Complex Regulation Research Articles

Sequence and epigenetic variations of R2R3-MYB transcription factors determine the diversity of taproot skin and flesh colors in different cultivated types of radish (Raphanus sativus L.).

This study found that three paralogous R2R3-MYB transcription factors exhibit functional divergence among different subspecies and cultivated types in radish. Cultivated radish taproots exhibit a wide range of color variations due to unique anthocyanin accumulation patterns in various tissues. This study investigated the universal principles of taproot color regulation that developed during domestication of different subspecies and cultivated types. The key candidate genes RsMYB1 and RsMYB2, which control anthocyanin accumulation in radish taproots, were identified using bulked segregant analysis in two genetic populations. We introduced the RsMYB1-RsF3'H-RsMYB1Met genetic model to elucidate the complex and unstable genetic regulation of taproot flesh color in Xinlimei radish. Furthermore, we analyzed the expression patterns of three R2R3-MYB transcription factors in lines with different taproot colors and investigated the relationship between RsMYB haplotypes and anthocyanin accumulation in a natural population of 56 germplasms. The results revealed that three paralogous RsMYBs underwent functional divergence during radish domestication, with RsMYB1 regulating the red flesh of Xinlimei radish, and RsMYB2 and RsMYB3 regulating the red skin of East Asian big long radish (R. sativus var. hortensis) and European small radish (R. sativus var. sativus), respectively. Moreover, RsMYB1-H1, RsMYB2-H10, and RsMYB3-H6 were identified as the primary haplotypes exerting regulatory functions on anthocyanin synthesis. These findings provide an understanding of the genetic mechanisms regulating anthocyanin synthesis in radish and offer a potential strategy for early prediction of color variations in breeding programs.

UHRF2 accumulates in early G1-phase after serum stimulation or mitotic exit to extend G1 and total cell cycle length.

Ubiquitin like with PHD and ring finger domains 2 (UHRF2) regulates the cell cycle and epigenetics as a multi-domain protein sharing homology with UHRF1. UHRF1 functions with DNMT1 to coordinate daughter strand methylation during DNA replication, but UHRF2 can't perform this function, and its roles during cell cycle progression are not well defined. UHRF2 role as an oncogene vs. tumor suppressor differs in distinct cell types. UHRF2 interacts with E2F1 to control Cyclin E1 (CCNE1) transcription. UHRF2 also functions in a reciprocal loop with Cyclin E/CDK2 during G1, first as a direct target of CDK2 phosphorylation, but also as an E3-ligase with direct activity toward both Cyclin E and Cyclin D. In this study, we demonstrate that UHRF2 is expressed in early G1 following either serum stimulation out of quiescence or in cells transiting directly out of M-phase, where UHRF2 protein is lost. Further, UHRF2 depletion in G2/M is reversed with a CDK1 specific inhibitor. UHRF2 controls expression levels of cyclins and CDK inhibitors and controls its own transcription in a negative-feedback loop. Deletion of UHRF2 using CRISPR/Cas9 caused a delay in passage through each cell cycle phase. UHRF2 loss culminated in elevated levels of cyclins but also the CDK inhibitor p27KIP1, which regulates G1 passage, to reduce retinoblastoma phosphorylation and increase the amount of time required to reach G1/S passage. Our data indicate that UHRF2 is a central regulator of cell-cycle pacing through its complex regulation of cell cycle gene expression and protein stability.

Relish-mediated C2H2 zinc finger protein IMZF restores Drosophila immune homeostasis via inhibiting the transcription of Imd/Tak1.

The dysregulation of intensity and duration in innate immunity can result in detrimental effects on the body, emphasizing the crucial need for precise regulation. However, the intricate and accurate nature of innate immunity implies the existence of numerous undiscovered innate immunomodulators, particularly transcription factors. In this study, we have identified a Drosophila C2H2 zinc finger protein CG18262, named Immune-mediated Zinc Finger protein (IMZF), capable of suppressing immune responses of Imd pathway. Mechanistically, IMZF serves as a transcription factor that represses the expression of Imd and Tak1. Intriguingly, our findings also reveal that Relish, an NF-κB transcription factor, positively regulates the expression of IMZF, consequently inhibiting the activation of Imd and Tak1 to prevent an exaggerated immune response. Additionally, we have elucidated the pivotal role played by the Relish-IMZF-Imd/Tak1 axis in restoring immune homeostasis of Drosophila Imd pathway. In summary, our findings not only unveil a novel C2H2 zinc finger immunoregulatory transcription factor, IMZF, along with its specific mechanism of immune regulation, but also shed light on the dual functionality of Relish in different stages of the immune response by modulating distinct effectors. This discovery provides new insights and enlightenment into the complex regulation of Drosophila innate immunity.

