Induction System Research Articles

Establishment of a callus induction system of Saxifraga stolonifera Meerb. and its response to different elicitors

Elicitation is a well-known strategy for increasing the accumulation of secondary metabolites in callus cultures. In this study, a callus induction system for Saxifraga stolonifera Meerb. was developed, and elicitors were used to enhance secondary metabolite production. Tender leaves of S. stolonifera were used as explants for in vitro culture. We examined callus induction characteristics of S. stolonifera and its response to various elicitors by assessing the sterilization conditions of the explants, varying hormone ratios, and determining the optimal concentrations of salicylic acid (SA), Sodium nitroprusside (SNP) and methyl jasmonate (MeJA). The results revealed that sterilizing the explants with 70% ethanol for 40 s, followed by 0.1% mercury chloride for 10 min, resulted in a survival rate of 82.22% during tissue culture. Following these sterilization methods, callus formation was successfully induced on Murashige and Skoog medium supplemented with 2 mg/L 6-benzylaminopurine and 0.15 mg/L naphthaleneacetic acid, achieving an induction rate of 88.88%. Additionally, the study identified SA as the most effective elicitor for increasing the levels of gallic acid, protocatechuic acid, and quercitrin in the callus, with optimal concentrations of salicylic acid being 60, 40, and 60 μM, respectively. SNP at 40 μM, 80 μM, and 20 μM concentrations was identified as the best elicitor for promoting the accumulation of bergenin, chlorogenic acid, and quercetin, respectively. This study established appropriate conditions for cultivating S. stolonifera callus and identified the optimal elicitor concentrations for maximizing secondary metabolite production.

OptoLacI: optogenetically engineered lactose operon repressor LacI responsive to light instead of IPTG.

Optogenetics' advancement has made light induction attractive for controlling biological processes due to its advantages of fine-tunability, reversibility, and low toxicity. The lactose operon induction system, commonly used in Escherichia coli, relies on the binding of lactose or isopropyl β-d-1-thiogalactopyranoside (IPTG) to the lactose repressor protein LacI, playing a pivotal role in controlling the lactose operon. Here, we harnessed the light-responsive light-oxygen-voltage 2 (LOV2) domain from Avena sativa phototropin 1 as a tool for light control and engineered LacI into two light-responsive variants, OptoLacIL and OptoLacID. These variants exhibit direct responsiveness to light and darkness, respectively, eliminating the need for IPTG. Building upon OptoLacI, we constructed two light-controlled E.coli gene expression systems, OptoE.coliLight system and OptoE.coliDark system. These systems enable bifunctional gene expression regulation in E.coli through light manipulation and show superior controllability compared to IPTG-induced systems. We applied the OptoE.coliDark system to protein production and metabolic flux control. Protein production levels are comparable to those induced by IPTG. Notably, the titers of dark-induced production of 1,3-propanediol (1,3-PDO) and ergothioneine exceeded 110% and 60% of those induced by IPTG, respectively. The development of OptoLacI will contribute to the advancement of the field of optogenetic protein engineering, holding substantial potential applications across various fields.

Restoration of degraded waterlogged saline soil through subsurface drainage: Spatiotemporal soil salinity assessment using electromagnetic induction (EMI) and geographic information system (GIS) techniques

AbstractReduction in soil salinity in the entire project area is a key indicator of proper functioning of the implemented subsurface drainage (SSD) projects. Electromagnetic induction (EMI) surveys were carried out to facilitate spatiotemporal assessment of soil salinity of a SSD project site in Kahni, Haryana (India). The surveys were conducted in two time frames: immediately after the start of SSD project in 2019 and after 2 years of successful operation in 2021. The collected data of 2 surveys was used to derive a depth‐specific calibrated correlation between the observed salinity (ECe) of collected soil samples and surveyed apparent electrical conductivity (ECa) observations in vertical (ECav) and horizontal (ECah) modes. Spatiotemporal analysis of soil salinity revealed notable changes in areas under different salinity classes over a period of 2 years, indicating the impact of SSD on soil desalinization. During the course of 2 years (2019–2021) of SSD operation, area under moderate salinity (4–8 dS m−1) in the top 0–0.15 m soil increased by 54.5% while it decreased by 40% and 6.6% under high (8–12 dS m−1) and very high (12–16 dS m−1) salinity classes. The rice and wheat yields were also enhanced by 24% and 7% respectively after 2 years operation of SSD in hitherto severally waterlogged saline soils. Overall findings of this study reinforce the scope of using EMI approach to quickly and effectively assess the efficacy of subsurface drainage. Creation of measurable soil salinity maps by proposed EMI and GIS techniques can help in planning and implementing site‐specific management strategies in saline soil reclamation projects.

