ABSTRACT
Caloric restriction and intermittent fasting prolong the lifespan and healthspan of model organisms and improve human health. The natural polyamine spermidine has been similarly linked to autophagy enhancement, geroprotection and reduced incidence of cardiovascular and neurodegenerative diseases across species borders. Here, we asked whether the cellular and physiological consequences of caloric restriction and fasting depend on polyamine metabolism. We report that spermidine levels increased upon distinct regimens of fasting or caloric restriction in yeast, flies, mice and human volunteers. Genetic or pharmacological blockade of endogenous spermidine synthesis reduced fasting-induced autophagy in yeast, nematodes and human cells. Furthermore, perturbing the polyamine pathway in vivo abrogated the lifespan- and healthspan-extending effects, as well as the cardioprotective and anti-arthritic consequences of fasting. Mechanistically, spermidine mediated these effects via autophagy induction and hypusination of the translation regulator eIF5A. In summary, the polyamine-hypusination axis emerges as a phylogenetically conserved metabolic control hub for fasting-mediated autophagy enhancement and longevity.
ABSTRACT
In-frame BRAF exon 12 deletions are increasingly identified in various tumor types. The resultant BRAFΔß3-αC oncoproteins usually lack five amino acids in the ß3-αC helix linker and sometimes contain de novo insertions. The dimerization status of BRAFΔß3-αC oncoproteins, their precise pathomechanism, and their direct druggability by RAF inhibitors (RAFi) has been under debate. Here, we functionally characterize BRAFΔLNVTAP>F and two novel mutants, BRAFdelinsFS and BRAFΔLNVT>F, and compare them with other BRAFΔß3-αC oncoproteins. We show that BRAFΔß3-αC oncoproteins not only form stable homodimers and large multiprotein complexes but also require dimerization. Nevertheless, details matter as aromatic amino acids at the deletion junction of some BRAFΔß3-αC oncoproteins, e.g., BRAFΔLNVTAP>F, increase their stability and dimerization propensity while conferring resistance to monomer-favoring RAFi such as dabrafenib or HSP 90/CDC37 inhibition. In contrast, dimer-favoring inhibitors such as naporafenib inhibit all BRAFΔß3-αC mutants in cell lines and patient-derived organoids, suggesting that tumors driven by such oncoproteins are vulnerable to these compounds.
Subject(s)
HSP90 Heat-Shock Proteins , Proto-Oncogene Proteins B-raf , Humans , Dimerization , Proto-Oncogene Proteins B-raf/genetics , Amino AcidsABSTRACT
Autophagy serves as a defense mechanism against intracellular pathogens, but several microorganisms exploit it for their own benefit. Accordingly, certain herpesviruses include autophagic membranes into their infectious virus particles. In this study, we analyzed the composition of purified virions of the Epstein-Barr virus (EBV), a common oncogenic γ-herpesvirus. In these, we found several components of the autophagy machinery, including membrane-associated LC3B-II, and numerous viral proteins, such as the capsid assembly proteins BVRF2 and BdRF1. Additionally, we showed that BVRF2 and BdRF1 interact with LC3B-II via their common protein domain. Using an EBV mutant, we identified BVRF2 as essential to assemble mature capsids and produce infectious EBV. However, BdRF1 was sufficient for the release of noninfectious viral envelopes as long as autophagy was not compromised. These data suggest that BVRF2 and BdRF1 are not only important for capsid assembly but together with the LC3B conjugation complex of ATG5-ATG12-ATG15L1 are also critical for EBV envelope release.
Subject(s)
Capsid , Epstein-Barr Virus Infections , Humans , Capsid/metabolism , Herpesvirus 4, Human/genetics , Herpesvirus 4, Human/metabolism , Viral Envelope/metabolism , Epstein-Barr Virus Infections/metabolism , Capsid Proteins/genetics , Capsid Proteins/metabolismABSTRACT
Hyperosmotic stress occurs in several diseases, but its long-term effects are largely unknown. We used sorbitol-treated human fibroblasts in 3D culture to study the consequences of hyperosmotic stress in the skin. Sorbitol regulated many genes, which help cells cope with the stress condition. The most robustly regulated gene encodes serine protease 35 (PRSS35). Its regulation by hyperosmotic stress was dependent on the kinases p38 and JNK and the transcription factors NFAT5 and ATF2. We identified different collagens and collagen-associated proteins as putative PRSS35 binding partners. This is functionally important because PRSS35 affected the extracellular matrix proteome, which limited cell proliferation. The in vivo relevance of these findings is reflected by the coexpression of PRSS35 and its binding partners in human skin wounds, where hyperosmotic stress occurs as a consequence of excessive water loss. These results identify PRSS35 as a key regulator of the matrisome under hyperosmotic stress conditions.
