Development, Establishment, and Validation of a Model for the Mineralization of Periodontium Remodelling Cells: Cementoblasts.

Chronic kidney disease (CKD) patients undergoing dialysis are at high risk of bone fractures. CKD-induced mineral and bone disorder is extended to periodontal disease due to changes in the ionic composition of saliva in CKD patients, dysregulating mineralization, hindering regeneration and thereby promoting the progression of dental complications. Despite the importance of cementum for overall oral health, the mechanisms that regulate its development and regeneration are not well comprehended, and a lack of sufficient in vitro experimental models has hindered research progress. In this study, the impact of experimental conditions on the calcification of cementoblasts was systematically investigated, aimed at establishing a standardized and validated model for the calcification of cementoblasts. The effects of phosphate, calcium, ascorbic acid, ß-glycerolphosphate, dexamethasone, and fetal calf serum on the calcification process of cementoblasts were analyzed over a wide range of concentrations and time points by investigating calcium content, cell viability, gene expression and kinase activity. Cementoblasts calcified in a concentration- and time-dependent manner with higher concentrations of supplements cause a higher degree of calcification but decreased cell viability. Phosphate and calcium have a significantly stronger effect on cementoblast calcification processes compared to osteogenic supplements: ascorbic acid, ß-glycerolphosphate, and dexamethasone induce calcification over a wide range of osteogenic signalling pathways, with osteopontin being a central target of gene regulation. Conversely, treatment with ascorbic acid, ß-glycerolphosphate, and dexamethasone leads to activating only selected pathways, especially promoting bone sialoprotein expression. The developed and validated cementoblast calcification protocol, incubating up to 60% confluent cementoblasts with 1.9 mmol L-1 of phosphate supplementation for a reasonable, multi-pathway calcification induction and 10 mmol L-1 ß-glycerolphosphate, 75 µmol L-1 ascorbic acid and 10 nmol L-1 dexamethasone for a reasonable osteogenic differentiation-based calcification induction, provides standard in vitro experimental models for better understanding cementoblast function and regeneration.

Functional requirement of the Arabidopsis importin-α nuclear transport receptor family in autoimmunity mediated by the NLR protein SNC1.

IMPORTIN-α3/MOS6 (MODIFIER OF SNC1, 6) is one of nine importin-α isoforms in Arabidopsis that recruit nuclear localization signal-containing cargo proteins to the nuclear import machinery. IMP-α3/MOS6 is required genetically for full autoimmunity of the nucleotide-binding leucine-rich repeat immune receptor mutant snc1 (suppressor of npr1-1, constitutive 1) and MOS6 also contributes to basal disease resistance. Here, we investigated the contribution of the other importin-α genes to both types of immune responses, and we analyzed potential interactions of all importin-α isoforms with SNC1. By using reverse-genetic analyses in Arabidopsis and protein-protein interaction assays in Nicotiana benthamiana, we provide evidence that among the nine α-importins in Arabidopsis, IMP-α3/MOS6 is the main nuclear transport receptor of SNC1, and that IMP-α3/MOS6 is required selectively for autoimmunity of snc1 and basal resistance to mildly virulent Pseudomonas syringae in Arabidopsis.

Mechanical Compression by Simulating Orthodontic Tooth Movement in an In Vitro Model Modulates Phosphorylation of AKT and MAPKs via TLR4 in Human Periodontal Ligament Cells.

Mechanical compression simulating orthodontic tooth movement in in vitro models induces pro-inflammatory cytokine expression in periodontal ligament (PDL) cells. Our previous work shows that TLR4 is involved in this process. Here, primary PDL cells are isolated and characterized to better understand the cell signaling downstream of key molecules involved in the process of sterile inflammation via TLR4. The TLR4 monoclonal blocking antibody significantly reverses the upregulation of phospho-AKT, caused by compressive force, to levels comparable to controls by inhibition of TLR4. Phospho-ERK and phospho-p38 are also modulated in the short term via TLR4. Additionally, moderate compressive forces of 2 g/cm2, a gold standard for static compressive mechanical stimulation, are not able to induce translocation of Nf-kB and phospho-ERK into the nucleus. Accordingly, we demonstrated for the first time that TLR4 is also one of the triggers for signal transduction under compressive force. The TLR4, one of the pattern recognition receptors, is involved through its specific molecular structures on damaged cells during mechanical stress. Our findings provide the basis for further research on TLR4 in the modulation of sterile inflammation during orthodontic therapy and periodontal remodeling.

