scholarly journals Low ABA concentration promotes root growth and hydrotropism through relief of ABA INSENSITIVE 1-mediated inhibition of plasma membrane H+-ATPase 2

2021 ◽  
Vol 7 (12) ◽  
pp. eabd4113
Author(s):  
Rui Miao ◽  
Wei Yuan ◽  
Yue Wang ◽  
Irene Garcia-Maquilon ◽  
Xiaolin Dang ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Root Growth ◽  
Experimental System ◽  
Aba Signaling ◽  
Wild Type ◽  
Normal Medium ◽  
C Terminus ◽  
Quadruple Mutant ◽  
Aba Insensitive ◽  
Aba Receptors

The hab1-1abi1-2abi2-2pp2ca-1 quadruple mutant (Qabi2-2) seedlings lacking key negative regulators of ABA signaling, namely, clade A protein phosphatases type 2C (PP2Cs), show more apoplastic H+ efflux in roots and display an enhanced root growth under normal medium or water stress medium compared to the wild type. The presence of low ABA concentration (0.1 micromolar), inhibiting PP2C activity via monomeric ABA receptors, enhances root apoplastic H+ efflux and growth of the wild type, resembling the Qabi2-2 phenotype in normal medium. Qabi2-2 seedlings also demonstrate increased hydrotropism compared to the wild type in obliquely-oriented hydrotropic experimental system, and asymmetric H+ efflux in root elongation zone is crucial for root hydrotropism. Moreover, we reveal that Arabidopsis ABA-insensitive 1, a key PP2C in ABA signaling, interacts directly with the C terminus of Arabidopsis plasma membrane H+-dependent adenosine triphosphatase 2 (AHA2) and dephosphorylates its penultimate threonine residue (Thr947), whose dephosphorylation negatively regulates AHA2.

scholarly journals A Screen for Genes That Function in Abscisic Acid Signaling in Arabidopsis thaliana

Genetics ◽  
2002 ◽  
Vol 161 (3) ◽  
pp. 1247-1255 ◽  
Author(s):  
Eiji Nambara ◽  
Masaharu Suzuki ◽  
Suzanne Abrams ◽  
Donald R McCarty ◽  
Yuji Kamiya ◽  
...  
Keyword(s):  
Abscisic Acid ◽  
Growth And Development ◽  
Plant Hormone ◽  
The Other ◽  
Aba Signaling ◽  
Wild Type ◽  
Plant Growth And Development ◽  
Diverse Range ◽  
Abscisic Acid Signaling ◽  
Aba Insensitive

Abstract The plant hormone abscisic acid (ABA) controls many aspects of plant growth and development under a diverse range of environmental conditions. To identify genes functioning in ABA signaling, we have carried out a screen for mutants that takes advantage of the ability of wild-type Arabidopsis seeds to respond to (−)-(R)-ABA, an enantiomer of the natural (+)-(S)-ABA. The premise of the screen was to identify mutations that preferentially alter their germination response in the presence of one stereoisomer vs. the other. Twenty-six mutants were identified and genetic analysis on 23 lines defines two new loci, designated CHOTTO1 and CHOTTO2, and a collection of new mutant alleles of the ABA-insensitive genes, ABI3, ABI4, and ABI5. The abi5 alleles are less sensitive to (+)-ABA than to (−)-ABA. In contrast, the abi3 alleles exhibit a variety of differences in response to the ABA isomers. Genetic and molecular analysis of these alleles suggests that the ABI3 transcription factor may perceive multiple ABA signals.

scholarly journals Palmitoylation of Sindbis Virus TF Protein Regulates Its Plasma Membrane Localization and Subsequent Incorporation into Virions

2016 ◽  
Vol 91 (3) ◽  
Author(s):  
Jolene Ramsey ◽  
Emily C. Renzi ◽  
Randy J. Arnold ◽  
Jonathan C. Trinidad ◽  
Suchetana Mukhopadhyay
Keyword(s):  
Plasma Membrane ◽  
Sindbis Virus ◽  
Molecular Mechanisms ◽  
Wild Type Virus ◽  
Membrane Localization ◽  
Clear Understanding ◽  
Wild Type ◽  
Plasma Membrane Localization ◽  
C Terminus ◽  
N Terminus