Subject(s)
Extracellular Matrix , Fibroblasts , Humans , Endopeptidases , Sorbitol , Serine ProteasesABSTRACT
The mitophagic degradation of mitochondrial matrix proteins in Saccharomyces cerevisiae was previously shown to be selective, reflecting a pre-engulfment sorting step within the mitochondrial network. This selectivity is regulated through phosphorylation of mitochondrial matrix proteins by the matrix kinases Pkp1 and Pkp2, which in turn appear to be regulated by the phosphatase Aup1/Ptc6. However, these same proteins also regulate the phosphorylation status and catalytic activity of the yeast pyruvate dehydrogenase complex, which is critical for mitochondrial metabolism. To understand the relationship between these two functions, we evaluated the role of the pyruvate dehydrogenase complex in mitophagic selectivity. Surprisingly, we identified a novel function of the complex in regulating mitophagic selectivity, which is independent of its enzymatic activity. Our data support a model in which the pyruvate dehydrogenase complex directly regulates the activity of its associated kinases and phosphatases. This regulatory interaction then determines the phosphorylation state of mitochondrial matrix proteins and their mitophagic fates.
Subject(s)
Pyruvate Dehydrogenase Complex , Saccharomyces cerevisiae Proteins , Phosphorylation , Pyruvate Dehydrogenase Complex/metabolism , Saccharomyces cerevisiae/metabolism , Mitochondria/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Mitochondrial Proteins/metabolismABSTRACT
The exporter of the auxin precursor indole-3-butyric acid (IBA), ABCG36/PDR8/PEN3, from the model plant Arabidopsis has recently been proposed to also function in the transport of the phytoalexin camalexin. Based on these bonafide substrates, it has been suggested that ABCG36 functions at the interface between growth and defense. Here, we provide evidence that ABCG36 catalyzes the direct, ATP-dependent export of camalexin across the plasma membrane. We identify the leucine-rich repeat receptor kinase, QIAN SHOU KINASE1 (QSK1), as a functional kinase that physically interacts with and phosphorylates ABCG36. Phosphorylation of ABCG36 by QSK1 unilaterally represses IBA export, allowing camalexin export by ABCG36 conferring pathogen resistance. As a consequence, phospho-dead mutants of ABCG36, as well as qsk1 and abcg36 alleles, are hypersensitive to infection with the root pathogen Fusarium oxysporum, caused by elevated fungal progression. Our findings indicate a direct regulatory circuit between a receptor kinase and an ABC transporter that functions to control transporter substrate preference during plant growth and defense balance decisions.
Subject(s)
Arabidopsis Proteins , Arabidopsis , ATP-Binding Cassette Transporters/metabolism , Arabidopsis/metabolism , Thiazoles/metabolism , Phytoalexins , Arabidopsis Proteins/metabolism , Plant Diseases/microbiology , Gene Expression Regulation, PlantABSTRACT
Geobacter sulfurreducens is a widely applied microorganism for the reduction of toxic metal salts, as an electron source for bioelectrochemical devices, and as a reagent for the synthesis of nanoparticles. In order to understand the influence of metal salts, and of electron transporting, multiheme c-cytochromes on the electron flux during respiration of G. sulfurreducens, the reduction kinetic of Fe3+, Co3+, V5+, Cr6+, and Mn7+ containing complexes were measured. Starting from the resting phase, each G. sulfurreducens cell produced an electron flux of 3.7 × 105 electrons per second during the respiration process. Reduction rates were within ± 30% the same for the 6 different metal salts, and reaction kinetics were of zero order. Decrease of c-cytochrome concentrations by downregulation and mutation demonstrated that c-cytochromes stabilized respiration rates by variation of their redox states. Increasing Fe2+/heme levels increased electron flux rates, and induced respiration flexibility. The kinetic effects parallel electrochemical results of G. sulfurreducens biofilms on electrodes, and might help to optimize bioelectrochemical devices.
