A Calcium-Dependent Protein Kinase Regulates the Defense Response in Citrus sinensis.

Citrus Huanglongbing (HLB), which is caused by 'Candidatus Liberibacter asiaticus' (CLas), is one of the most destructive citrus diseases worldwide, and defense-related Citrus sinensis gene resources remain largely unexplored. Calcium signaling plays an important role in diverse biological processes. In plants, a few calcium-dependent protein kinases (CDPKs/CPKs) have been shown to contribute to defense against pathogenic microbes. The genome of C. sinensis encodes dozens of CPKs. In this study, the role of C. sinensis calcium-dependent protein kinases (CsCPKs) in C. sinensis defense was investigated. Silencing of CsCPK6 compromised the induction of defense-related genes in C. sinensis. Expression of a constitutively active form of CsCPK6 (CsCPK6CA) triggered the activation of defense-related genes in C. sinensis. Complementation of CsCPK6 rescued the defense-related gene induction in an Arabidopsis thaliana cpk4/11 mutant, indicating that CsCPK6 carries CPK activity and is capable of functioning as a CPK in Arabidopsis. Moreover, an effector derived from CLas inhibits defense induced by the expression of CsCPK6CA and autophosphorylation of CsCPK6, which suggests the involvement of CsCPK6 and calcium signaling in defense. These results support a positive role for CsCPK6 in C. sinensis defense against CLas, and the autoinhibitory regulation of CsCPK6 provides a potential genome-editing target for improving C. sinensis defense. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Plant cellular messengers mobilized to defend.

Phosphatidic acid (PA) and reactive oxygen species (ROS) are cellular messengers that relay signals to regulate diverse biological processes. In recent issues of Cell Host & Microbe and Cell, Qi et al. and Kong et al., respectively, investigate diacylglycerol kinase 5-mediated PA in regulating ROS signaling and plant immunity.

A complete MAP kinase cascade controls hyphopodium formation and virulence of Verticillium dahliae.

Phytopathogens develop specialized infection-related structures to penetrate plant cells during infection. Different from phytopathogens that form appressoria or haustoria, the soil-borne root-infecting fungal pathogen Verticillium dahliae forms hyphopodia during infection, which further differentiate into penetration pegs to promote infection. The molecular mechanisms underlying the regulation of hyphopodium formation in V. dahliae remain poorly characterized. Mitogen-activated protein kinases (MAPKs) are highly conserved cytoplasmic kinases that regulate diverse biological processes in eukaryotes. Here we found that deletion of VdKss1, out of the five MAPKs encoded by V. dahliae, significantly impaired V. dahliae hyphopodium formation, in vitro penetration, and pathogenicity in cotton plants. Constitutive activation of MAPK kinase (MAPKK) VdSte7 and MAPK kinase kinase (MAPKKK) VdSte11 specifically activate VdKss1. Deletion of VdSte7 or VdSte11 resulted in a phenotype similar to that of the mutant with VdKss1 deletion. Thus, this study demonstrates that VdSte11-VdSte7-VdKss1 is a core MAPK cascade that regulates hyphopodium formation and pathogenicity in V. dahliae. Supplementary Information: The online version contains supplementary material available at 10.1007/s42994-023-00102-y.

Pathogenesis mechanisms of phytopathogen effectors.

Plants commonly face the threat of invasion by a wide variety of pathogens and have developed sophisticated immune mechanisms to defend against infectious diseases. However, successful pathogens have evolved diverse mechanisms to overcome host immunity and cause diseases. Different cell structures and unique cellular organelles carried by plant cells endow plant-specific defense mechanisms, in addition to the common framework of innate immune system shared by both plants and animals. Effectors serve as crucial virulence weapons employed by phytopathogens to disarm the plant immune system and promote infection. Here we summarized the many diverse strategies by which phytopathogen effectors overcome plant defense and prospected future perspectives. This article is categorized under: Infectious Diseases > Molecular and Cellular Physiology.

Chitin Synthase Genes Are Differentially Required for Growth, Stress Response, and Virulence in Verticillium dahliae.