The mutated cytoplasmic fragile X mental retardation 1 (FMR1)-interacting protein 2 (CYFIP2 S968F) regulates cocaine-induced reward behaviour and plasticity in the nucleus accumbens.

Cytoplasmic fragile X mental retardation 1 (FMR1)-interacting protein 2 (CYFIP2), as a component of the Wiskott-Aldrich syndrome protein family verprolin-homologous protein (WAVE) regulatory complex, is involved in actin polymerization, contributing to neuronal development and structural plasticity. Mutating serine-968 to phenylalanine (S968F) in CYFIP2 causes an altered cocaine response in mice. The neuronal mechanisms underlying this response remain unknown. We performed cocaine reward-related behavioural tests and examined changes in synaptic protein phenotypes and neuronal morphology in the nucleus accumbens (NAc), using CYFIP2 S968F knock-in mice to investigate the role of CYFIP2 in regulating cocaine reward. CYFIP2 S968F mutation attenuated cocaine-induced behavioural sensitization and conditioned place preference. Cocaine-induced c-Fos was not observed in the NAc of CYFIP2 S968F knock-in mice. However, c-Fos induction was still evident in the medial prefrontal cortex (mPFC). CYFIP2 S968F mutation altered cocaine-associated CYFIP2 signalling, glutamatergic protein expression and synaptic density in the NAc following cocaine exposure. To further determine the role of CYFIP2 in NAc neuronal activity and the mPFC projecting to the NAc activity-mediating reward response, we used optogenetic tools to stimulate the NAc or mPFC-NAc pathway and observed that optogenetic activation of the NAc or mPFC-NAc pathway induced reward-related behaviours. This effect was not observed in the S968F mutation in CYFIP2. These results suggest that CYFIP2 plays a role in controlling cocaine-mediated neuronal function and structural plasticity in the NAc, and that CYFIP2 could serve as a target for regulating cocaine reward.

A role for the C. elegans Argonaute protein CSR-1 in small nuclear RNA 3' processing.

The Integrator is a multi-subunit protein complex that catalyzes the maturation of snRNA transcripts via 3' cleavage, a step required for snRNA incorporation with snRNP for spliceosome biogenesis. Here we developed a GFP based in vivo snRNA misprocessing reporter as a readout of Integrator function and performed a genome-wide RNAi screen for Integrator regulators. We found that loss of the Argonaute encoding csr-1 gene resulted in widespread 3' misprocessing of snRNA transcripts that is accompanied by a significant increase in alternative splicing. Loss of the csr-1 gene down-regulates the germline expression of Integrator subunits 4 and 6 and is accompanied by a reduced protein translation efficiency of multiple Integrator catalytic and non-catalytic subunits. Through isoform and motif mutant analysis, we determined that CSR-1's effect on snRNA processing is dependent on its catalytic slicer activity but does not involve the CSR-1a isoform. Moreover, mRNA-sequencing revealed high similarity in the transcriptome profile between csr-1 and Integrator subunit knockdown via RNAi. Together, our findings reveal CSR-1 as a new regulator of the Integrator complex and implicate a novel role of this Argonaute protein in snRNA 3' processing.

The capsular polysaccharide obstructs wall teichoic acid functions in Staphylococcus aureus.

The cell envelope of Staphylococcus aureus contains two major secondary cell wall glycopolymers: capsular polysaccharide (CP) and wall teichoic acid (WTA). Both the CP and the WTA are attached to the cell wall and play distinct roles in S. aureus colonization, pathogenesis, and bacterial evasion of host immune defenses. We aimed to investigate whether CP interferes with WTA-mediated properties. Strains with natural heterogeneous expression of CP, strains with homogeneous high CP expression and CP-deficient strains were compared to WTA deficient controls regarding WTA dependent phage binding, cell adhesion, IgG deposition, and virulence in vivo. WTA-mediated phage adsorption, specific antibody deposition and cell adhesion were negatively correlated with CP expression. WTA, but not CP, enhanced the bacterial burden in a mouse abscess model, while CP overexpression resulted in intermediate virulence in vivo. CP protects the bacteria from WTA-dependent opsonization and phage binding. This protection comes at the cost of diminished adhesion to host cells. The highly complex regulation and mostly heterogeneous expression of CP has probably evolved to ensure the survival and optimal physiological adaptation of the bacterial population as a whole.