The truncated NLR protein TIR-NBS13 is a MOS6/IMPORTIN-α3 interaction partner required for plant immunity.

Importin-α proteins mediate the translocation of nuclear localization signal (NLS)-containing proteins from the cytoplasm into the nucleus through nuclear pore complexes (NPCs). Genetically, Arabidopsis IMPORTIN-α3/MOS6 (MODIFIER OF SNC1, 6) is required for basal plant immunity and constitutive disease resistance activated in autoimmune mutant snc1 (suppressor of npr1-1, constitutive 1), suggesting that MOS6 plays a role in the nuclear import of proteins involved in plant defense signaling. Here, we sought to identify and characterize defense-regulatory cargo proteins and interaction partners of MOS6. We conducted both in silico database analyses and affinity purification of functional epitope-tagged MOS6 from pathogen-challenged stable transgenic plants coupled with mass spectrometry. We show that among the 13 candidate MOS6 interactors we selected for further functional characterization, the TIR-NBS-type protein TN13 is required for resistance against Pseudomonas syringae pv. tomato (Pst) DC3000 lacking the type-III effector proteins AvrPto and AvrPtoB. When expressed transiently in N. benthamiana leaves, TN13 co-immunoprecipitates with MOS6, but not with its closest homolog IMPORTIN-α6, and localizes to the endoplasmic reticulum (ER), consistent with a predicted N-terminal transmembrane domain in TN13. Our work uncovered the truncated NLR protein TN13 as a component of plant innate immunity that selectively binds to MOS6/IMPORTIN-α3 in planta. We speculate that the release of TN13 from the ER membrane in response to pathogen stimulus, and its subsequent nuclear translocation, is important for plant defense signal transduction.

The putative kinase substrate MUSE7 negatively impacts the accumulation of NLR proteins.

Stringent modulation of immune signaling in plants is necessary to enable a rapid response to pathogen attack without spurious defense activation. To identify genes involved in plant immunity, a forward genetic screen for enhancers of the autoimmune snc1 (suppressor of npr1, constitutive 1) mutant was conducted. The snc1 mutant contains a gain-of-function mutation in a gene encoding a NOD-like receptor (NLR) protein. The isolated muse7 (mutant, snc1-enhancing, 7) mutant was shown to confer a reversion to autoimmune phenotypes in the wild-type-like mos4 (modifier of snc1, 4) snc1 background. Positional cloning revealed that MUSE7 encodes an evolutionarily conserved putative kinase substrate of unknown function. The muse7 single mutants display enhanced resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000. While transcription of SNC1 is not enhanced, elevated SNC1 protein accumulation is associated with mutations in muse7. Accumulation of two additional NLR proteins, RPS2 (RESISTANCE TO PSEUDOMONAS SYRINGAE 2) and RPM1 (RESISTANCE TO PSEUDOMONAS SYRINGAE pv. MACULICOLA 1), was also observed in muse7 plants. Although proteasome-mediated degradation of NLR proteins is a well studied event in plant immunity, no interactions were detected between MUSE7 and selected components of this pathway. This study has demonstrated a role for MUSE7 in modulating plant immune responses through negatively affecting NLR accumulation, and will benefit future studies of MUSE7 homologs in other species.

Nucleoporin-Regulated MAP Kinase Signaling in Immunity to a Necrotrophic Fungal Pathogen.