Crop wilt disease caused by Verticillium dahliae usually leads to serious yield loss. Chitin, an important component of most fungal cell walls, functions to maintain the rigidity of cell walls and septa. Chitin synthesis mainly relies on the activity of chitin synthase (CHS). Eight CHS genes have been predicted in V. dahliae. In this study, we characterized the functions of these genes in terms of growth, stress responses, penetration, and virulence. Results showed that VdCHS5 is important for conidia germination and resistance to hyperosmotic stress. Conidial production is significantly decreased in Vdchs1, Vdchs4, and Vdchs8 mutants. VdCHS1, VdCHS2, VdCHS4, VdCHS6, VdCHS7, and VdCHS8 genes are important for cell wall integrity, while all mutants are important for cell membrane integrity. All of the VdCHS genes, except for VdCHS3, are required for the full pathogenicity of V. dahliae to Arabidopsis thaliana and cotton plants. The in vitro and in vivo penetration of Vdchs1, Vdchs4, Vdchs6, and Vdchs7 mutants was impaired, while that of the other mutants was normal. Overall, our results indicate that the VdCHS genes exert diverse functions to regulate the growth and development, conidial germination, conidial production, stress response, penetration, and virulence in V. dahliae.

TOUCH 3 and CALMODULIN 1/4/6 cooperate with calcium-dependent protein kinases to trigger calcium-dependent activation of CAM-BINDING PROTEIN 60-LIKE G and regulate fungal resistance in plants.

Plants utilize localized cell-surface and intracellular receptors to sense microbes and activate the influx of calcium, which serves as an important second messenger in eukaryotes to regulate cellular responses. However, the mechanisms through which plants decipher calcium influx to activate immune responses remain largely unknown. Here, we show that pathogen-associated molecular patterns (PAMPs) trigger calcium-dependent phosphorylation of CAM-BINDING PROTEIN 60-LIKE G (CBP60g) in Arabidopsis (Arabidopsis thaliana). CALCIUM-DEPENDENT PROTEIN KINASE5 (CPK5) phosphorylates CBP60g directly, thereby enhancing its transcription factor activity. TOUCH 3 (TCH3) and its homologs CALMODULIN (CAM) 1/4/6 and CPK4/5/6/11 are required for PAMP-induced CBP60g phosphorylation. TCH3 interferes with the auto-inhibitory region of CPK5 and promotes CPK5-mediated CBP60g phosphorylation. Furthermore, CPKs-mediated CBP60g phosphorylation positively regulates plant resistance to soil-borne fungal pathogens. These lines of evidence uncover a novel calcium signal decoding mechanism during plant immunity through which TCH3 relieves auto-inhibition of CPK5 to phosphorylate and activate CBP60g. The findings reveal cooperative interconnections between different types of calcium sensors in eukaryotes.

Evaluation of autofluorescence visualization system in the delineation of oral squamous cell carcinoma surgical margins.

INTRODUCTION: Delineating the margins of Oral squamous cell carcinoma (OSCC) is a critical step for optimaltumor resection. The aim of this study was to evaluate the accuracy of lesion surgical margin identification using autofluorescence visualization. MATERIALS AND METHODS: Thirty patients with OSCC were included in this study. For each lesion, the fluorescence loss boundary was determined using VELscope before ablative surgical resection (with a 1.5-2cm safety margin) was performed. A total of 126 samples were obtained from 30 surgical specimens, each containing the tissue from the fluorescence loss boundary to surgical margin. The status of each sample was determined by oral pathologists and the staining intensities of Ki-67, E-cadherin, and Vimentin at the fluorescence loss boundary and surgical margin were evaluated by immunohistochemistry. RESULTS: Fluorescence loss regions were identified in all patients. Of the 126 samples collected, HE staining identified 77 normal epithelia (61.1%), 26 mild dysplasia (20.6%), 17 severe dysplasia (13.4%) and 6 carcinomas in situ (4.9%). A significant correlation was found between the differentiation grade of tumor cells and the pathological status of the surgical marginal specimens (P<0.05). Forty-two of the 126 samples were randomly selected for further immunohistochemical staining. No significant differences were seen in Ki-67, E-cadherin, or Vimentin expression at the fluorescence loss boundary or surgical margin, however, the proteins' expression level was positively correlated with the degree of dysplasia (P<0.01). CONCLUSION: Autofluorescence visualization has potential as a simple surgical margin setting device for OSCC and may help delineate the superficial area of OSCC with acceptable accuracy. However, when considering the inherent limitations of this system, we suggest that the approach should only be applied under certain conditions, such as when dealing with superficial, well-differentiated lesions.