ABSTRACT
Spermatogenesis is a highly specialized differentiation process driven by a dynamic gene expression program and ending with the production of mature spermatozoa. Whereas hundreds of genes are known to be essential for male germline proliferation and differentiation, the contribution of several genes remains uncharacterized. The predominant expression of the latest globin family member, androglobin (Adgb), in mammalian testis tissue prompted us to assess its physiological function in spermatogenesis. Adgb knockout mice display male infertility, reduced testis weight, impaired maturation of elongating spermatids, abnormal sperm shape, and ultrastructural defects in microtubule and mitochondrial organization. Epididymal sperm from Adgb knockout animals display multiple flagellar malformations including coiled, bifid or shortened flagella, and erratic acrosomal development. Following immunoprecipitation and mass spectrometry, we could identify septin 10 (Sept10) as interactor of Adgb. The Sept10-Adgb interaction was confirmed both in vivo using testis lysates and in vitro by reciprocal co-immunoprecipitation experiments. Furthermore, the absence of Adgb leads to mislocalization of Sept10 in sperm, indicating defective manchette and sperm annulus formation. Finally, in vitro data suggest that Adgb contributes to Sept10 proteolysis in a calmodulin-dependent manner. Collectively, our results provide evidence that Adgb is essential for murine spermatogenesis and further suggest that Adgb is required for sperm head shaping via the manchette and proper flagellum formation.
Subject(s)
Globins , Infertility, Male , Animals , Fertility , Globins/metabolism , Infertility, Male/genetics , Male , Mammals , Mice , Mice, Knockout , Semen , Sperm Tail , Spermatids/metabolism , Spermatozoa , Testis/metabolismABSTRACT
Interactions between macrophages, cardiac cells and the extracellular matrix are crucial for cardiac repair following myocardial infarction (MI). We hypothesized that cell-based treatments might modulate these interactions. After validating that bone marrow cells (BMC) associated with fibrin lowered the infarct extent and improved cardiac function, we interrogated the influence of fibrin, as a biologically active scaffold, on the secretome of BMC and the impact of their association on macrophage fate and cardiomyoblast proliferation. In vitro, BMC were primed with fibrin (F-BMC). RT-PCR and proteomic analyses showed that fibrin profoundly influenced the gene expression and the secretome of BMCs. Consequently, the secretome of F-BMC increased the spreading of cardiomyoblasts and showed an alleviated immunomodulatory capacity. Indeed, the proliferation of anti-inflammatory macrophages was augmented, and the phenotype of pro-inflammatory switched as shown by downregulated Nos2, Il6 and IL1b and upregulated Arg1, CD163, Tgfb and IL10. Interestingly, the secretome of F-BMC educated-macrophages stimulated the incorporation of EdU in cardiomyoblasts. In conclusion, our study provides evidence that BMC/fibrin-based treatment improved cardiac structure and function following MI. In vitro proofs-of-concept reveal that the F-BMC secretome increases cardiac cell size and promotes an anti-inflammatory response. Thenceforward, the F-BMC educated macrophages sequentially stimulated cardiac cell proliferation.
ABSTRACT
The biogenesis of eukaryotic ribosomes involves the ordered assembly of around 80 ribosomal proteins. Supplying equimolar amounts of assembly-competent ribosomal proteins is complicated by their aggregation propensity and the spatial separation of their location of synthesis and pre-ribosome incorporation. Recent evidence has highlighted that dedicated chaperones protect individual, unassembled ribosomal proteins on their path to the pre-ribosomal assembly site. Here, we show that the co-translational recognition of Rpl3 and Rpl4 by their respective dedicated chaperone, Rrb1 or Acl4, reduces the degradation of the encoding RPL3 and RPL4 mRNAs in the yeast Saccharomyces cerevisiae. In both cases, negative regulation of mRNA levels occurs when the availability of the dedicated chaperone is limited and the nascent ribosomal protein is instead accessible to a regulatory machinery consisting of the nascent-polypeptide-associated complex and the Caf130-associated Ccr4-Not complex. Notably, deregulated expression of Rpl3 and Rpl4 leads to their massive aggregation and a perturbation of overall proteostasis in cells lacking the E3 ubiquitin ligase Tom1. Taken together, we have uncovered an unprecedented regulatory mechanism that adjusts the de novo synthesis of Rpl3 and Rpl4 to their actual consumption during ribosome assembly and, thereby, protects cells from the potentially detrimental effects of their surplus production.