TEX13B is essential for metabolic reprogramming during germ cell differentiation.

What is the functional significance of Tex13b in male germ cell development and differentiation? Tex13b regulates male germ cell differentiation by metabolic reprogramming during spermatogenesis. Studies in mice and humans suggest that TEX13B is a transcription factor and is exclusively expressed in germ cells. We sequenced the coding regions of TEX13B in 628 infertile men and 427 ethnically matched fertile control men. Further, to identify the molecular function of Tex13b, we created a Tex13b knockout and conditional overexpression system in GC-1spg (hereafter, GC-1) cells. Our recent exome sequencing study identified novel candidate genes for male infertility. TEX13B was found to be one of the potential candidates, hence we explored the role of TEX13B in male infertility within a large infertile case-control cohort. We performed functional analyses of Tex13b in a GC-1 cell line using CRISPR-Cas9. We differentially labelled the cell proteins by stable isotope labelling of amino acids in cell culture (SILAC) and performed mass spectrometry-based whole-cell proteomics to identify the differential protein regulation in knockout cells compared to wild-type cells. We found that Tex13b knockout leads to downregulation of the OXPHOS complexes and upregulation of glycolysis genes, which was further validated by western blotting. These results were further confirmed by respirometry analysis in Tex13b knockout cells. Further, we also performed a conditional overexpression of TEX13B in GC-1 cells and studied the expression of OXPHOS complex proteins by western blotting. We identified a rare variant, rs775429506 (p.Gly237Glu), exclusively in two non-obstructive-azoospermia (NOA) men, that may genetically predispose these men for infertility. Further, we demonstrated that Tex13b functions in the transcription regulation of OXPHOS complexes. N/A. We examined the function of Tex13b in GC-1 in vitro by knocking out and conditional overexpression, for understanding the function of Tex13b in germ cells. Unfortunately, this could not be replicated in either an animal model or in patient-derived tissue due to the non-availability of an animal model or patient's testis biopsies. This study identified that Tex13b plays an important role in male germ cell development and differentiation. The findings of this study would be useful for screening infertile males with spermatogenic failure and counselling them before the implementation of assisted reproduction technique(s). Funding was provided by the Council of Scientific and Industrial Research (CSIR) under the network project (BSC0101 and MLP0113) and SERB, the Department of Science and Technology, Government of India (J C Bose Fellowship: JCB/2019/000027). The authors do not have any competing interest.

An Analysis of How Logistics Integration Affects Organizational Performance in Kenya's Clearing and Forwarding Sector. A Structural Equation Modelling Approach.

The performance of logistics service providers in Kenya is marred by poor customer satisfaction, stemming from delays experienced in port clearance, disintegrated clearance systems, complex regulations, inefficient document processing and lack of integrated information systems. This study examined the effect of logistics integration on organizational performance among the Clearing and Forwarding firms in Kenya. It adopted a cross-sectional design and collected primary data through structured questionnaires. Simple and stratified random sampling were used to select respondents and ethical approvals obtained before collecting data using structured questionnaires. Data were collected from 298 employees from the operations, customer service, and finance departments. Using SPSS and SPSS AMOS 29, the study used descriptive statistics and structural equation modelling. The study findings revealed a disparity in the level of logistics integration among different Clearing and Forwarding firms in Kenya. This variance in logistics integration significantly impacts the overall organizational performance of these firms. From SEM analysis, the study established that logistics integration has a positive and significant effect (Path coefficient = 0.791, C.R = 5.664, P-value < 0.001) on the organizational performance of C&F companies in Kenya. The study concludes that enhancing logistics integration will lead to a substantial improvement in the organizational performance of C&F companies in Kenya. Thus, managers responsible for strategy should aim to collaborate with strategic partners such as state corporations managing ports and customs processes (KRA and Kenya Ports Authority), shipping lines, transportation companies, and other key stakeholders in the C&F supply chain in integrating logistics systems and processes.

Comparison of CcrM-dependent methylation in Caulobacter crescentus and Brucella abortus by nanopore sequencing.