Pathogen-responsive mitogen-activated protein kinase (MAPK or MPK) cascades relay signals from activated immune receptors across the nuclear envelope to intranuclear targets. However, in plants, little is known about the spatial control of MAPK signaling. Here, we report that the Arabidopsis (Arabidopsis thaliana) nuclear pore complex protein Nup88/MOS7 is essential for immunity to the necrotrophic fungus Botrytis cinerea The mos7-1 mutation, causing a four-amino acid deletion, compromises B. cinerea-induced activation of the key immunoregulatory MAPKs MPK3/MPK6 and reduces MPK3 protein levels posttranscriptionally. Furthermore, MOS7 contributes to retaining a sufficient MPK3 abundance in the nucleus, which is required for full immunity to B. cinerea Finally, we present a structural model of MOS7 and show that the mos7-1 mutation compromises interactions with Nup98a/b, two phenylalanine-glycine repeat nucleoporins implicated in maintaining the selective nuclear pore complex permeability barrier. Together, our analysis uncovered MOS7 and Nup98 as novel components of plant immunity toward a necrotrophic pathogen and provides mechanistic insights into how these nucleoporins coordinate nucleocytoplasmic transport to mount a robust immune response.

An E4 ligase facilitates polyubiquitination of plant immune receptor resistance proteins in Arabidopsis.

Proteins with nucleotide binding and leucine-rich repeat domains (NLRs) serve as immune receptors in animals and plants that recognize pathogens and activate downstream defense responses. As high accumulation of NLRs can result in unwarranted autoimmune responses, their cellular concentrations must be tightly regulated. However, the molecular mechanisms of this process are poorly detailed. The F-box protein Constitutive expressor of PR genes 1 (CPR1) was previously identified as a component of a Skp1, Cullin1, F-box protein E3 complex that targets NLRs, including Suppressor of NPR1, Constitutive 1 (SNC1) and Resistance to Pseudomonas syringae 2 (RPS2), for ubiquitination and further protein degradation. From a forward genetic screen, we identified Mutant, snc1-enhancing 3 (MUSE3), an E4 ubiquitin ligase involved in polyubiquitination of its protein targets. Knocking out MUSE3 in Arabidopsis thaliana results in increased levels of NLRs, including SNC1 and RPS2, whereas overexpressing MUSE3 together with CPR1 enhances polyubiquitination and protein degradation of these immune receptors. This report on the functional role of an E4 ligase in plants provides insight into the scarcely understood NLR degradation pathway.

Probing formation of cargo/importin-α transport complexes in plant cells using a pathogen effector.

Importin-αs are essential adapter proteins that recruit cytoplasmic proteins destined for active nuclear import to the nuclear transport machinery. Cargo proteins interact with the importin-α armadillo repeat domain via nuclear localization sequences (NLSs), short amino acids motifs enriched in Lys and Arg residues. Plant genomes typically encode several importin-α paralogs that can have both specific and partially redundant functions. Although some cargos are preferentially imported by a distinct importin-α it remains unknown how this specificity is generated and to what extent cargos compete for binding to nuclear transport receptors. Here we report that the effector protein HaRxL106 from the oomycete pathogen Hyaloperonospora arabidopsidis co-opts the host cell's nuclear import machinery. We use HaRxL106 as a probe to determine redundant and specific functions of importin-α paralogs from Arabidopsis thaliana. A crystal structure of the importin-α3/MOS6 armadillo repeat domain suggests that five of the six Arabidopsis importin-αs expressed in rosette leaves have an almost identical NLS-binding site. Comparison of the importin-α binding affinities of HaRxL106 and other cargos in vitro and in plant cells suggests that relatively small affinity differences in vitro affect the rate of transport complex formation in vivo. Our results suggest that cargo affinity for importin-α, sequence variation at the importin-α NLS-binding sites and tissue-specific expression levels of importin-αs determine formation of cargo/importin-α transport complexes in plant cells.

Two activities of long-chain acyl-coenzyme A synthetase are involved in lipid trafficking between the endoplasmic reticulum and the plastid in Arabidopsis.