Living cells are packed full of molecules known as proteins, which perform many vital tasks the cells need to survive and grow. Machines called ribosomes inside the cells use template molecules called messenger RNAs (or mRNAs for short) to produce proteins. The newly-made proteins then have to travel to a specific location in the cell to perform their tasks. Some newly-made proteins are prone to forming clumps, so cells have other proteins known as chaperones that ensure these clumps do not form. The ribosomes themselves are made up of several proteins, some of which are also prone to clumping as they are being produced. To prevent this from happening, cells control how many ribosomal proteins they make, so there are just enough to form the ribosomes the cell needs at any given time. Previous studies found that, in yeast, two ribosomal proteins called Rpl3 and Rpl4 each have their own dedicated chaperone to prevent them from clumping. However, it remained unclear whether these chaperones are also involved in regulating the levels of Rpl3 and Rpl4. To address this question, Pillet et al. studied both of these dedicated chaperones in yeast cells. The experiments showed that the chaperones bound to their target proteins (either units of Rpl3 or Rpl4) as they were being produced on the ribosomes. This protected the template mRNAs the ribosomes were using to produce these proteins from being destroyed, thus allowing further units of Rpl3 and Rpl4 to be produced. When enough Rpl3 and Rpl4 units were made, there were not enough of the chaperones to bind them all, leaving the mRNA templates unprotected. This led to the destruction of the mRNA templates, which decreased the numbers of Rpl3 and Rpl4 units being produced. The work of Pillet et al. reveals a feedback mechanism that allows yeast to tightly control the levels of Rpl3 and Rpl4. In the future, these findings may help us understand diseases caused by defects in ribosomal proteins, such as Diamond-Blackfan anemia, and possibly also neurodegenerative diseases caused by clumps of proteins forming in cells. The next step will be to find out whether the mechanism uncovered by Pillet et al. also exists in human and other mammalian cells.
Subject(s)
Ribosomal Proteins , Saccharomyces cerevisiae Proteins , Proteostasis , Ribosomal Proteins/genetics , Ribosomal Proteins/metabolism , Ribosomes/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolismABSTRACT
Mitogen-activated protein kinase (MAPK) cascades transmit environmental signals and induce stress and defence responses in plants. These signalling cascades are negatively controlled by specific Ser/Thr protein phosphatases of the type 2C (PP2C) and dual-specificity phosphatase (DSP) families that inactivate stress-induced MAPKs; however, the interplay between phosphatases of these different types has remained unknown. This work reveals that different Arabidopsis MAPK phosphatases, the PP2C-type AP2C1 and the DSP-type MKP1, exhibit both specific and overlapping functions in plant stress responses. Each single mutant, ap2c1 and mkp1, and the ap2c1 mkp1 double mutant displayed enhanced stress-induced activation of the MAPKs MPK3, MPK4, and MPK6, as well as induction of a set of transcription factors. Moreover, ap2c1 mkp1 double mutants showed an autoimmune-like response, associated with increased levels of the stress hormones salicylic acid and ethylene, and of the phytoalexin camalexin. This phenotype was reduced in the ap2c1 mkp1 mpk3 and ap2c1 mkp1 mpk6 triple mutants, suggesting that the autoimmune-like response is due to MAPK misregulation. We conclude that the evolutionarily distant MAPK phosphatases AP2C1 and MKP1 contribute crucially to the tight control of MAPK activities, ensuring appropriately balanced stress signalling and suppression of autoimmune-like responses during plant growth and development.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Humans , Mitogen-Activated Protein Kinases/genetics , Mitogen-Activated Protein Kinases/metabolism , Phosphoprotein Phosphatases/genetics , Phosphoprotein Phosphatases/metabolism , Phosphorylation , Protein Tyrosine Phosphatases/genetics , Protein Tyrosine Phosphatases/metabolismABSTRACT
The eukaryotic TORC1 kinase assimilates diverse environmental cues, including growth factors and nutrients, to control growth by tuning anabolic and catabolic processes. In yeast, TORC1 stimulates protein synthesis in response to abundant nutrients primarily through its proximal effector kinase Sch9. Conversely, TORC1 inhibition following nutrient limitation unlocks various distally controlled kinases (e.g., Atg1, Gcn2, Npr1, Rim15, Slt2/Mpk1, and Yak1), which cooperate through poorly defined circuits to orchestrate the quiescence program. To better define the signaling landscape of the latter kinases, we use in vivo quantitative phosphoproteomics. Through pinpointing known and uncharted Npr1, Rim15, Slt2/Mpk1, and Yak1 effectors, our study examines the architecture of the distally controlled TORC1 kinase network. Accordingly, this is built on a combination of discrete, convergent, and multilayered feedback regulatory mechanisms, which likely ensure homeostatic control of and/or robust responses by TORC1 and its effector kinases under fluctuating nutritional conditions.