ABSTRACT Palmitoylation is a reversible, posttranslational modification that helps target proteins to cellular membranes. The alphavirus small membrane proteins 6K and TF have been reported to be palmitoylated and to positively regulate budding. 6K and TF are isoforms that are identical in their N termini but unique in their C termini due to a −1 ribosomal frameshift during translation. In this study, we used cysteine (Cys) mutants to test differential palmitoylation of the Sindbis virus 6K and TF proteins. We modularly mutated the five Cys residues in the identical N termini of 6K and TF, the four additional Cys residues in TF's unique C terminus, or all nine Cys residues in TF. Using these mutants, we determined that TF palmitoylation occurs primarily in the N terminus. In contrast, 6K is not palmitoylated, even on these shared residues. In the C-terminal Cys mutant, TF protein levels increase both in the cell and in the released virion compared to the wild type. In viruses with the N-terminal Cys residues mutated, TF is much less efficiently localized to the plasma membrane, and it is not incorporated into the virion. The three Cys mutants have minor defects in cell culture growth but a high incidence of abnormal particle morphologies compared to the wild-type virus as determined by transmission electron microscopy. We propose a model where the C terminus of TF modulates the palmitoylation of TF at the N terminus, and palmitoylated TF is preferentially trafficked to the plasma membrane for virus budding. IMPORTANCE Alphaviruses are a reemerging viral cause of arthritogenic disease. Recently, the small 6K and TF proteins of alphaviruses were shown to contribute to virulence in vivo. Nevertheless, a clear understanding of the molecular mechanisms by which either protein acts to promote virus infection is missing. The TF protein is a component of budded virions, and optimal levels of TF correlate positively with wild-type-like particle morphology. In this study, we show that the palmitoylation of TF regulates its localization to the plasma membrane, which is the site of alphavirus budding. Mutants in which TF is not palmitoylated display drastically reduced plasma membrane localization, which effectively prevents TF from participating in budding or being incorporated into virus particles. Investigation of the regulation of TF will aid current efforts in the alphavirus field searching for approaches to mitigate alphaviral disease in humans.

scholarly journals Yeast P4-ATPases Drs2p and Dnf1p Are Essential Cargos of the NPFXD/Sla1p Endocytic Pathway

2007 ◽  
Vol 18 (2) ◽  
pp. 487-500 ◽  
Author(s):  
Ke Liu ◽  
Zhaolin Hua ◽  
Joshua A. Nepute ◽  
Todd R. Graham
Keyword(s):  
Plasma Membrane ◽  
Protein Transport ◽  
Endocytic Pathway ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Sensitive Mutant ◽  
C Terminus ◽  
P Type ◽  
Golgi Network ◽  
Endocytic Pathways

Drs2p family P-type ATPases (P4-ATPases) are required in multiple vesicle-mediated protein transport steps and are proposed to be phospholipid translocases (flippases). The P4-ATPases Drs2p and Dnf1p cycle between the exocytic and endocytic pathways, and here we define endocytosis signals required by these proteins to maintain a steady-state localization to internal organelles. Internalization of Dnf1p from the plasma membrane uses an NPFXD endocytosis signal and its recognition by Sla1p, part of an endocytic coat/adaptor complex with clathrin, Pan1p, Sla2p/End4p, and End3p. Drs2p has multiple endocytosis signals, including two NPFXDs near the C terminus and PEST-like sequences near the N terminus that may mediate ubiquitin (Ub)-dependent endocytosis. Drs2p localizes to the trans-Golgi network in wild-type cells and accumulates on the plasma membrane when both the Ub- and NPFXD-dependent endocytic mechanisms are inactivated. Surprisingly, the pan1-20 temperature-sensitive mutant is constitutively defective for Ub-dependent endocytosis but is not defective for NPFXD-dependent endocytosis at the permissive growth temperature. To sustain viability of pan1-20, Drs2p must be endocytosed through the NPFXD/Sla1p pathway. Thus, Drs2p is an essential endocytic cargo in cells compromised for Ub-dependent endocytosis. These results demonstrate an essential role for endocytosis in retrieving proteins back to the Golgi, and they define critical cargos of the NPFXD/Sla1p system.