An analysis of semaphorin-mediated cellular interactions in the Caenorhabditis elegans epidermis using the IR-LEGO single-cell gene induction system.

One of the major functions of the semaphorin signaling system is the regulation of cell shape. In the nematode Caenorhabditis elegans, membrane-bound semaphorins SMP-1/2 (SMPs) regulate the morphology of epidermal cells via their receptor plexin, PLX-1. In the larval male tail of the SMP-PLX-1 signaling mutants, the border between two epidermal cells, R1.p and R2.p, is displaced anteriorly, resulting in the anterior displacement of the anterior-most ray, ray 1, in the adult male. To elucidate how the intercellular signaling mediated by SMPs regulates the position of the intercellular border, we performed mosaic gene expression analyses by using infrared laser-evoked gene operator (IR-LEGO). We show that PLX-1 expressed in R1.p and SMP-1 expressed in R2.p are required for the proper positioning of ray 1. The result suggests that SMP signaling promotes extension, rather than retraction, of R1.p. This is in contrast to a previous finding that SMPs mediate inhibition of cell extension of vulval precursor cells, another group of epidermal cells of C. elegans, indicating the context dependence of cell shape control via the semaphorin signaling system.

Expression of soluble moloney murine leukemia virus-reverse transcriptase in Escherichia coli BL21 star (DE3) using autoinduction system.

Autoinduction systems in Escherichia coli can control the production of proteins without the addition of a particular inducer. In the present study, we optimized the heterologous expression of Moloney Murine Leukemia Virus derived Reverse Transcriptase (MMLV-RT) in E. coli. Among 4 autoinduction media, media Imperial College resulted the highest MMLV-RT overexpression in E. coli BL21 Star (DE3) with incubation time 96h. The enzyme was produced most optimum in soluble fraction of lysate cells. The MMLV-RT was then purified using the Immobilized Metal Affinity Chromatography method and had specific activity of 629.4U/mg. The system resulted lower specific activity and longer incubation of the enzyme than a classical Isopropyl ß-D-1-thiogalactopyranoside (IPTG)-induction system. However, the autoinduction resulted higher yield of the enzyme than the conventional induction (27.8%). Techno Economic Analysis revealed that this method could produce MMLV-RT using autoinduction at half the cost of MMLV-RT production by IPTG-induction. Bioprocessing techniques are necessary to conduct to obtain higher quality of MMLV-RT under autoinduction system.

Research on the function of CsMYB36 based on an effective hair root transformation system

ABSTRACT The hairy root induction system was used to efficiently investigate gene expression and function in plant root. Cucumber is a significant vegetable crop worldwide, with shallow roots, few lateral roots, and weak root systems, resulting in low nutrient absorption and utilization efficiency. Identifying essential genes related to root development and nutrient absorption is an effective way to improve the growth and development of cucumbers. However, genetic mechanisms underlying cucumber root development have not been explored. Here, we report a novel, rapid, effective hairy root transformation system. Compared to the in vitro cotyledon transformation method, this method shortened the time needed to obtain transgenic roots by 13 days. Furthermore, we combined this root transformation method with CRISPR/Cas9 technology and validated our system by exploring the expression and function of CsMYB36, a pivotal gene associated with root development and nutrient uptake. The hairy root transformation system established in this study provides a powerful method for rapidly identifying essential genes related to root development in cucumber and other horticultural crop species. This advancement holds promise for expediting research on root biology and molecular breeding strategies, contributing to the broader understanding and improvements crop growth and development.