Subject(s)
Gene Expression Regulation, Fungal/drug effects , Mechanistic Target of Rapamycin Complex 1/pharmacology , Phosphoproteins/metabolism , Protein Kinases/chemistry , Proteome/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Proteome/analysis , Saccharomyces cerevisiae/drug effectsABSTRACT
In cooperative systems exhibiting division of labor, such as microbial communities, multicellular organisms, and social insect colonies, individual units share costs and benefits through both task specialization and exchanged materials. Socially exchanged fluids, like seminal fluid and milk, allow individuals to molecularly influence conspecifics. Many social insects have a social circulatory system, where food and endogenously produced molecules are transferred mouth-to-mouth (stomodeal trophallaxis), connecting all the individuals in the society. To understand how these endogenous molecules relate to colony life, we used quantitative proteomics to investigate the trophallactic fluid within colonies of the carpenter ant Camponotus floridanus. We show that different stages of the colony life cycle circulate different types of proteins: young colonies prioritize direct carbohydrate processing; mature colonies prioritize accumulation and transmission of stored resources. Further, colonies circulate proteins implicated in oxidative stress, ageing, and social insect caste determination, potentially acting as superorganismal hormones. Brood-caring individuals that are also closer to the queen in the social network (nurses) showed higher abundance of oxidative stress-related proteins. Thus, trophallaxis behavior could provide a mechanism for distributed metabolism in social insect societies. The ability to thoroughly analyze the materials exchanged between cooperative units makes social insect colonies useful models to understand the evolution and consequences of metabolic division of labor at other scales.
Division of labor is essential for cooperation, because groups can achieve more when individuals specialize in different tasks. This happens across the natural world, from different cells in organisms performing specific roles, to the individuals in an ant colony carrying out diverse duties. In both of these systems, individuals work together to ensure the survival of the collective unit the body or the colony instead of competing against each other. One of the main ways division of labor is evident within these two systems is regarding reproduction. Both in the body and in an ant colony, only one or a few individual units can reproduce, while the rest provide support. In the case of ant colonies, only queens and males reproduce, while the young workers nurse the brood and older workers forage for food. This intense cooperation requires close communication between individual units in the case of some species of ants, by sharing fluids mouth-to-mouth. These fluids contain food but also many molecules produced by the ants themselves, including proteins. Given that both individuals and the colony as a whole change as they age with workers acquiring new roles, and new queens and males only reared once the colony is mature it is likely that the proteins transmitted in the fluid also change. To better understand whether the lifecycles of individuals and the age of the colony affect the fluids shared by carpenter ants Camponotus floridanus, Hakala et al. examined the ant-produced proteins in these fluids. This revealed differences in the proteins shared by young and mature colonies, and young nurse ants and older forager ants. In young colonies, the fluids contained proteins involved in fast sugar processing; while in mature colonies, the fluids contained more proteins to store nutrients, which help insect larvae grow into larger individuals, like queens. Young worker ants, who spend their time nursing the brood, produced more anti-aging proteins. This may be because these ants are in close contact with the queen, who lives much longer than the rest of the ants in the colony. Taken together, these observations suggest that ants divide the labor of metabolism, as well as work and reproduction. Dividing the labor of metabolism among individuals is one more similarity between ants and the cells of a multicellular organism, like a fly or a human. Division of labor allows the sharing of burden, with some individuals lightening the load of others. Understanding how ants achieve this by sharing fluids could shed new light on this complex exchange at other scales or in other organisms. By matching proteins to life stages, researchers have a starting point to examine individual molecules in more detail.