SATB2 overexpression promotes oral squamous cell carcinoma progression by up-regulating NOX4.

While atypical expression of special AT-rich sequence-binding protein 2 (SATB2) has been approved associated with tumor progression, metastasis and unfavourable prognosis in various carcinomas. However, in oral squamous cell carcinoma (OSCC), both the expressive state and associated functions of SATB2's are still undefined. Here we show that, in clinical samples from a retrospective cohort of 58 OSCC patients, high expression of SATB2 is associated with poor prognosis of OSCC patients. In this study, we investigated SATB2 is highly expressed in OSCC tissues and cell lines, which can promote OSCC cells' proliferation, migration, invasion and tumor growth. According to sequencing results based on previous literature, we identified NOX4 is a bona fide downstream target of SATB2, when it was knockdown, OSCC's proliferation can be partially suppressed. Furthermore, NOX4 knockdown inhibits tumorigenicity, which can be rescued partially by ectopic expression of SATB2 in HNSCC cell line, and vice versa. Collectively, our findings not only indicate overexpression of SATB2 triggers the proliferative, migratory and invasive mechanisms which are important in the malignant phenotype of OSCC, but also identify NOX4 as the downstream gene for SATB2. These findings indicate that SATB2 may play a key role in OSCC tumorigenicity and may be a future target for the development of new therapeutic regimens.

Regulatory role of receptor-like cytoplasmic kinases in early immune signaling events in plants.

Receptor-like cytoplasmic kinases (RLCKs) play crucial roles in regulating plant development and immunity. Conserved pathogen-associated molecular patterns (PAMPs) derived from microbes are recognized by plant pattern recognition receptors to activate PAMP-triggered immunity (PTI). Microbial effectors, whose initial function is to promote virulence, are recognized by plant intracellular nucleotide-binding domain and leucine-rich repeat receptors (NLRs) to initiate effector-triggered immunity (ETI). Both PTI and ETI trigger early immune signaling events including the production of reactive oxygen species, induction of calcium influx and activation of mitogen-activated protein kinases. Research progress has revealed the important roles of RLCKs in the regulation of early PTI signaling. Accordingly, RLCKs are often targeted by microbial effectors that are evolved to evade PTI via diverse modulations. In some cases, modulation of RLCKs by microbial effectors triggers the activation of NLRs. This review covers the mechanisms by which RLCKs engage diverse substrates to regulate early PTI signaling and the regulatory roles of RLCKs in triggering NLR activation. Accumulating evidence suggests evolutionary links and close connections between PAMP- and effector-triggered early immune signaling that are mediated by RLCKs. As key immune regulators, RLCKs can be considered targets with broad prospects for the improvement of plant resistance via genetic engineering.

An Approach to Robust INS/UWB Integrated Positioning for Autonomous Indoor Mobile Robots.