Comparison of incremental encoder digital signal processing techniques for theinduction motor flux-torque vector control systems

The articlepresents the results of a study on the effectiveness of varioustechniques forsynchronous reference frame angular position numerical calculation in flux-torque vector control system of induction motor.Investigation was caried out taking into account the discrete nature of the angular speedsignal obtained using an incremental encoder. In this workfor investigation by simulation useda direct torque vector control system, which, in the presence of an ideal rotor angular speedsignal, ensures direct asymptotic field orientation, asymptotic tracking of torque-flux referencetrajectories, as well as asymptotic decouplingtorque and flux subsystems. The parameters of the inductionmotor and encoder used in the study correspond to those existing in traction electromechanical systems of city trolleybuses. It is shown that the discrete nature of the angular speedsignal, which usedin synchronous reference frameposition equation of flux-torque vector control systems, introduces field orientation errors and leads to current and torque ripples, which in a real system increase acoustic noise and can cause mechanical vibrations and resonance phenomena. An analysis of possible ways to reduce the influence of the speed signal discreteness on flux-torque control is performed, and a method for practical implementation of the synchronous reference frame angular position numerical calculationis proposed. This method allows ensuring conditions for more precisefield orientation and, by using an additional filter for the angular speedsignal, reducing the level of current and torque ripplesto negligibly small values without affecting the field orientation processes. The proposed solution can be used in the development of high dynamicflux-torque vector control systems for inductionmotors using incremental encoders, including for electric vehicles.

Autonomous differentiation of transgenic cells requiring no external hormone application: the endogenous gene expression and phytohormone behaviors.

The ectopic overexpression of developmental regulator (DR) genes has been reported to improve the transformation in recalcitrant plant species because of the promotion of cellular differentiation during cell culture processes. In other words, the external plant growth regulator (PGR) application during the tissue and cell culture process is still required in cases utilizing DR genes for plant regeneration. Here, the effect of Arabidopsis BABY BOOM (BBM) and WUSCHEL (WUS) on the differentiation of tobacco transgenic cells was examined. We found that the SRDX fusion to WUS, when co-expressed with the BBM-VP16 fusion gene, significantly influenced the induction of autonomous differentiation under PGR-free culture conditions, with similar effects in some other plant species. Furthermore, to understand the endogenous background underlying cell differentiation toward regeneration, phytohormone and RNA-seq analyses were performed using tobacco leaf explants in which transgenic cells were autonomously differentiating. The levels of active auxins, cytokinins, abscisic acid, and inactive gibberellins increased as cell differentiation proceeded toward organogenesis. Gene Ontology terms related to phytohormones and organogenesis were identified as differentially expressed genes, in addition to those related to polysaccharide and nitrate metabolism. The qRT-PCR four selected genes as DEGs supported the RNA-seq data. This differentiation induction system and the reported phytohormone and transcript profiles provide a foundation for the development of PGR-free tissue cultures of various plant species, facilitating future biotechnological breeding.

One-step creation of CMS lines using a BoCENH3-based haploid induction system in Brassica crop

Heterosis utilization in a large proportion of crops depends on the use of cytoplasmic male sterility (CMS) tools, requiring the development of homozygous fertile lines and CMS lines1. Although doubled haploid (DH) technology has been developed for several crops to rapidly generate fertile lines2,3, CMS lines are generally created by multiple rounds of backcrossing, which is time consuming and expensive4. Here we describe a method for generating both homozygous fertile and CMS lines through in vivo paternal haploid induction (HI). We generated in-frame deletion and restored frameshift mutants of BoCENH3 in Brassica oleracea using the CRISPR/Cas9 system. The mutants induced paternal haploids by outcrossing. We subsequently generated HI lines with CMS cytoplasm, which enabled the generation of homozygous CMS lines in one step. The BoCENH3-based HI system provides a new DH technology to accelerate breeding in Brassica and other crops.