Bacteria rely on DNA methylation for restriction-modification systems and epigenetic control of gene expression. Here, we use direct detection of methylated bases by nanopore sequencing to monitor global DNA methylation in Alphaproteobacteria, where use of this technique has not yet been reported. One representative of this order, Caulobacter crescentus, relies on DNA methylation to control cell cycle progression, but it is unclear whether other members of this order, such as Brucella abortus, depend on the same systems. We addressed these questions by first measuring CcrM-dependent DNA methylation in Caulobacter and showing excellent correlation between nanopore-based detection and previously published results. We then directly measure the impact of Lon-mediated CcrM degradation on the epigenome, verifying that loss of Lon results in pervasive methylation. We also show that the AlkB demethylase has no global impact on DNA methylation during normal growth. Next, we report on the global DNA methylation in B. abortus for the first time and find that CcrM-dependent methylation is reliant on Lon but impacts the two chromosomes differently. Finally, we explore the impact of the MucR transcription factor, known to compete with CcrM methylation, on the Brucella methylome and share the results with a publicly available visualization package. Our work demonstrates the utility of nanopore-based sequencing for epigenome measurements in Alphaproteobacteria and reveals new features of CcrM-dependent methylation in a zoonotic pathogen.IMPORTANCEDNA methylation plays an important role in bacteria, maintaining genome integrity and regulating gene expression. We used nanopore sequencing to directly measure methylated bases in Caulobacter crescentus and Brucella abortus. In Caulobacter, we showed that stabilization of the CcrM methyltransferase upon loss of the Lon protease results in prolific methylation and discovered that the putative methylase AlkB is unlikely to have a global physiological effect. We measured genome-wide methylation in Brucella for the first time, revealing a similar role for CcrM in cell-cycle methylation but a more complex regulation by the Lon protease than in Caulobacter. Finally, we show how the virulence factor MucR impacts DNA methylation patterns in Brucella.

Genome-wide profiling of angiogenic cis-regulatory elements unravels cis-regulatory SNPs for vascular abnormality

Angiogenesis is extensively involved in embryonic development and requires complex regulation networks, whose defects can cause a variety of vascular abnormalities. Cis-regulatory elements control gene expression at all developmental stages, but they have not been studied or profiled in angiogenesis yet. In this study, we exploited public DNase-seq and RNA-seq datasets from a VEGFA-stimulated in vitro angiogenic model, and carried out an integrated analysis of the transcriptome and chromatin accessibility across the entire process. Totally, we generated a bank of 47,125 angiogenic cis-regulatory elements with promoter (marker by H3K4me3) and/or enhancer (marker by H3K27ac) activities. Motif enrichment analysis revealed that these angiogenic cis-regulatory elements interacted preferentially with ETS family TFs. With this tool, we performed an association study using our WES data of TAPVC and identified rs199530718 as a cis-regulatory SNP associated with disease risk. Altogether, this study generated a genome-wide bank of angiogenic cis-regulatory elements and illustrated its utility in identifying novel cis-regulatory SNPs for TAPVC, expanding new horizons of angiogenesis as well as vascular abnormality genetics.

The histone methyltransferase SUV420H2 regulates brown and beige adipocyte thermogenesis.

Activation of brown adipose tissue (BAT) thermogenesis increases energy expenditure and alleviates obesity. Here we discover that histone methyltransferase suppressor of variegation 4-20 homolog 2 (Suv420h2) expression parallels that of Ucp1 in brown and beige adipocytes and that Suv420h2 knockdown significantly reduces, whereas Suv420h2 overexpression significantly increases Ucp1 levels in brown adipocytes. Suv420h2 knockout (H2KO) mice exhibit impaired cold-induced thermogenesis and are prone to diet-induced obesity. In contrast, mice with specific overexpression of Suv420h2 in adipocytes display enhanced cold-induced thermogenesis and are resistant to diet-induced obesity. Further study shows that Suv420h2 catalyzes H4K20 trimethylation at eukaryotic translation initiation factor 4E-binding protein 1 (4e-bp1) promoter, leading to down-regulated expression of 4e-bp1, a negative regulator of the translation initiation complex. This in turn up-regulates PGC1α protein levels, which is associated with increased expression of thermogenic program. We conclude that Suv420h2 is a key regulator of brown/beige adipocyte development and thermogenesis.

NtG3BPL1 confers resistance to chilli veinal mottle virus through promoting the degradation of 6K2 in tobacco.