The key to successful positioning of autonomous mobile robots in complicated indoor environments lies in the strong anti-interference of the positioning system and accurate measurements from sensors. Inertial navigation systems (INS) are widely used for indoor mobile robots because they are not susceptible to external interferences and work properly, but the positioning errors may be accumulated over time. Thus ultra wideband (UWB) is usually adopted to compensate the accumulated errors due to its high ranging precision. Unfortunately, UWB is easily affected by the multipath effects and non-line-of-sight (NLOS) factor in complex indoor environments, which may degrade the positioning performance. To solve above problems, this paper proposes an effective system framework of INS/UWB integrated positioning for autonomous indoor mobile robots, in which our modeling approach is simple to implement and a Sage⁻Husa fuzzy adaptive filter (SHFAF) is proposed. Due to the favorable property (i.e., self-adaptive adjustment) of SHFAF, the difficult problem of time-varying noise in complex indoor environments is considered and solved explicitly. Moreover, outliers can be detected and corrected by the proposed sliding window estimation with fading coefficients. This facilitates the positioning performance improvement for indoor mobile robots. The benefits of what we propose are illustrated by not only simulations but more importantly experimental results.

Cellophane surface-induced gene, VdCSIN1, regulates hyphopodium formation and pathogenesis via cAMP-mediated signalling in Verticillium dahliae.

The soil-borne vascular pathogen Verticillium dahliae infects many dicotyledonous plants to cause devastating wilt diseases. During colonization, V. dahliae spores develop hyphae surrounding the roots. Only a few hyphae that adhere tightly to the root surface form hyphopodia at the infection site, which further differentiate into penetration pegs to facilitate infection. The molecular mechanisms controlling hyphopodium formation in V. dahliae remain unclear. Here, we uncovered a cellophane surface-induced gene (VdCSIN1) as a regulator of V. dahliae hyphopodium formation and pathogenesis. Deletion of VdCSIN1 compromises hyphopodium formation, hyphal development and pathogenesis. Exogenous application of cyclic adenosine monophosphate (cAMP) degradation inhibitor or disruption of the cAMP phosphodiesterase gene (VdPDEH) partially restores hyphopodium formation in the VdΔcsin1 mutant. Moreover, deletion of VdPDEH partially restores the pathogenesis of the VdΔcsin1 mutant. These findings indicate that VdCSIN1 regulates hyphopodium formation via cAMP-mediated signalling to promote host colonization by V. dahliae.

Farmers' perceived efficacy of adaptive behaviors to climate change in the Loess Plateau, China.

The impact of climate change is very significant to farmers who depend on natural resources for livelihood. It is essential to have a better understanding of farmers' assessments of the efficacy of adaptive behaviors for formulating appropriate adaptation policies and improving farmers' ability to adapt to climate change. Based on survey data from interviews with farmers in the Loess Plateau, the features of farmers' perceived adaptation efficacy are analyzed. Three multiple linear regression models are used to analyze farmers' perceived efficacy of adaptive behaviors and identify factors influencing those assessments in terms of farmers' demographical and social factors, their perception of climate change, their perception of climate change effects and the average temperature and precipitation from 2005 to 2015 in this area. The results are as follows: (1) Generally, the adaptive behaviors with high perceived adaptation efficacy were used by most respondents. Measures with relatively low perceived adaptation efficacy were not commonly adopted, such as migration, buying insurance, changing planting and harvesting time. (2) The factors affect the perceived efficacy of adaption behavior in decreasing order are as follows: perception of climate change, the average precipitation, and demographical and social factors. Perception of local natural disasters, perception of planting and harvesting time, crop yield and diseases and insect pests caused by climate change were found to affect farmers' adaptation assessments. The key demographical and social factors influencing farmers' assessments were non-farming income, farming income, farmland quantity, gender, the frequency of watching TV and going to the market.

The Xanthomonas effector XopK harbours E3 ubiquitin-ligase activity that is required for virulence.

Xanthomonas oryzae pv. oryzae is the causative agent of rice bacterial leaf blight. While the type III secretion system of X. oryzae pv. oryzae is essential for virulence, the biochemical activities and virulence mechanisms of non-transcription activator-like (non-TAL) effectors delivered by this system are largely unknown. Here, by screening for non-TAL effectors that contribute to X. oryzae pv. oryzae virulence, we revealed that Xanthomonas outer protein K (XopK) inhibits pathogen-associated molecular pattern-triggered immunity upstream of mitogen-activated protein kinase cascades. Specifically, XopK interacted with and directly ubiquitinated rice somatic embryogenic receptor kinase 2 (OsSERK2), resulting in its degradation. Accordingly, mutation of a putative ubiquitin-conjugation enzyme (E2) binding site abolished XopK-induced degradation of OsSERK2 and compromised XopK-dependent virulence. As crucial immune regulators associated with a multitude of immune receptors, SERKs have been shown to be perturbed by Pseudomonas effectors via different mechanisms. Our study revealed a distinct perturbation mechanism of SERK activity via ubiquitination achieved by Xanthomonas non-TAL effector.