Investigation of graphene-perovskite integration for enhanced absorption of solar thermal absorber for industrial heating applications

Advanced materials are critical for enhancing the efficiency of solar absorbers. Hence, the present study looks at the unique interaction between graphene and perovskite materials, a combination that has the potential to change the landscape of solar energy harvesting. The Graphene–Perovskite Integrated Solar Absorber (GPISA) structure harvests more than 98 % in UV and VIS regions ranging from 200 nm and goes up to 4000 nm of the solar spectrum using the Finite Element Method (FEM). Not only does it boast a broader absorption range from 200 to 4000 nm, covering virtually the entire solar spectrum, but it also achieves an impressive average absorption of 95.50 %. Additionally, the comprehensive analysis of its performance, encompasses optimized parameterization, exhaustive mapping of electric and magnetic field intensities, and in-depth investigation of both transverse magnetic (TM) and transverse electric (TE) field behaviors. This correlates to a weighted absorption rate of more than 97 % in the AM1.5 solar spectrum, as well as endurance to varied incoming angles (0°–80°) without considerable absorption loss, making it suitable for real-world direct sunlight absorptance. The GPISA is significantly more cost-effective due to its 3240 nm thickness. Overall, GPISA's exceptional combination of broad absorption, high efficiency, and compact design firmly positions it as a frontrunner in the advancement of efficient, high-performance solar energy harvesting. GPISA has efficient uses in solar induction systems as well as solar air, water heating, and other industrial heating applications.

An Intelligent Optimized Control System for Induction Heating Application

In the modern era, induction heating (IH) technology is utilized for industrial functions like medical applications and domestic applications. Furthermore, efficient uses such as control topologies, magnetic component design, and power electronics have high demand as well as cost-effective with respect to advanced technology. However, IH technology directly targets the power converter and coil so it has wasted higher amount of heat. Also, high power dissipation in an IH system leads to high energy costs and reduced system performance. Hence, various approaches are developed to minimize power dissipation. However, they cannot reduce power dissipation in the induction coil. Thus, a novel Decision-based Squirrel Control System was developed in this article to decrease the power wastage in induction coil optimally. The developed model’s squirrel fitness function monitors power wastage and compensates them by tuning it. The developed model is executed in MATLAB/Simulink, and the results are determined. In terms of rising time, settling time, overshoot, and error rate. Here, the proposed work can achieve 0.075s resting time, 1.02s settling time, 0.05s overshoot, and 0.29% of error rate. The comparative assessment proves that the presented approach attained better results than others.

Engineered Saccharomyces cerevisiae as a Biosynthetic Platform of Nucleotide Sugars.

Glycosylation of biomolecules can greatly alter their physicochemical properties, cellular recognition, subcellular localization, and immunogenicity. Glycosylation reactions rely on the stepwise addition of sugars using nucleotide diphosphate (NDP)-sugars. Making these substrates readily available will greatly accelerate the characterization of new glycosylation reactions, elucidation of their underlying regulation mechanisms, and production of glycosylated molecules. In this work, we engineered Saccharomyces cerevisiae to heterologously express nucleotide sugar synthases to access a wide variety of uridine diphosphate (UDP)-sugars from simple starting materials (i.e., glucose and galactose). Specifically, activated glucose, uridine diphosphate d-glucose (UDP-d-Glc), can be converted to UDP-d-glucuronic acid (UDP-d-GlcA), UDP-d-xylose (UDP-d-Xyl), UDP-d-apiose (UDP-d-Api), UDP-d-fucose (UDP-d-Fuc), UDP-l-rhamnose (UDP-l-Rha), UDP-l-arabinopyranose (UDP-l-Arap), and UDP-l-arabinofuranose (UDP-l-Araf) using the corresponding nucleotide sugar synthases of plant and microbial origins. We also expressed genes encoding the salvage pathway to directly activate free sugars to achieve the biosynthesis of UDP-l-Arap and UDP-l-Araf. We observed strong inhibition of UDP-d-Glc 6-dehydrogenase (UGD) by the downstream product UDP-d-Xyl, which we circumvented using an induction system (Tet-On) to delay the production of UDP-d-Xyl to maintain the upstream UDP-sugar pool. Finally, we performed a time-course study using strains containing the biosynthetic pathways to produce five non-native UDP-sugars to elucidate their time-dependent interconversion and the role of UDP-d-Xyl in regulating UDP-sugar metabolism. These engineered yeast strains are a robust platform to (i) functionally characterize sugar synthases in vivo, (ii) biosynthesize a diverse selection of UDP-sugars, (iii) examine the regulation of intracellular UDP-sugar interconversions, and (iv) produce glycosylated secondary metabolites and proteins.