The RNA regulatory network is a complex and dynamic regulation in plant cells involved in mRNA modification, translation, and degradation. Ras-GAP SH3 domain-binding protein (G3BP) is a scaffold protein for the assembly of stress granules (SGs) and is considered an antiviral component in mammals. However, the function of G3BP during virus infection in plants is still largely unknown. In this study, four members of the G3BP-like proteins (NtG3BPLs) were identified in Nicotiana tabacum and the expression levels of NtG3BPL1 were upregulated during chilli veinal mottle virus (ChiVMV) infection. NtG3BPL1 was localized in the nucleus and cytoplasm, forming cytoplasmic granules under transient high-temperature treatment, whereas the abundance of cytoplasmic granules was decreased under ChiVMV infection. Overexpression of NtG3BPL1 inhibited ChiVMV infection and delayed the onset of symptoms, whereas knockout of NtG3BPL1 promoted ChiVMV infection. In addition, NtG3BPL1 directly interacted with ChiVMV 6K2 protein, whereas 6K2 protein had no effect on NtG3BPL1-derived cytoplasmic granules. Further studies revealed that the expression of NtG3BPL1 reduced the chloroplast localization of 6K2-GFP and the NtG3BPL1-6K2 interaction complex was localized in the cytoplasm. Furthermore, NtG3BPL1 promoted the degradation of 6K2 through autophagy pathway, and the accumulation of 6K2 and ChiVMV was affected by autophagy activation or inhibition in plants. Taken together, our results demonstrate that NtG3BPL1 plays a positive role in tobacco resistance against ChiVMV infection, revealing a novel mechanism of plant G3BP in antiviral strategy.

The immune cells have complex causal regulation effects on cancers

BackgroundThere was a large body of evidence linking immune cells to cancer risk. However, the causal relationship between immune cells, cancer, and what genes play an important role is unclear. MethodsIn this study, we performed comprehensive two-sample Mendelian randomization analysis (TSMR) to determine the causal relationship between immune cells and common cancers. We also performed Multimarker Analysis of Genomic Annotation (MAGMA) on immune cells causally associated with cancer to identify their relevant genes and used data summary-based MR (SMR) analysis to investigate the causal relationship between their gene expression, methylation, and cancer, and further used drug prediction and molecular docking to validate the medicinal value of the targets. Finally, reverse TSMR analysis was performed on cancer and immune cells to rule out reverse causality. ResultsAfter FDR correction (PFDR < 0.05), the results showed that 2 immune cells were associated with lung cancer risk, and 1 immune cell was significantly associated with pancreatic cancer risk. The expression of OSBPL10, CHD4, SMDT1, PHETA2, and NAGA was positively and causally related to the risk of lung cancer by SMR analysis and HEIDI test. We also found that increased expression of ANP32E decreased the risk of pancreatic cancer and that the methylation level of OSBPL10, CHD4, SULF2, CENPM, and CYP2D6 had a causal association with lung cancer. The methylation level of FCGR3A was causally associated with pancreatic cancer. The results of molecular docking indicated a strong affinity between the drugs and proteins that possessed existing structural information. ConclusionThis data-driven Mendelian randomization (MR) study demonstrates the causal role of immune cells in cancers. In addition, this study identifies candidate genes that may be potential anti-cancer drug targets.

Investigator Participation in Clinical Research: An Examination of Barriers and Facilitators in the Indian Clinical Trials Settings

Background: This study aims to improve clinical research by identifying the barriers and facilitators for investigators conducting trials. We explore investigator demographics, challenges faced, and factors promoting engagement. Additionally, we examine how investigators’ background influences their experiences with both barriers and facilitators, ultimately aiming to foster a patient-centric clinical trial ecosystem. Methods: A quantitative, exploratory study investigated barriers and facilitators in clinical trials among 132 investigators in India. A self-designed questionnaire captured experiences with challenges, strategies, and facilitators. Pilot testing ensured tool validity and reliability. Ethical considerations were addressed, and data analysis will identify key factors impacting investigator engagement. This study aims to improve the clinical research ecosystem in India. Results: The study aims to identify the facilitators and barriers experienced by the research investigators. An online questionnaire survey was used in this cross-sectional investigation. The study's population consists of 132 respondents, with 68 (51.5%) males and 64 (48.5%) females. Clinical Research Coordinators (CRCs) (41.7%), Research Associates (RA’s) (27.3%), Supporting staff (19.7%), Co-Investigators (Co-I) (6.8%), and Principal Investigators (PIs) (4.5%) which were the majority of respondents. The most common barriers found among the principal investigators regarding the limited resources (p=0.014) towards the clinical trials. Inadequate training (p=0.461), Complex regulations (p=0.625), and Communication barriers (p=0.853) did not seem like barriers to the clinical research staff. Whereas some of the staff faced barriers (p=0.118) with the complex regulations and Inadequate training on the other hand most of the respondents considered the same statements as facilitators (p=0.963) with their daily working activities. Conclusion: The Barriers and Facilitators experienced by the clinical investigators, It was found that there is no difference in viewpoint. As per the results, Research associates agreed more with the facilitators and the principal investigators faced barriers.