In plants, fatty acids are synthesized within the plastid and need to be distributed to the different sites of lipid biosynthesis within the cell. Free fatty acids released from the plastid need to be converted to their corresponding coenzyme A thioesters to become metabolically available. This activation is mediated by long-chain acyl-coenzyme A synthetases (LACSs), which are encoded by a family of nine genes in Arabidopsis (Arabidopsis thaliana). So far, it has remained unclear which of the individual LACS activities are involved in making plastid-derived fatty acids available to cytoplasmic glycerolipid biosynthesis. Because of its unique localization at the outer envelope of plastids, LACS9 was regarded as a candidate for linking plastidial fatty export and cytoplasmic use. However, data presented in this study show that LACS9 is involved in fatty acid import into the plastid. The analyses of mutant lines revealed strongly overlapping functions of LACS4 and LACS9 in lipid trafficking from the endoplasmic reticulum to the plastid. In vivo labeling experiments with lacs4 lacs9 double mutants suggest strongly reduced synthesis of endoplasmic reticulum-derived lipid precursors, which are required for the biosynthesis of glycolipids in the plastids. In conjunction with this defect, double-mutant plants accumulate significant amounts of linoleic acid in leaf tissue.

Analyses of wrky18 wrky40 plants reveal critical roles of SA/EDS1 signaling and indole-glucosinolate biosynthesis for Golovinomyces orontii resistance and a loss-of resistance towards Pseudomonas syringae pv. tomato AvrRPS4.

Simultaneous mutation of two WRKY-type transcription factors, WRKY18 and WRKY40, renders otherwise susceptible wild-type Arabidopsis plants resistant towards the biotrophic powdery mildew fungus Golovinomyces orontii. Resistance in wrky18 wrky40 double mutant plants is accompanied by massive transcriptional reprogramming, imbalance in salicylic acid (SA) and jasmonic acid (JA) signaling, altered ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1) expression, and accumulation of the phytoalexin camalexin. Genetic analyses identified SA biosynthesis and EDS1 signaling as well as biosynthesis of the indole-glucosinolate 4MI3G as essential components required for loss-of-WRKY18 WRKY40-mediated resistance towards G. orontii. The analysis of wrky18 wrky40 pad3 mutant plants impaired in camalexin biosynthesis revealed an uncoupling of pre- from postinvasive resistance against G. orontii. Comprehensive infection studies demonstrated the specificity of wrky18 wrky40-mediated G. orontii resistance. Interestingly, WRKY18 and WRKY40 act as positive regulators in effector-triggered immunity, as the wrky18 wrky40 double mutant was found to be strongly susceptible towards the bacterial pathogen Pseudomonas syringae DC3000 expressing the effector AvrRPS4 but not against other tested Pseudomonas strains. We hypothesize that G. orontii depends on the function of WRKY18 and WRKY40 to successfully infect Arabidopsis wild-type plants while, in the interaction with P. syringae AvrRPS4, they are required to mediate effector-triggered immunity.

Histone lysine methyltransferase-related neurodevelopmental disorders: current knowledge and saRNA future therapies.

Neurodevelopmental disorders encompass a group of debilitating diseases presenting with motor and cognitive dysfunction, with variable age of onset and disease severity. Advances in genetic diagnostic tools have facilitated the identification of several monogenic chromatin remodeling diseases that cause Neurodevelopmental disorders. Chromatin remodelers play a key role in the neuro-epigenetic landscape and regulation of brain development; it is therefore not surprising that mutations, leading to loss of protein function, result in aberrant neurodevelopment. Heterozygous, usually de novo mutations in histone lysine methyltransferases have been described in patients leading to haploinsufficiency, dysregulated protein levels and impaired protein function. Studies in animal models and patient-derived cell lines, have highlighted the role of histone lysine methyltransferases in the regulation of cell self-renewal, cell fate specification and apoptosis. To date, in depth studies of histone lysine methyltransferases in oncology have provided strong evidence of histone lysine methyltransferase dysregulation as a determinant of cancer progression and drug resistance. As a result, histone lysine methyltransferases have become an important therapeutic target for the treatment of different cancer forms. Despite recent advances, we still lack knowledge about the role of histone lysine methyltransferases in neuronal development. This has hampered both the study and development of precision therapies for histone lysine methyltransferases-related Neurodevelopmental disorders. In this review, we will discuss the current knowledge of the role of histone lysine methyltransferases in neuronal development and disease progression. We will also discuss how RNA-based technologies using small-activating RNAs could potentially provide a novel therapeutic approach for the future treatment of histone lysine methyltransferase haploinsufficiency in these Neurodevelopmental disorders, and how they could be first tested in state-of-the-art patient-derived neuronal models.