Investigation of the Effect of High-Frequency Induction Sintering on Phase Structure and Microstructure of SiC Reinforced Aluminum Matrix Composites

In this study, SiC-reinforced aluminum matrix composites were powder metallurgically (PM) prepared and sintered using high-frequency induction system (HFIS). The samples with different ratios of SiC (wt.%10, 20 and 40) added to the aluminum matrix were sintered at 660, 800, and 1000 °C. In addition, Al/SiC composites were compared by sintering with the conventional sintering (CS) method under similar sintering conditions. The heating rate for the sintering process using HFIS was 500 °C/min, while the CS method used a heating rate of 10 °C/min. The effect of the temperature and SiC ratio on the density, hardness, phase structure, and microstructure of composites was investigated. The optimum sintering temperature was determined according to the SiC additive amount. When 10%, 20%, and 40% SiC by weight were added to the aluminum matrix in the sintering process with HFIS, the required sintering temperatures were determined as 660, 800, and 1000 °C, respectively. While new phases were not formed as a result of short-term HFIS sintering, a high-temperature Al4C3 phase was detected in CS sintering. HFIS sintered Al/SiC composite samples were obtained in Al and SiC phases with high density and hardness ranging from 43-118 HV. In the high-temperature sintering process with HFIS, the formation of Al4C3 was prevented and its physical and mechanical properties were improved.

Analysis of the dynamic characteristics of an eccentric cylinder in axial flow

The simple pendulum model is a fundamental and widely studied concept in physics, its movement follows a tendency to stabilize towards the side with lower gravitational potential energy. Consequently, the simple pendulum structures have been extensive applications in petroleum, aerospace machinery, and others, particularly in gravity induction and actuator systems. The utilization of this model offers versatile solutions for complex engineering challenges. The research object of this study is an eccentric cylindrical stabilized platform. A nonlinear dynamic model of the eccentric cylindrical stable platform in axial flow with variable damping is established. Due to the complexity of the analytical solution of the simple pendulum motion, we adopt the method of numerical analysis to building the simulation and analysis model. Combined with the fluid calculation and dynamics simulation methods, the motion characteristics of the eccentric cylindrical model under steady and unsteady flow fields are analyzed by adjusting the rotational speed, inclination angle, and initial pendulum angle. The experimental platform is built to analyze the motion law of the eccentric cylinder under the action of axial flow, and the experimental results of the motion law of the eccentric cylindrical platform are consistent with the numerical calculation results.

An Efficient Agrobacterium-Mediated Genetic Transformation System for Gene Editing in Strawberry (Fragaria × ananassa).

The octoploid-cultivated strawberry variety Benihope (Fragaria × ananassa Duch cv. Benihope) is an important commercial plant. It is highly susceptible to different diseases, which ultimately leads to a reduction in yield. Gene-editing methods, such as CRISPR/Cas9, demonstrate potential for improving disease resistance in the strawberry cv. Benihope. Establishing a plant regeneration system suitable for CRISPR/Cas9 gene editing is crucial for obtaining transgenic plants on a large scale. This research established a callus induction and plant regeneration system for Agrobacterium-mediated CRISPR/Cas9 gene editing in strawberry cv. Benihope by evaluating multiple types of explants and various plant growth regulators throughout the entire tissue culture process. The results showed that the efficiency of callus induction is strongly influenced by the type of explant and is highly sensitive to the combination of plant growth regulators. Among the different plant growth regulators employed, thidiazuron (TDZ), in combination with 2,4-dichlorophenoxyacetic acid (2,4-D), effectively induced callus formation and plant regeneration from explants derived from nutrient tissues such as runner tips and crowns. In addition, the regeneration experiment demonstrated that the addition of polyvinylpyrrolidone (PVPP) to the shoot regeneration medium could inhibit tissue browning. The gene-edited plants in which some or all of the Fvb7-1, Fvb7-2, Fvb7-3, and Fvb7-4 genes in the MLO (Mildew resistance Locus O) gene family were knocked out by CRISPR/Cas9 system were obtained by applying the plant regeneration system developed in this study.