scholarly journals The fungal sesquiterpenoid pyrenophoric acid B uses the plant ABA biosynthetic pathway to inhibit seed germination

2019 ◽  
Vol 70 (19) ◽  
pp. 5487-5494 ◽  
Author(s):  
Jorge Lozano-Juste ◽  
Marco Masi ◽  
Alessio Cimmino ◽  
Suzette Clement ◽  
Maria A Fernández ◽  
...  
Keyword(s):  
Seed Germination ◽  
Seedling Establishment ◽  
Biosynthetic Pathway ◽  
Aba Signaling ◽  
Wild Type ◽  
Invasive Weed ◽  
Seed Mortality ◽  
Signaling Components ◽  
Aba Biosynthesis ◽  
Aba Receptors

Abstract Pyrenophoric acid (P-Acid), P-Acid B, and P-Acid C are three phytotoxic sesquiterpenoids produced by the ascomycete seed pathogen Pyrenophora semeniperda, a fungus proposed as a mycoherbicide for biocontrol of cheatgrass, an extremely invasive weed. When tested in cheatgrass bioassays, these metabolites were able to delay seed germination, with P-Acid B being the most active compound. Here, we have investigated the cross-kingdom activity of P-Acid B and its mode of action, and found that it activates the abscisic acid (ABA) signaling pathway in order to inhibit seedling establishment. P-Acid B inhibits seedling establishment in wild-type Arabidopsis thaliana, while several mutants affected in the early perception as well as in downstream ABA signaling components were insensitive to the fungal compound. However, in spite of structural similarities between ABA and P-Acid B, the latter is not able to activate the PYR/PYL family of ABA receptors. Instead, we have found that P-Acid B uses the ABA biosynthesis pathway at the level of alcohol dehydrogenase ABA2 to reduce seedling establishment. We propose that the fungus P. semeniperda manipulates plant ABA biosynthesis as a strategy to reduce seed germination, increasing its ability to cause seed mortality and thereby increase its fitness through higher reproductive success.

scholarly journals O-GlcNAcylation of co-activator-associated arginine methyltransferase 1 regulates its protein substrate specificity

2015 ◽  
Vol 466 (3) ◽  
pp. 587-599 ◽  
Author(s):  
Purin Charoensuksai ◽  
Peter Kuhn ◽  
Lu Wang ◽  
Nathan Sherer ◽  
Wei Xu
Keyword(s):  
Substrate Specificity ◽  
Cellular Localization ◽  
Wild Type ◽  
Post Translational Modification ◽  
Arginine Methyltransferase ◽  
C Terminus ◽  
Arginine Residues ◽  
Quadruple Mutant ◽  
Comprehensive Mapping ◽  
Methyltransferase 1

Co-activator-associated arginine methyltransferase 1 (CARM1) asymmetrically di-methylates proteins on arginine residues. CARM1 was previously known to be modified through O-linked-β-N-acetylglucosaminidation (O-GlcNAcylation). However, the site(s) of O-GlcNAcylation were not mapped and the effects of O-GlcNAcylation on biological functions of CARM1 were undetermined. In the present study, we describe the comprehensive mapping of CARM1 post-translational modification (PTM) using top-down MS. We found that all detectable recombinant CARM1 expressed in human embryonic kidney (HEK293T) cells is automethylated as we previously reported and that about 50% of this automethylated CARM1 contains a single O-linked-β-N-acetylglucosamine (O-GlcNAc) moiety [31]. The O-GlcNAc moiety was mapped by MS to four possible sites (Ser595, Ser598, Thr601 and Thr603) in the C-terminus of CARM1. Mutation of all four sites [CARM1 quadruple mutant (CARM1QM)] markedly decreased O-GlcNAcylation, but did not affect protein stability, dimerization or cellular localization of CARM1. Moreover, CARM1QM elicits similar co-activator activity as CARM1 wild-type (CARM1WT) on a few transcription factors known to be activated by CARM1. However, O-GlcNAc-depleted CARM1 generated by wheat germ agglutinin (WGA) enrichment, O-GlcNAcase (OGA) treatment and mutation of putative O-GlcNAcylation sites displays different substrate specificity from that of CARM1WT. Our findings suggest that O-GlcNAcylation of CARM1 at its C-terminus is an important determinant for CARM1 substrate specificity.