The plant-specific transcription factors CBP60g and SARD1 are targeted by a Verticillium secretory protein VdSCP41 to modulate immunity.

The vascular pathogen Verticillium dahliae infects the roots of plants to cause Verticillium wilt. The molecular mechanisms underlying V. dahliae virulence and host resistance remain elusive. Here, we demonstrate that a secretory protein, VdSCP41, functions as an intracellular effector that promotes V. dahliae virulence. The Arabidopsis master immune regulators CBP60g and SARD1 and cotton GhCBP60b are targeted by VdSCP41. VdSCP41 binds the C-terminal portion of CBP60g to inhibit its transcription factor activity. Further analyses reveal a transcription activation domain within CBP60g that is required for VdSCP41 targeting. Mutations in both CBP60g and SARD1 compromise Arabidopsis resistance against V. dahliae and partially impair VdSCP41-mediated virulence. Moreover, virus-induced silencing of GhCBP60b compromises cotton resistance to V. dahliae. This work uncovers a virulence strategy in which the V. dahliae secretory protein VdSCP41 directly targets plant transcription factors to inhibit immunity, and reveals CBP60g, SARD1 and GhCBP60b as crucial components governing V. dahliae resistance.

Tracking of Maneuvering Complex Extended Object with Coupled Motion Kinematics and Extension Dynamics Using Range Extent Measurements.

The key to successful maneuvering complex extended object tracking (MCEOT) using range extent measurements provided by high resolution sensors lies in accurate and effective modeling of both the extension dynamics and the centroid kinematics. During object maneuvers, the extension dynamics of an object with a complex shape is highly coupled with the centroid kinematics. However, this difficult but important problem is rarely considered and solved explicitly. In view of this, this paper proposes a general approach to modeling a maneuvering complex extended object based on Minkowski sum, so that the coupled turn maneuvers in both the centroid states and extensions can be described accurately. The new model has a concise and unified form, in which the complex extension dynamics can be simply and jointly characterized by multiple simple sub-objects' extension dynamics based on Minkowski sum. The proposed maneuvering model fits range extent measurements very well due to its favorable properties. Based on this model, an MCEOT algorithm dealing with motion and extension maneuvers is also derived. Two different cases of the turn maneuvers with known/unknown turn rates are specifically considered. The proposed algorithm which jointly estimates the kinematic state and the object extension can also be easily implemented. Simulation results demonstrate the effectiveness of the proposed modeling and tracking approaches.

Expansion of pathogen recognition specificity in plants using pattern recognition receptors and artificially designed decoys.

Pathogen/microbe-associated molecular patterns (PAMPs/MAMPs) are recognized by plant pattern recognition receptors (PRRs) localized on the cell surface to activate immune responses. This PAMP-triggered immunity (PTI) confers resistance to a broad range of pathogenic microbes and, therefore, has a great potential for genetically engineering broad-spectrum resistance by transferring PRRs across plant families. Pathogenic effectors secreted by phytopathogens often directly target and inhibit key components of PTI signaling pathways via diverse biochemical mechanisms. In some cases, plants have evolved to produce decoy proteins that mimic the direct virulence target, which senses the biochemical activities of pathogenic effectors. This kind of perception traps the effectors of erroneous targeting and results in the activation of effector-triggered immunity (ETI) instead of suppressing PTI. This mechanism suggests that artificially designed decoy proteins could be used to generate new recognition specificities in a particular plant. In this review, we summarize recent advances in research investigating PAMP recognition by PRRs and virulence effector surveillance by decoy proteins. Successful expansion of recognition specificities, conferred by the transgenic expression of EF-Tu receptor (EFR) and AvrPphB susceptible 1 (PBS1) decoys, has highlighted the considerable potential of PRRs and artificially designed decoys to expand plant resistance spectra and the need to further identify novel PRRs and decoys.