Understanding fenpropathrin-induced pulmonary toxicity: What apoptosis, inflammation, and pyreptosis reveal analyzing cross-links at the molecular, immunohistochemical, and immunofluorescent levels

Fenpropathrin (FN), a pyrethroid has been linked to potential pulmonary toxic effects to humans via incident direct or indirect ingestion. Thus, we aimed to the investigate the underlying mechanisms of lung toxicity upon exposure to FN in the rat model, besides studying whether curcumin (CCM) and curcumin-loaded chitosan nanoformulation (CCM-Chs) can mitigate FN-induced lung damage. Six distinct groups, namely, control, CCM, CCM-Chs, FN, and CCM + FN, CCM-Chs + FN were assigned separately. The inflammatory, apoptotic, and oxidative stress states, histological, immunohistochemical, and immunofluorescence examination of different markers within the pulmonary tissue were applied. The results revealed that the FN-induced tissue damage might be caused by the oxidative stress induction and depressed antioxidant glutathione system in the lungs of rats. Furthermore, FN upregulated the expression of genes related to inflammation, and pyroptosis, and elevated the immunoreactivity of Caspase-3, tumor necrosis factor-α, vimentin, and 4-Hydroxynonenal in pulmonary tissues of FN-exposed rats compared to the control. CCM and CCM-Chs mitigated the FN-induced disturbances, while remarkably, CCM-Chs showed better potency than CCM in mitigating the FN-induced toxicity. In conclusion, this study shows the prominent preventive ability of CCM-Chs more than CCM in combatting the pulmonary toxicity induced by FN. This may be beneficial in developing therapeutic and preventive strategies against FN-induced pulmonary toxicity.

Optimization of an Efficient Direct Shoot Induction System and Histological Evaluation of Shoot Formation in Cucurbita maxima Duch

Improving plant regeneration ability and shortening regeneration time can promote the development of genetic transformation breeding technology for horticultural crops. We optimized several culture conditions, including explant type, mother plant genotype, and medium, to improve shoot formation in winter squash (Cucurbita maxima Duch.). Histological analysis of the occurrence of shoots was also carried out. The results indicate that cotyledon was the most suitable explant for inducing the shoot regeneration of winter squash. We found that ‘Jin-li’ had a shorter shoot induction time and a higher average number of shoots. The highest induction rate of 95.23% among the five lines. The average shoot induction rate of five lines was the highest (84.85%) on Murashige and Skoog (MS) medium supplemented with 2.0 mg/L 6-benzylaminopurine (6-BA) and 0.2 mg/L indole-3-acetic acid (IAA). We also found that there was an interaction between genotypes and induction media, and their interaction had a greater impact on the shoot induction rate than individual effects. Histological observation revealed that the induced shoots of winter squash cotyledons originated from subepidermal cells. We also found that the optimal medium for de novo root regeneration was 1/2 MS. We acclimatized and cultivated regenerated plants and harvested their fruits, which maintained the characteristics of mother plants. These findings lay an important foundation for further research on direct shoot regeneration and accelerate its application in winter squash genetic transformation.

Exploration of the Native Sucrose Operon Enables the Development of an Inducible T7 Expression System in Paenibacillus polymyxa.