Subject(s)
Ants/physiology , Biomarkers/metabolism , Animals , Ants/metabolism , Social BehaviorABSTRACT
The evolutionarily conserved ULK1 kinase complex acts as gatekeeper of canonical autophagy and regulates induction of autophagosome biogenesis. To better understand control of ULK1 and analyze whether ULK1 has broader functions that are also linked to the later steps of autophagy, we perform comprehensive phosphoproteomic analyses. Combining in vivo with in vitro data, we identify numerous direct ULK1 target sites within autophagy-relevant proteins that are critical for autophagosome maturation and turnover. In addition, we highlight an intimate crosstalk between ULK1 and several phosphatase complexes. ULK1 is not only a PP2A target but also directly phosphorylates the regulatory PP2A subunit striatin, activating PP2A and serving as positive feedback to promote autophagy-dependent protein turnover. Thus, ULK1 and phosphatase activities are tightly coordinated to robustly regulate protein degradation by autophagy.
Subject(s)
Autophagy-Related Protein-1 Homolog/metabolism , Autophagy/physiology , Calmodulin-Binding Proteins/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Membrane Proteins/metabolism , Nerve Tissue Proteins/metabolism , Protein Phosphatase 2/metabolism , Autophagosomes/metabolism , Autophagy-Related Proteins/metabolism , Humans , Phosphorylation , Protein Processing, Post-Translational/physiologyABSTRACT
Recessive dystrophic epidermolysis bullosa (RDEB), a genetic skin blistering disease, is a paradigmatic condition of tissue fragility-driven multi-organ fibrosis. Here, longitudinal analyses of the tissue proteome through the course of naturally developing disease in RDEB mice revealed that increased pro-inflammatory immunity associates with fibrosis evolution. Mechanistically, this fibrosis is a consequence of altered extracellular matrix organization rather than that of increased abundance of major structural proteins. In a humanized system of disease progression, we targeted inflammatory cell fibroblast communication with Ang-(1-7)-an anti-inflammatory heptapeptide of the renin-angiotensin system, which reduced the fibrosis-evoking aptitude of RDEB cells. In vivo, systemic administration of Ang-(1-7) efficiently attenuated progression of multi-organ fibrosis and increased survival of RDEB mice. Collectively, our study shows that selective down-modulation of pro-inflammatory immunity may mitigate injury-induced fibrosis. Furthermore, together with published data, our data highlight molecular diversity among fibrotic conditions. Both findings have direct implications for the design of therapies addressing skin fragility and fibrosis.
Subject(s)
Epidermolysis Bullosa Dystrophica , Animals , Collagen Type VII , Epidermolysis Bullosa Dystrophica/pathology , Fibroblasts/pathology , Fibrosis , MiceABSTRACT
The circadian clock regulates many biochemical and physiological pathways, and lack of clock genes, such as Period (Per) 2, affects not only circadian activity rhythms, but can also modulate feeding and mood-related behaviors. However, it is not known how cell-type specific expression of Per2 contributes to these behaviors. In this study, we find that Per2 in glial cells is important for balancing mood-related behaviors, without affecting circadian activity parameters. Genetic and adeno-associated virus-mediated deletion of Per2 in glial cells of mice leads to reduced despair and anxiety. This is paralleled by an increase of the GABA transporter 2 (Gat2/Slc6a13) and Dopamine receptor D3 (Drd3) mRNA, and a reduction of glutamate levels in the nucleus accumbens (NAc). Interestingly, neuronal Per2 knock-out also reduces despair, but does not influence anxiety. The change in mood-related behavior is not a result of a defective molecular clock, as glial Bmal1 deletion has no effect on neither despair nor anxiety. Exclusive deletion of Per2 in glia of the NAc reduced despair, but had no influence on anxiety. Our data provide strong evidence for an important role of glial Per2 in regulating mood-related behavior.
