs1 and the bak1 null mutants.
A temporal model of these interactions is proposed. flg22 binding induces nearly instantaneous dimerization between FLS2 and BAK1. Phosphorylated BAK1 interacts with and enables AtRGS1 to move away from BIR1 and AtRGS1 becomes phosphorylated leading to its endocytosis thus leading to de-repression by permitting AtGPA1 to exchange GDP for GTP. Finally, the G protein complex becomes dissociated thus AGB1 interacts with its effector proteins leading to changes in reactive oxygen species and calcium.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Investigators studying G protein-coupled signaling--often called the best-understood pathway in the world owing to intense research in medical fields--have adopted plants as a new model to explore ...the plasticity and evolution of G signaling. Much research on plant G signaling has not disappointed. Although plant cells have most of the core elements found in animal G signaling, differences in network architecture and intrinsic properties of plant G protein elements make G signaling in plant cells distinct from the animal paradigm. In contrast to animal G proteins, plant G proteins are self-activating, and therefore regulation of G activation in plants occurs at the deactivation step. The self-activating property also means that plant G proteins do not need and therefore do not have typical animal G protein-coupled receptors. Targets of activated plant G proteins, also known as effectors, are unlike effectors in animal cells. The simpler repertoire of G signal elements in Arabidopsis makes G signaling easier to manipulate in a multicellular context.
Size-fractionated samples of airborne particulate matter have been collected in a number of campaigns at Marylebone Road, London and simultaneously at background sites either in Regents Park or at ...North Kensington. Analysis of these samples has enabled size distributions of total mass and of a number of elements to be determined, and roadside increments attributable to nonexhaust emissions arising from traffic activity have been calculated. Taking a novel approach, the combined use of size distribution information and tracer elements has allowed the separate estimation of the contributions of brake dust, tire dust, and resuspension to particle mass in the range 0.9–11.5 μm aerodynamic diameter and mean contributions (±s.e.) at the Marylebone Road sampling site are estimated as resuspended dust 38.1 ± 9.7%, brake dust 55.3 ± 7.0%, and tire dust 10.7 ± 2.3%, (accounting for a total of 104.1% of coarse particle mass in the traffic increment above background).
Full text
Available for:
IJS, KILJ, NUK, PNG, UL, UM
We present a model for the effects of ligands on information transmission in G-Protein Coupled Receptor (GPCR) complexes. The model is built
entirely on principles of statistical mechanics and tenets ...of information transmission theory and was validated in part using agonist-induced effector activity and signaling bias for the angiotensin- and adrenergic-mediated signaling pathways, with
observations of phosphorylation sites on the C tail of the GPCR complex, and single-cell information-transmission experiments. The model extends traditional kinetic models that form the basis for many existing models of GPCR signaling. It is based on maximizing the rates of entropy production and information transmission through the GPCR complex. The model predicts that (1) phosphatase-catalyzed reactions, as opposed to kinase-catalyzed reactions, on the C-tail and internal loops of the GPCR are responsible for controlling the signaling activity, (2) signaling favors the statistical balance of the number of switches in the ON state and the number in the OFF state, and (3) biased-signaling response depends discontinuously on ligand concentration.
The
(sp
)-aryl sulfonate functional group is found in bioactive molecules, but their synthesis can involve extreme temperatures (>190 °C or flash vacuum pyrolysis) and strongly acidic reaction ...conditions. Inspired by the 1917 Tyrer industrial process for a sulfa dye that involved an aniline
(sp
)-SO
intermediate
to a
(sp
)-SO
rearranged product, we investigated tributylsulfoammonium betaine (TBSAB) as a milder
-sulfamation to
-sulfonate relay reagent. Initial investigations of a stepwise route involving TBSAB on selected anilines at room temperature enabled the isolation of
(sp
)-sulfamate. Subsequent thermal rearrangement demonstrated the intermediary of a sulfamate
to the sulfonate; however, it was low-yielding. Investigation of the
-sulfamate to
--sulfonate mechanism through control experiments with variation at the heteroatom positions and kinetic isotope experiments (KIE
) confirmed the formation of a key
(sp
)-SO
intermediate and further confirmed an
molecular mechanism. Furthermore, compounds without an accessible nitrogen (or oxygen) lone pair did not undergo sulfamation- (or sulfation) -to-sulfonation under these conditions. A one-pot sulfamation and thermal sulfonation reaction was ultimately developed and explored on a range of aniline and heterocyclic scaffolds with high conversions, including
(sp
)-sulfamates (
(sp
)-sulfates) and
(sp
)-sulfonates, in up to 99 and 80% (and 88% for a phenolic example) isolated yield, respectively. Encouragingly, the ability to modulate the