The use of Paenibacillus polymyxa as an industrial producer is limited by the lack of suitable synthetic biology tools. In this study, we identified a native sucrose operon in P. polymyxa. Its structural and functional relationship analysis revealed the presence of multiple regulatory elements, including four ScrR-binding sites and a catabolite-responsive element (CRE). In P. polymyxa, we established a cascade T7 expression system involving an integrated T7 RNA polymerase (T7P) regulated by the sucrose operon and a T7 promoter. It enables controllable gene expression by sucrose and regulatory elements, and a 5-fold increase in expression efficiency compared with the original sucrose operon was achieved. Further deletion of SacB in P. polymyxa resulted in a 38.95% increase in the level of thermophilic lipase (TrLip) production using the cascade T7 induction system. The results highlight the effectiveness of sucrose regulation as a novel synthetic biology tool, which facilitates exploring gene circuits and enables their dynamic regulation.

Time-resolved RNA-seq analysis to unravel the in vivo competence induction by Streptococcus pneumoniae during pneumonia-derived sepsis.

Competence development in Streptococcus pneumoniae (pneumococcus) is tightly intertwined with virulence. In addition to genes encoding genetic transformation machinery, the competence regulon also regulates the expression of allolytic factors, bacteriocins, and cytotoxins. Pneumococcal competence system has been extensively interrogated in vitro where the short transient competent state upregulates the expression of three distinct phases of "early," "late," and "delayed" genes. Recently, we have demonstrated that the pneumococcal competent state develops naturally in mouse models of pneumonia-derived sepsis. To unravel the underlying adaptive mechanisms driving the development of the competent state, we conducted a time-resolved transcriptomic analysis guided by the spatiotemporal live in vivo imaging system of competence induction during pneumonia-derived sepsis. Mouse lungs infected by the serotype 2 strain D39 expressing a competent state-specific reporter gene (D39-ssbB-luc) were subjected to RNA sequencing guided by monitoring the competence development at 0, 12, 24, and, at the moribund state, >40 hours post-infection (hpi). Transcriptomic analysis revealed that the competence-specific gene expression patterns in vivo were distinct from those under in vitro conditions. There was significant upregulation of early, late, and some delayed phase competence-specific genes as early as 12 hpi, suggesting that the pneumococcal competence regulon is important for adaptation to the lung environment. Additionally, members of the histidine triad (pht) gene family were sharply upregulated at 12 hpi followed by a steep decline throughout the rest of the infection cycle, suggesting that Pht proteins participate in the early adaptation to the lung environment. Further analysis revealed that Pht proteins execute a metal ion-dependent regulatory role in competence induction.IMPORTANCEThe induction of pneumococcal competence for genetic transformation has been extensively studied in vitro but poorly understood during lung infection. We utilized a combination of live imaging and RNA sequencing to monitor the development of a competent state during acute pneumonia. Upregulation of competence-specific genes was observed as early as 12 hour post-infection, suggesting that the pneumococcal competence regulon plays an important role in adapting pneumococcus to the stressful lung environment. Among others, we report novel finding that the pneumococcal histidine triad (pht) family of genes participates in the adaptation to the lung environment and regulates pneumococcal competence induction.

SHR and SCR coordinate root patterning and growth early in the cell cycle

Precise control of cell division is essential for proper patterning and growth during the development of multicellular organisms. Coordination of formative divisions that generate new tissue patterns with proliferative divisions that promote growth is poorly understood. SHORTROOT (SHR) and SCARECROW (SCR) are transcription factors that are required for formative divisions in the stem cell niche of Arabidopsis roots1,2. Here we show that levels of SHR and SCR early in the cell cycle determine the orientation of the division plane, resulting in either formative or proliferative cell division. We used 4D quantitative, long-term and frequent (every 15 min for up to 48 h) light sheet and confocal microscopy to probe the dynamics of SHR and SCR in tandem within single cells of living roots. Directly controlling their dynamics with an SHR induction system enabled us to challenge an existing bistable model3 of the SHR–SCR gene-regulatory network and to identify key features that are essential for rescue of formative divisions in shr mutants. SHR and SCR kinetics do not align with the expected behaviour of a bistable system, and only low transient levels, present early in the cell cycle, are required for formative divisions. These results reveal an uncharacterized mechanism by which developmental regulators directly coordinate patterning and growth.