Dysfunction of Avo3, an essential component of target of rapamycin complex 2, induces ubiquitin-proteasome-dependent downregulation of Avo2 in Saccharomyces cerevisiae

The ubiquitin-proteasome system (UPS) plays a key role in maintaining cellular protein homeostasis and participates in modulating various cellular functions. Target of rapamycin (TOR), a highly conserved Ser/Thr kinase found across species from yeasts to humans, forms two multi-protein complexes, TORC1 and TORC2, to orchestrate cellular processes crucial for optimal growth, survival, and stress responses. While UPS-mediated regulation of mammalian TOR complexes has been documented, the ubiquitination of yeast TOR complexes remains largely unexplored. Here we report a functional interplay between the UPS and TORC2 in Saccharomyces cerevisiae. Using avo3-2ts, a temperature-sensitive mutant of the essential TORC2 component Avo3 exhibiting TORC2 defects at restrictive temperatures, we obtained evidence for UPS-dependent protein degradation and downregulation of the TORC2 component Avo2. Our results established the involvement of the E3 ubiquitin ligase Ubr1 and its catalytic activity in mediating Avo2 degradation in cells with defective Avo3. Coimmunoprecipitation revealed the interaction between Avo2 and Ubr1, indicating Avo2 as a potential substrate of Ubr1. Furthermore, depleting Ubr1 rescued the growth of avo3-2ts cells at restrictive temperatures, suggesting an essential role of Avo2 in sustaining cell viability under heat stress and/or TORC2 dysfunction. This study uncovers a role of UPS in yeast TORC2 regulation, highlighting the impact of protein degradation control on cellular signaling.

The complex regulation of Slo1 potassium channels from a structural perspective.

Fast and regulated potassium efflux by Slo1 channels is crucial in many tissues in animals including neurons, the kidney and smooth muscle. During the last decade, structures have revealed many details about the gating mechanism and regulation of these large and complex molecular machines. This review summarizes these findings and the current knowledge about the intricate regulation of these important channels. Slo1 integrates sensing of the membrane potential via a voltage-sensor domain that undergoes subtle but significant structural rearrangements with a calcium-induced expansion of parts of the intracellular gating ring. Together, these two signals synergistically lead to changes in the conformation and chemical nature of the pore domain, allowing potassium ions to be translocated. In many native tissues, Slo1 channels are assembled with at least three classes of auxiliary subunits that change the gating kinetics or allow the channel to open also in absence of one of the two signals. Finally, Slo1 is inhibited, activated or deregulated by natural toxins and synthetic compounds, underlining the importance of the channel for the organism and as a potential target for drugs and other molecules.

Identifying key barriers to joining an energy community using AHP

Energy communities (ECs) offer a promising solution for achieving sustainable and decentralized energy systems. However, the successful establishment and operation of ECs requires overcoming barriers that can hinder stakeholder participation. Existing research has primarily focused on incentives and motivations to join ECs, thus neglecting a comprehensive understanding of the key barriers affecting EC stakeholders in European Union (EU) countries. This paper aims to fill this research gap by identifying and ranking the barriers to joining ECs in the EU context with focus on Spain and Italy. To accomplish this, a framework of barriers was developed based on 20 in-depth interviews with diverse stakeholders of established ECs and a survey (n = 56). The barriers identified were categorized into four types: (a) financial, (b) regulatory and bureaucratic, (c) technical and practical, and (d) social and cultural. The analytical hierarchical process (AHP) methodology was employed to estimate and rank these barriers. The findings highlight that the most significant barrier categories are regulatory and bureaucratic, and financial. Specifically, regulatory complexity and legal limitations emerge as the top-ranked barriers among the obstacles identified to joining ECs by research participants.