scholarly journals Wild-type and mutant ferroportins do not form oligomers in transfected cells

2006 ◽  
Vol 396 (2) ◽  
pp. 265-275 ◽  
Author(s):  
Ana Sofia Gonçalves ◽  
Françoise Muzeau ◽  
Rand Blaybel ◽  
Gilles Hetet ◽  
Fathi Driss ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Human Kidney ◽  
Cell Types ◽  
Dominant Negative ◽  
Wild Type ◽  
Dominant Form ◽  
Hek 293 ◽  
C Terminus ◽  
293 Cells ◽  
Iron Export

Ferroportin [FPN; Slc40a1 (solute carrier family 40, member 1)] is a transmembrane iron export protein expressed in macrophages and duodenal enterocytes. Heterozygous mutations in the FPN gene result in an autosomal dominant form of iron overload disorder, type-4 haemochromatosis. FPN mutants either have a normal iron export activity but have lost their ability to bind hepcidin, or are defective in their iron export function. The mutant protein has been suggested to act as a dominant negative over the wt (wild-type) protein by multimer formation. Using transiently transfected human epithelial cell lines expressing mouse FPN modified by the addition of a haemagglutinin or c-Myc epitope at the C-terminus, we show that the wtFPN is found at the plasma membrane and in Rab5-containing endosomes, as are the D157G and Q182H mutants. However, the delV162 mutant is mostly intracellular in HK2 cells (human kidney-2 cells) and partially addressed at the cell surface in HEK-293 cells (human embryonic kidney 293 cells). In both cell types, it is partially associated with the endoplasmic reticulum and with Rab5-positive vesicles. However, this mutant is complex-glycosylated like the wt protein. D157G and G323V mutants have a defective iron export capacity as judged by their inability to deplete the intracellular ferritin content, whereas Q182H and delV162 have normal iron export function and probably have lost their capacity to bind hepcidin. In co-transfection experiments, the delV162 mutant does not co-localize with the wtFPN, does not prevent its normal targeting to the plasma membrane and cannot be immunoprecipitated in the same complex, arguing against the formation of FPN hetero-oligomers.

scholarly journals PYL9 is involved in the regulation of ABA signaling during tomato fruit ripening

2019 ◽  
Vol 70 (21) ◽  
pp. 6305-6319 ◽  
Author(s):  
Wenbin Kai ◽  
Juan Wang ◽  
Bin Liang ◽  
Ying Fu ◽  
Yu Zheng ◽  
...  
Keyword(s):  
Fruit Ripening ◽  
Tomato Fruit ◽  
Primary Root ◽  
Dependent Manner ◽  
Aba Signaling ◽  
Wild Type ◽  
Cell Wall Modification ◽  
Altered Expression ◽  
Tomato Fruit Ripening ◽  
Aba Receptors

Abstract Abscisic acid (ABA) regulates fruit ripening, yet little is known about the exact roles of ABA receptors in fruit. In this study, we reveal the role of SlPYL9, a tomato pyrabactin resistance (PYR)/pyrobactin resistance-like (PYL)/regulatory component of ABA receptors (RCAR) protein, as a positive regulator of ABA signaling and fruit ripening. SlPYL9 inhibits protein phosphatase-type 2C (PP2C2/6) in an ABA dose-dependent way, and it interacts physically with SlPP2C2/3/4/5 in an ABA-dependent manner. Expression of SlPYL9 was observed in the seeds, flowers, and fruits. Overexpression and suppression of SlPYL9 induced a variety of phenotypes via altered expression of ABA signaling genes (SlPP2C1/2/9, SlSnRK2.8, SlABF2), thereby affecting expression of ripening-related genes involved in ethylene release and cell wall modification. SlPYL9-OE/RNAi plants showed a typical ABA hyper-/hypo-sensitive phenotype in terms of seed germination, primary root growth, and response to drought. Fruit ripening was significantly accelerated in SlPYL9-OE by 5–7 d as a result of increased endogenous ABA accumulation and advanced release of ethylene compared with the wild-type. In the SlPYL9-RNAi lines, fruit ripening was delayed, mesocarp thickness was enhanced, and petal abscission was delayed compared with the wild-type, resulting in conical/oblong and gourd-shaped fruits. These results suggest that SlPYL9 is involved in ABA signaling, thereby playing a role in the regulation of flower abscission and fruit ripening in tomato.