Contributory roles of two l-lactate dehydrogenases for l-lactic acid production in thermotolerant Bacillus coagulans.

Thermotolerant Bacillus coagulans is considered to be a more promising producer for bio-chemicals, due to its capacity to withstand harsh conditions. Two L-lactate dehydrogenase (LDH) encoding genes (ldhL1 and ldhL2) and one D-LDH encoding gene (ldhD) were annotated from the B. coagulans DSM1 genome. Transcriptional analysis revealed that the expression of ldhL2 was undetectable while the ldhL1 transcription level was much higher than that of ldhD at all growth phases. Deletion of the ldhL2 gene revealed no difference in fermentation profile compared to the wild-type strain, while ldhL1 single deletion or ldhL1ldhL2 double deletion completely blocked L-lactic acid production. Complementation of ldhL1 in the above knockout strains restored fermentation profiles to those observed in the wild-type strain. This study demonstrates ldhL1 is crucial for L-lactic acid production and NADH balance in B. coagulans DSM1 and lays the fundamental for engineering the thermotolerant B. coagulans strain as a platform chemicals producer.

Diammonium phosphate stimulates transcription of L-lactate dehydrogenase leading to increased L-lactate production in the thermotolerant Bacillus coagulans strain.

Exploration of cost-effective fermentation substrates for efficient lactate production is an important economic objective. Although some organic nitrogen sources are also cheaper, inorganic nitrogen salts for lactate fermentation have additional advantages in facilitating downstream procedures and significantly improving the commercial competitiveness of lactate production. In this study, we first established an application of diammonium phosphate to replace yeast extract with a reduced 90 % nitrogen cost for a thermotolerant Bacillus coagulans strain. In vivo enzymatic and transcriptional analyses demonstrated that diammonium phosphate stimulates the gene expression of L-lactate dehydrogenase, thus providing higher specific enzyme activity in vivo and increasing L-lactic acid production. This new information provides a foundation for establishing a cost-effective process for polymer-grade L-lactic acid production in an industrial setting.

Comparative Proteomic Insights into the Lactate Responses of Halophilic Salinicoccus roseus W12.

Extremophiles use adaptive mechanisms to survive in extreme environments, which is of great importance for several biotechnological applications. A halophilic strain, Salinicoccus roseus W12, was isolated from salt lake in Inner Mongolia, China in this study. The ability of the strain to survive under high sodium conditions (including 20% sodium lactate or 25% sodium chloride, [w/v]) made it an ideal host to screen for key factors related to sodium lactate resistance. The proteomic responses to lactate were studied using W12 cells cultivated with or without lactate stress. A total of 1,656 protein spots in sodium lactate-treated culture and 1,843 spots in NaCl-treated culture were detected by 2-dimensional gel electrophoresis, and 32 of 120 significantly altered protein spots (fold change > 2, p < 0.05) were identified by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry. Among 21 successfully identified spots, 19 proteins were upregulated and 2 were downregulated. The identified proteins are mainly involved in metabolism, cellular processes and signaling, and information storage and processing. Transcription studies confirmed that most of the encoding genes were upregulated after the cells were exposed to lactate in 10 min. Cross-protecting and energy metabolism-related proteins played an important role in lactate tolerance for S. roseus W12.

Genome Sequence of Bacillus coagulans P38, an Efficient Polymer-Grade l-Lactate Producer from Cellulosic Substrates.

Bacillus coagulans P38 is an efficient polymer-grade l-lactic acid producer from a cellulosic carbon source. Here, the draft 3.37-Mb genome sequence of this potential strain may provide useful information to further improve the strain performance for higher titers and, importantly, to understand the mechanism of its high tolerance for 2-furfural.