Subject(s)
Affect , Behavior, Animal , Neuroglia/metabolism , Period Circadian Proteins/genetics , Sequence Deletion , Animals , Astrocytes/metabolism , Breeding , Circadian Rhythm , Dependovirus/genetics , Gene Expression , Genetic Association Studies , Genetic Vectors/genetics , Mice , Phenotype , Transduction, GeneticABSTRACT
Most land plants entertain a mutualistic symbiosis known as arbuscular mycorrhiza with fungi (Glomeromycota) that provide them with essential mineral nutrients, in particular phosphate (Pi), and protect them from biotic and abiotic stress. Arbuscular mycorrhizal (AM) symbiosis increases plant productivity and biodiversity and is therefore relevant for both natural plant communities and crop production. However, AM fungal populations suffer from intense farming practices in agricultural soils, in particular Pi fertilization. The dilemma between natural fertilization from AM symbiosis and chemical fertilization has raised major concern and emphasizes the need to better understand the mechanisms by which Pi suppresses AM symbiosis. Here, we test the hypothesis that Pi may interfere with AM symbiosis via the phytohormone gibberellic acid (GA) in the Solanaceous model systems Petunia hybrida and Nicotiana tabacum. Indeed, we find that GA is inhibitory to AM symbiosis and that Pi may cause GA levels to increase in mycorrhizal roots. Consistent with a role of endogenous GA as an inhibitor of AM development, GA-defective N. tabacum lines expressing a GA-metabolizing enzyme (GA methyltransferase-GAMT) are colonized more quickly by the AM fungus Rhizoglomus irregulare, and exogenous Pi is less effective in inhibiting AM colonization in these lines. Systematic gene expression analysis of GA-related genes reveals a complex picture, in which GA degradation by GA2 oxidase plays a prominent role. These findings reveal potential targets for crop breeding that could reduce Pi suppression of AM symbiosis, thereby reconciling the advantages of Pi fertilization with the diverse benefits of AM symbiosis.
Subject(s)
Gibberellins/metabolism , Mycorrhizae/physiology , Nicotiana/physiology , Petunia/physiology , Phosphates/metabolism , Gene Expression Regulation, Plant , Plant Growth Regulators/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Plant Roots/genetics , Plant Roots/microbiology , Plants, Genetically Modified , Signal Transduction , SymbiosisABSTRACT
Daily recurring events can be predicted by animals based on their internal circadian timing system. However, independently from the suprachiasmatic nuclei (SCN), the central pacemaker of the circadian system in mammals, restriction of food access to a particular time of day elicits food anticipatory activity (FAA). This suggests an involvement of other central and/or peripheral clocks as well as metabolic signals in this behavior. One of the metabolic signals that is important for FAA under combined caloric and temporal food restriction is ß-hydroxybutyrate (ßOHB). Here we show that the monocarboxylate transporter 1 (Mct1), which transports ketone bodies such as ßOHB across membranes of various cell types, is involved in FAA. In particular, we show that lack of the Mct1 gene in the liver, but not in neuronal or glial cells, reduces FAA in mice. This is associated with a reduction of ßOHB levels in the blood. Our observations suggest an important role of ketone bodies and its transporter Mct1 in FAA under caloric and temporal food restriction.
ABSTRACT
Indole-3-acetic acid (IAA) is the most common, naturally occurring phytohormone that regulates cell division, differentiation, and senescence in plants. The capacity to synthesize IAA is also widespread among plant-associated bacterial and fungal species, which may use IAA as an effector molecule to define their relationships with plants or to coordinate their physiological behavior through cell-cell communication. Fungi, including many species that do not entertain a plant-associated life style, are also able to synthesize IAA, but the physiological role of IAA in these fungi has largely remained enigmatic. Interestingly, in this context, growth of the budding yeast Saccharomyces cerevisiae is sensitive to extracellular IAA. Here, we use a combination of various genetic approaches including chemical-genetic profiling, SAturated Transposon Analysis in Yeast (SATAY), and genetic epistasis analyses to identify the mode-of-action by which IAA inhibits growth in yeast. Surprisingly, these analyses pinpointed the target of rapamycin complex 1 (TORC1), a central regulator of eukaryotic cell growth, as the major growth-limiting target of IAA. Our biochemical analyses further demonstrate that IAA inhibits TORC1 both in vivo and in vitro. Intriguingly, we also show that yeast cells are able to synthesize IAA and specifically accumulate IAA upon entry into stationary phase. Our data therefore suggest that IAA contributes to proper entry of yeast cells into a quiescent state by acting as a metabolic inhibitor of TORC1.