Knockout of bone sialoprotein in cementoblasts cell lines affects specific gene expression in unstimulated and mechanically stimulated conditions.

One of the major components in cementum extracellular matrix is bone sialoprotein (BSP). BSP knockout (Ibsp) mice were reported to have a nonfunctional hypo-mineralized cementum, as well as detachment and disorganization of the periodontal ligament tissue. However, studies investigating the influence of Ibsp in cementoblasts are missing yet. This study investigates the influences of Bsp in three cementoblasts cell lines (OCCM.30-WT,IbspΔNterm, and IbspKAE). The mRNA expression of cementoblast and osteoclast markers (Col1a1, Alpl, Ocn, Runx2, Ctsk, Rankl and Opg) and the cell morphology were compared. Additionally, a functional monocyte adhesion assay was performed. To understand the influence of external stimuli, the effect of Ibsp was investigated under static compressive force, mimicking the compression side of orthodontic tooth movement. Cementoblasts with genotype IbspΔNterm and IbspKAE showed slight differences in cell morphology compared to OCCM.30-WT, as well as different gene expression. Under compressive force, the Ibsp cell lines presented expression pattern markers similar to the OCCM.30-WT cell line. However, Cathepsin K was strongly upregulated in IbspΔNterm cementoblasts under compressive force. This study provides insight into the role of BSP in cementoblasts and explores the influence of BSP on periodontal ligament tissues. BSP markers in cementoblasts seem to be involved in the regulation of cementum organization as an important factor for a functional periodontium. In summary, our findings provide a basis for investigations regarding molecular biology interactions of BSP in cementoblasts, and a supporting input for understanding the periodontal and cellular cementum remodeling.

Potential donor-dependent regulative effects of endogenous sclerostin expression and mineralization potential in primary human PDL cells in vitro.

OBJECTIVES: The glycoprotein sclerostin is mostly expressed in osteocytes and plays a central role in human bone metabolism. However, sclerostin and the corresponding SOST gene have been found in periodontal ligament cells under mineralizing conditions as well. The present study aimed to investigate, whether there was a correlation between endogenous SOST expression, the corresponding gene, and mineralization potential in human periodontal ligament cells and to identify different sclerostin expression and secretion patterns in cells derived from individual donors. MATERIAL AND METHODS: Primary human periodontal ligament cells of three different donors were cultivated under control or mineralizing conditions for 6, 13, 15 and 18 days, respectively. Calcium deposits were stained with alizarin red and quantified afterwards. Quantitative expression analysis of the SOST gene encoding sclerostin was performed using quantitative reverse transcription polymerase chain reaction (RT-PCR). Additionally, intracellular sclerostin expression was analyzed using Western blotting and extracellular sclerostin secretion was quantified using Enzyme-linked Immunosorbent Assay (ELISA). RESULTS: Alizarin red staining identified calcium deposits in periodontal ligament cells under mineralizing conditions beginning from day 13, relative SOST expression occurred on day 6. Whereas staining continued to increase in donor 1 on day 15, it remained stable in donors 2 and 3. Conversely, baseline SOST expression was significantly lower in donor 1 compared to donors 2 and 3. Western blotting and ELISA revealed increased intra- and extracellular sclerostin expression at day 13 under mineralizing conditions. Donor 3 exhibited the highest overall sclerostin levels. CONCLUSIONS: Our data emphasize donor-specific characteristics in differentiation potential and sclerostin expression patterns in primary human periodontal ligament cells. Sclerostin might play a central role in modulating osteogenic differentiation in periodontal ligament cells as part of a negative feedback mechanism in avoiding excessive mineralization.

Differentiation of human induced pluripotent stem cells into cortical neural stem cells.