selectivity of the products obtained was observed under thermal control. A sulfonated analog of the intravenous anesthetic propofol was isolated (88% yield), demonstrating a proof-of-concept modification of a licensed drug alongside a range of nitrogen- and sulfur-containing heterocyclic fragments used in drug discovery.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The Beckmann rearrangement of ketoximes to their corresponding amides, using a Brønsted acid-mediated fragmentation and migration sequence, has found wide-spread industrial application. We postulated ...that the development of a methodology to access ylideneamino sulfates using tributylsulfoammonium betaine (TBSAB) would afford isolable Beckmann-type intermediates and competent partners for subsequent rearrangement cascades. The ylideneamino sulfates generated, isolated as their tributylammonium salts, are sufficiently activated to undergo Beckmann rearrangement without additional reagent activation. The generation of sulfuric acid in situ from the ylideneamino sulfate giving rise to a routine Beckmann rearrangement and additional amide bond cleavage to the corresponding aniline was detrimental to reaction success. The screening of bases revealed inexpensive sodium bicarbonate to be an effective additive to prevent classic Brønsted acid-mediated fragmentation and achieve optimal conversions of up to 99%.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
In eukaryotes, the combinatorial usage of cis-regulatory elements enables the assembly of composite genetic switches to integrate multifarious, convergent signals within a single promoter. Plants as ...sessile organisms, incapable of seeking for optimal conditions, rely on the use of this resource to adapt to changing environments. Emerging evidence suggests that the transcriptional responses of plants to stress are associated with epigenetic processes that govern chromatin accessibility. However, the extent at which specific chromatin modifications contribute to gene regulation has not been assessed.
In the present work, we combined methyl-sensitive-cut counting and RNA-seq to follow the transcriptional and epigenetic response of plants to simulated drought. Comprehensive genome wide evidence supports the notion that the methylome is widely reactive to water potential. The predominant changes in methylomes were loci in the promoters of genes encoding for proteins suited to cope with the environmental challenge.
These selective changes in the methylome with corresponding changes in gene transcription suggest drought sets in motion an instructive mechanism guiding epigenetic machinery toward specific effectors genes.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Plasma membrane-localized leucine-rich repeat receptor-like kinases directly activates G protein complex via interaction with seven transmembrane domain Regulator of G-protein Signaling 1 (AtRGS1) ...protein. Brassinosteroid insensitive 1 (BRI1) LIKE3 (BRL3) phosphorylates AtRGS1 in vitro. FRET analysis showed that BRL3 and AtRGS1 interaction dynamics change in response to glucose and flg22. Both BRL3 and AtRGS1 function in glucose sensing and brl3 and rgs1-2 single mutants are hyposensitive to high glucose as well as the brl3/rgs1 double mutant. BRL3 and AtRGS1 function in the same pathway linked to high glucose sensing. Hypocotyl elongation, another sugar-mediated pathway, is also implicated to be partially mediated by BRL3 and AtRGS1 because rgs1-2, brl3-2 and brl3-2/ rgs1-2 mutants share the long hypocotyl phenotype. BRL3 and AtRGS1 modulate the flg22-induced ROS burst and block one or more components positively regulating ROS production because the brl3/rgs1 double mutant has ~60% more ROS production than wild type while rgs1-2 has a partial ROS burst impairment and brl3 has slightly more ROS production. Here, we proposed a simple model where both BRL3 and AtRGS1 are part of a fine-tuning mechanism sensing glucose and flg22 to prevent excess ROS burst and control growth inhibition.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
This Personal Account describes the author's groups’ research in the field of electrosynthetic anodic oxidation, beginning with initial trial and error attempts with the Shono oxidation. Early ...setbacks with complex rotameric amide mixtures, provided the ideal environment for the discovery of the Oxa‐Shono reaction‐Osp2‐Csp3 bond cleavage of esters‐providing two useful products in one‐step: aldehyde selective oxidation level products and a mild de‐esterification method to afford carboxylic acids in the process. The development of the Oxa‐Shono reaction provided the impetus for the discovery of other electrically propelled‐Nsp2‐Csp2 and Nsp2‐Csp3‐bond breaking reactions in bioactive amide and sulfonamide systems. Understanding the voltammetric behaviour of the molecule under study, switching between controlled current‐ or controlled potential‐ electrolysis, and restricting electron flow (the reagent), affords exquisite control over the reaction outcomes in batch and flow. Importantly, this bio‐inspired advance in electrosynthetic dealkylation chemistry mimics the metabolic outcomes observed in nature.
In this account, I describe our group's investigations into electrosynthesis from fundamental electrode materials to cutting‐edge flow technologies. With an emphasis on voltammetry methods to discover new electrically driven reactions, dialling‐in new reactivity into the classic Shono‐type anodic oxidation
Full text
Available for:
FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
PDF