INVESTIGATING BARRIERS IN GREEN SUPPLY CHAIN MANAGEMENT

The Green Supply Chain Management (GSCM) has emerged in today’s scenario in response to the growing concern of environmental sustainability. Green Supply Chain is a term that has it’s importance in the entire supply chain starting from the designing of the product to raw material procurement to manufacturing of the product to distribution and until the end-of-life of the product. It’s main aim is to minimize the negative impact of the supply chain activities and also to maximize the efficiency of resources along with reduction of the waste generation. There are a number of challenges in implementing green practices in the organization, which include the need for cooperation, collaboration and coordination among the partners of supply chain and also the need to identify the cost-effective green practices and technologies. There are a number of strategies which have been proposed to address the challenges faced. These include packaging which is eco-friendly, the concept of reverse logistics etc. The organizations which adopt the green supply chain practices in their organizations, gain various types of benefits from it. These benefits include savings in the costs as there is reduction in the consumption and also waste generation. The brand image of the organization is enhanced by the adoption of green practices and there is increase in the customer loyalty etc. Companieswhich use green supply chain management in their supply chain include Johnson and Johnson, Ford, Adidas, Nestle, Coca-Cola, Tesla etc. The adoption of green supply chain management (GSCM) faces multiple challenges including financial barriers, lack of awareness and supplier engagement, and regulatory complexity, while cultural resistance and technological limitations also impede its implementation. Success relies on top management support and overcoming these hurdles to develop sustainable practices across the entire supply chain.

Multiple pathways impact the swarming motility of Pseudomonas fluorescens Pf0-1.

Swarming motility in pseudomonads typically requires both a functional flagellum and the production/secretion of a biosurfactant. Published work has shown that the wild-type Pseudomonas fluorescens Pf0-1 is swarming deficient due to a point mutation in the gacA gene, which until recently was thought to inactivate rather than attenuate the Gac/Rsm pathway. As a result, little is known about the underlying mechanisms that regulate swarming motility by P. fluorescens Pf0-1. Here, we demonstrate that a ΔrsmA ΔrsmE ΔrsmI mutant, which phenotypically mimics Gac/Rsm pathway overstimulation, is proficient at swarming motility. RsmA and RsmE appear to play a key role in this regulation. Transposon mutagenesis of the ΔrsmA ΔrsmE ΔrsmI mutant identified multiple factors that impact swarming motility, including pathways involved in flagellar synthesis and biosurfactant production/secretion. We find that loss of genes linked to biosurfactant Gacamide A biosynthesis or secretion impacts swarming motility, as does loss of the alternative sigma factor FliA, which results in a defect in flagellar function. Collectively, these findings provide evidence that P. fluorescens Pf0-1 can swarm if the Gac/Rsm pathway is activated, highlight the regulatory complexity of swarming motility in this strain, and demonstrate that the cyclic lipopeptide Gacamide A is utilized as a biosurfactant for swarming motility.IMPORTANCESwarming motility is a coordinated process that allows communities of bacteria to collectively move across a surface. For P. fluorescens Pf0-1, this phenotype is notably absent in the parental strain, and to date, little is known about the regulation of swarming in this strain. Here, we identify RsmA and RsmE as key repressors of swarming motility via modulating the levels of biosurfactant production/secretion. Using transposon mutagenesis and subsequent genetic analyses, we further identify potential regulatory mechanisms of swarming motility and link Gacamide A biosynthesis and transport machinery to swarming motility.

Evaluation of Supply Chain Management Challenges of multinational companies in West Africa under the ECOWAS Trade Liberalization Scheme (ETLS)

This paper examined the Supply Chain Management challenges of multinational companies in West Africa under the ECOWAS trade liberalization scheme. The objective of the study includes to identify the various ECOWAS's regional integration protocols, Assess the level of implementation of ETLS among members state and evaluation of the various negative factors affecting full implementation of ETL, among others. The methodologies adopted includes Qualitative and quantitative method of data collection including literature review, expert interview, survey and case studies over a sample of 150 multinational companies (MNC). The results reveal that Complex and Non-Transparent Regulations exists among the MNC in the West African region, existence of Inadequate Infrastructure and Logistics, Non-Harmonized Trade Facilitation Measures as well as weak institutional co-ordination and enforcements. The study concludes that ETLS presents both opportunities and challenges for MNCs operating in West Africa and recommends among others the Simplification and Harmonization of Trade Regulations, investment in infrastructures and logistics as well as strengthening of institutional co-ordinations and enforcements.