Efficient and effective methods for converting human induced pluripotent stem cells into differentiated derivatives are critical for performing robust, large-scale studies of development and disease modelling, and for providing a source of cells for regenerative medicine. Here, we describe a 14-day neural differentiation protocol which allows for the scalable, simultaneous differentiation of multiple iPSC lines into cortical neural stem cells We currently employ this protocol to differentiate and compare sets of engineered iPSC lines carrying loss of function alleles in developmental disorder associated genes, alongside isogenic wildtype controls. Using RNA sequencing (RNA-Seq), we can examine the changes in gene expression brought about by each disease gene knockout, to determine its impact on neural development and explore mechanisms of disease. The 10-day Neural Induction period uses the well established dual-SMAD inhibition approach combined with Wnt/ß-Catenin inhibition to selectively induce formation of cortical NSCs. This is followed by a 4-day Neural Maintenance period facilitating NSC expansion and rosette formation, and NSC cryopreservation. We also describe methods for thawing and passaging the cryopreserved NSCs, which are useful in confirming their viability for further culture. Routine implementation of immunocytochemistry Quality Control confirms the presence of PAX6-positive and/or FOXG1-positive NSCs and the absence of OCT4-positive iPSCs after differentiation. RNA-Seq, flow cytometry, immunocytochemistry (ICC) and RT-qPCR provide additional confirmation of robust presence of NSC markers in the differentiated cells. The broader utility and application of our protocol is demonstrated by the successful differentiation of wildtype iPSC lines from five additional independent donors. This paper thereby describes an efficient method for the production of large numbers of high purity cortical NSCs, which are widely applicable for downstream research into developmental mechanisms, further differentiation into postmitotic cortical neurons, or other applications such as large-scale drug screening experiments.

Paternal effects in Arabidopsis indicate that offspring can influence their own size.

The existence of genetic variation in offspring size in plants and animals is puzzling because offspring size is often strongly associated with fitness and expected to be under stabilizing selection. An explanation for variation in seed size is conflict between parents and between parents and offspring. However, for this hypothesis to be true, it must be shown that the offspring genotype can affect its own size. The existence of paternal effects would support this hypothesis, but these have rarely been shown. Using a diallel cross among four natural accessions of Arabidopsis thaliana we show that maternal, paternal and positional effects jointly influence seed size, number and the frequency of seed abortion. We found that seed abortion (%) depends on the combination of maternal and paternal genotypes, suggesting the existence of mate choice or epistatic incompatibility among accessions of A. thaliana. In addition, since paternal genotype explains approximately 10 per cent of the variation in seed size, we propose that A. thaliana's offspring must influence the amount of resources allocated to themselves. Identification of paternal effects in Arabidopsis should facilitate dissection of the genetic mechanisms involved in paternal effects.

Decodability of Reward Learning Signals Predicts Mood Fluctuations.

Our mood often fluctuates without warning. Recent accounts propose that these fluctuations might be preceded by changes in how we process reward. According to this view, the degree to which reward improves our mood reflects not only characteristics of the reward itself (e.g., its magnitude) but also how receptive to reward we happen to be. Differences in receptivity to reward have been suggested to play an important role in the emergence of mood episodes in psychiatric disorders [1-16]. However, despite substantial theory, the relationship between reward processing and daily fluctuations of mood has yet to be tested directly. In particular, it is unclear whether the extent to which people respond to reward changes from day to day and whether such changes are followed by corresponding shifts in mood. Here, we use a novel mobile-phone platform with dense data sampling and wearable heart-rate and electroencephalographic sensors to examine mood and reward processing over an extended period of one week. Subjects regularly performed a trial-and-error choice task in which different choices were probabilistically rewarded. Subjects' choices revealed two complementary learning processes, one fast and one slow. Reward prediction errors [17, 18] indicative of these two processes were decodable from subjects' physiological responses. Strikingly, more accurate decodability of prediction-error signals reflective of the fast process predicted improvement in subjects' mood several hours later, whereas more accurate decodability of the slow process' signals predicted better mood a whole day later. We conclude that real-life mood fluctuations follow changes in responsivity to reward at multiple timescales.