scholarly journals Arabidopsis LSH8 Positively Regulates ABA Signaling by Changing the Expression Pattern of ABA-Responsive Proteins

2021 ◽  
Vol 22 (19) ◽  
pp. 10314
Author(s):  
Jinpeng Zou ◽  
Zhifang Li ◽  
Haohao Tang ◽  
Li Zhang ◽  
Jingdu Li ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Seed Germination ◽  
Signaling Pathway ◽  
Expression Pattern ◽  
Large Scale ◽  
Aba Signaling ◽  
Wild Type ◽  
Floral Organogenesis ◽  
Early Seedling ◽  
Aba Insensitive

Phytohormone ABA regulates the expression of numerous genes to significantly affect seed dormancy, seed germination and early seedling responses to biotic and abiotic stresses. However, the function of many ABA-responsive genes remains largely unknown. In order to improve the ABA-related signaling network, we conducted a large-scale ABA phenotype screening. LSH, an important transcription factor family, extensively participates in seedling development and floral organogenesis in plants, but whether its family genes are involved in the ABA signaling pathway has not been reported. Here we describe a new function of the transcription factor LSH8 in an ABA signaling pathway. In this study, we found that LSH8 was localized in the nucleus, and the expression level of LSH8 was significantly induced by exogenous ABA at the transcription level and protein level. Meanwhile, seed germination and root length measurements revealed that lsh8 mutant lines were ABA insensitive, whereas LSH8 overexpression lines showed an ABA-hypersensitive phenotype. With further TMT labeling quantitative proteomic analysis, we found that under ABA treatment, ABA-responsive proteins (ARPs) in the lsh8 mutant presented different changing patterns with those in wild-type Col4. Additionally, the number of ARPs contained in the lsh8 mutant was 397, six times the number in wild-type Col4. In addition, qPCR analysis found that under ABA treatment, LSH8 positively mediated the expression of downstream ABA-related genes of ABI3, ABI5, RD29B and RAB18. These results indicate that in Arabidopsis, LSH8 is a novel ABA regulator that could specifically change the expression pattern of APRs to positively mediate ABA responses.

scholarly journals A CACNA1A variant associated with trigeminal neuralgia alters the gating of Cav2.1 channels

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Eder Gambeta ◽  
Maria A. Gandini ◽  
Ivana A. Souza ◽  
Laurent Ferron ◽  
Gerald W. Zamponi
Keyword(s):  
Trigeminal Neuralgia ◽  
Missense Mutation ◽  
Neurotransmitter Release ◽  
Trigeminal System ◽  
Wild Type ◽  
Calcium Dependent ◽  
Whole Cell Patch Clamp ◽  
C Terminus ◽  
Dependent Inactivation

AbstractA novel missense mutation in the CACNA1A gene that encodes the pore forming α1 subunit of the CaV2.1 voltage-gated calcium channel was identified in a patient with trigeminal neuralgia. This mutation leads to a substitution of proline 2455 by histidine (P2455H) in the distal C-terminus region of the channel. Due to the well characterized role of this channel in neurotransmitter release, our aim was to characterize the biophysical properties of the P2455H variant in heterologously expressed CaV2.1 channels. Whole-cell patch clamp recordings of wild type and mutant CaV2.1 channels expressed in tsA-201 cells reveal that the mutation mediates a depolarizing shift in the voltage-dependence of activation and inactivation. Moreover, the P2455H mutant strongly reduced calcium-dependent inactivation of the channel that is consistent with an overall gain of function. Hence, the P2455H CaV2.1 missense mutation alters the gating properties of the channel, suggesting that associated changes in CaV2.1-dependent synaptic communication in the trigeminal system may contribute to the development of trigeminal neuralgia.

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