MOS6 and TN13 in plant immunity.

The Arabidopsis nuclear transport receptor IMPORTIN-α3/MOS6 (MODIFIER OF SNC1, 6) is required for constitutive defense responses of the auto-immune mutant snc1 (suppressor of npr1-1, constitutive 1) and contributes to basal disease resistance, suggesting a role in nuclear import of defense-regulatory cargo proteins. We recently showed that MOS6 selectively interacts with TN13, a TIR-NBS protein involved in basal resistance to Pseudomonas syringae pv. tomato (Pst) DC3000 lacking the effectors AvrPto and AvrPtoB. Consistent with a predicted N-terminal transmembrane domain, TN13 localizes to the endoplasmic reticulum (ER) and the nuclear envelope (NE) where it interacts with MOS6 in a transient expression assay. Here, we propose a model that summarizes the subcellular localization, association and function of TN13 and MOS6 in plant defense signaling.

Changes in motor cortex excitability for the trained and non-trained hand after long-term unilateral motor training.

Repetitive unilateral upper limb motor training does not only affect behavior but also increases excitability of the contralateral primary motor cortex (M1). The behavioral gain is partially transferred to the non-trained side. Changes in M1 intracortical facilitation (ICF) might as well be observed for both hand sides. We measured ICF of both left and right abductor pollicis brevis muscles (APB) before and after a two-week period of arm ability training (AAT) of the left hand in 13 strongly right handed healthy volunteers. Performance with AAT-tasks improved for both the left trained and right untrained hand. ICF for the untrained hand decreased over training while it remained unchanged for the left trained hand. Decrease of ICF for the right hand was moderately associated with an increase of AAT-performance for the untrained right hand. We conclude that ICF-imbalance between dominant and non-dominant hand is sensitive to long-term motor training: training of the non-dominant hand results in a decrease of ICF of the dominant hand. The ICF-decrease is associated with a transfer of training-induced improvement of performance from the non-dominant to the dominant hand.

Hop-on hop-off: importin-α-guided tours to the nucleus in innate immune signaling.

Nuclear translocation of immune regulatory proteins and signal transducers is an essential process in animal and plant defense signaling against pathogenic microbes. Import of proteins containing a nuclear localization signal (NLS) into the nucleus is mediated by nuclear transport receptors termed importins, typically dimers of a cargo-binding α-subunit and a ß-subunit that mediates translocation through the nuclear pore complex. Here, we review recent reports of importin-α cargo specificity and mutant phenotypes in plant- and animal-microbe interactions. Using homology modeling of the NLS-binding cleft of nine predicted Arabidopsis α-importins and analyses of their gene expression patterns, we discuss functional redundancy and specialization within this transport receptor family. In addition, we consider how pathogen effector proteins that promote infection by manipulating host cell nuclear processes might compete with endogenous cargo proteins for nuclear uptake.

Mitochondrial AtPAM16 is required for plant survival and the negative regulation of plant immunity.

Proteins containing nucleotide-binding and leucine-rich repeat domains (NB-LRRs) serve as immune receptors in plants and animals. Negative regulation of immunity mediated by NB-LRR proteins is crucial, as their overactivation often leads to autoimmunity. Here we describe a new mutant, snc1-enhancing (muse) forward genetic screen, targeting unknown negative regulators of NB-LRR-mediated resistance in Arabidopsis. From the screen, we identify MUSE5, which is renamed as AtPAM16 because it encodes the ortholog of yeast PAM16, part of the mitochondrial inner membrane protein import motor. Consistently, AtPAM16-GFP localizes to the mitochondrial inner membrane. AtPAM16L is a paralog of AtPAM16. Double mutant Atpam16-1 Atpam16l is lethal, indicating that AtPAM16 function is essential. Single mutant Atpam16 plants exhibit a smaller size and enhanced resistance against virulent pathogens. They also display elevated reactive oxygen species (ROS) accumulation. Therefore, AtPAM16 seems to be involved in importing a negative regulator of plant immunity into mitochondria, thus protecting plants from over-accumulation of ROS and preventing autoimmunity.