Components Of Pathway Research Articles

Oroxylin A-induced Trained Immunity Promotes LC3-associated Phagocytosis in Macrophage in Protecting Mice Against Sepsis.

Sepsis is defined as a dysregulated host response to infection that leads to multiorgan failure. Innate immune memory, i.e., "trained immunity", can result in stronger immune responses and provide protection against various infections. Many biological agents, including β-glucan, can induce trained immunity, but these stimuli may cause uncontrolled inflammation. Oroxylin A (OA) is an active flavonoid compound that is derived from Scutellaria baicalensis.OA is an agonist for inducing trained immunity in vivo and in vitro, and β-glucan was used as a positive control. The protective effects of OA-induced trained immunity were evaluated in mouse models that were established by either lipopolysaccharide (LPS) administration or caecal ligation and puncture (CLP). The expression of inflammatory factors and signaling pathway components involved in trained immunity was evaluated in vitro using qRT‒PCR, western blotting (WB) and enzyme-linked immunosorbent assay (ELISA). Flow cytometry and confocal microscopy were used to examine reactive oxygen species (ROS) levels and phagocytosis in trained macrophages. A PCR array was used to screen genes that were differentially expressed in trained macrophages.Here, we revealed that OA alleviated sepsis via trained immunity. OA-treated macrophages displayed increased glycolysis and mTOR phosphorylation, and mTOR inhibitors suppressed OA-induced trained immunity by effectively reprogramming macrophages. The PCR array revealed key genes in the mTOR signaling pathway in OA-treated macrophages. Furthermore, OA targeted the Dectin-1-syk axis to promote LC3-associated phagocytosis (LAP) by trained macrophages, thereby enhancing the ability of these macrophages to protect against infection. This ability could be transferred to a new host via the adoptive transfer of peritoneal macrophages.This study is the first to provide new insights into the potential of OA-induced trained immunity to be used as a strategy to protect mice against sepsis by promoting LAP by macrophages.

Hypothesis: evidence that the PRS gene products of Saccharomyces cerevisiae support both PRPP synthesis and maintenance of cell wall integrity

The gene products of PRS1-PRS5 in Saccharomyces cerevisiae are responsible for the production of PRPP (5-phospho-D-ribosyl-α-1-pyrophosphate). However, it has been demonstrated that they are also involved in the cell wall integrity (CWI) signalling pathway as shown by protein–protein interactions (PPIs) with, for example Slt2, the MAP kinase of the CWI pathway. The following databases: SGD, BioGRID and Hit Predict, which collate PPIs from various research papers, have been scrutinized for evidence of PPIs between Prs1-Prs5 and components of the CWI pathway. The level of certainty in PPIs was verified by interaction scores available in the Hit Predict database revealing that well-documented interactions correspond with higher interaction scores and can be graded as high confidence interactions based on a score > 0.28, an annotation score ≥ 0.5 and a method-based high confidence score level of ≥ 0.485. Each of the Prs1-Prs5 polypeptides shows some degree of interaction with the CWI pathway. However, Prs5 has a vital role in the expression of FKS2 and Rlm1, previously only documented by reporter assay studies. This report emphasizes the importance of investigating interactions using more than one approach since every method has its limitations and the use of different methods, as described herein, provides complementary experimental and statistical data, thereby corroborating PPIs. Since the experimental data described so far are consistent with a link between PRPP synthetase and the CWI pathway, our aim was to demonstrate that these data are also supported by high-throughput bioinformatic analyses promoting our hypothesis that two of the five PRS-encoding genes contain information required for the maintenance of CWI by combining data from our targeted approach with relevant, unbiased data from high-throughput analyses.

RRP8, associated with immune infiltration, is a prospective therapeutic target in hepatocellular carcinoma

BackgroundRibosomal RNA Processing 8 (RRP8) is a nucleolar Rossman fold-like methyltransferase that exhibits increased expression in many malignant tumours. However, the role of RRP8 in hepatocellular carcinoma (HCC) is still uncertain. We explored the relationships between RRP8 and prognosis and immune infiltration, as well as the putative pathological function and mechanism of RRP8 in HCC.MethodsAnalysis of RRP8 expression across cancers was performed by using multiple databases. Associations between RRP8 expression and clinicopathological factors were further examined. Gene enrichment analysis was used to identify various putative biological activities and regulatory networks of RRP8 in HCC. The relationship between RRP8 expression and immune infiltration was confirmed by single-sample gene set enrichment analysis (ssGSEA). Univariate and multivariate Cox regression analyses were conducted to assess the impact of clinical variables on patient outcomes. Furthermore, a nomogram was constructed to estimate survival probability based on multivariate Cox regression analysis. Functional validation of RRP8 in HCC was performed with two different systems: doxycycline-inducible shRNA knockdown and CRISPR-Cas9 knockout.ResultsRRP8 was markedly overexpressed in HCC clinical specimens compared to adjacent normal tissues. Further analysis demonstrated that RRP8 was directly connected to multiple clinical characteristics and strongly associated with various immune markers in HCC. Moreover, elevated RRP8 expression indicated an unfavourable prognosis. Our functional studies revealed that both knockdown and knockout of RRP8 dramatically attenuated liver cancer cells to proliferate and migrate. Knockout of RRP8 decreased the phosphorylation of MEK1/2 and β-catenin-(Y654) signalling pathway components; downregulated downstream signalling effectors, including Cyclin D1 and N-cadherin; and upregulated E-cadherin.ConclusionsRRP8 is strongly implicated in immune infiltration and could be a potential therapeutic target in HCC.

Virus-vectoring thrips regulate the excessive multiplication of tomato spotted wilt virus using their antiviral immune responses.

The tomato spotted wilt virus (TSWV) is a member of the Tospoviridae family and has an negative/ambisense single-stranded RNA genome. Frankliniella occidentalis and F. intonsa are known to be dominant pests in Capsicum annuum (hot pepper) and can cause damage to the plant either directly by feeding, or indirectly by transmitting TSWV in a persistent and propagative manner, resulting in serious economic damage. This study compared the immune responses of two different thrips species against TSWV infection by transcriptome analysis, which then allowed the assessment of antiviral responses using RNA interference (RNAi). Both adult thrips shared about 90 % of the transcripts in non-viruliferous conditions. Most signal components of the immune pathways were shared by these two thrips species, and their expression levels fluctuated differentially in response to TSWV infection at early immature stages. The functional assays using RNAi treatments indicated that the Toll and JAK/STAT pathways were associated with the antiviral responses, but the IMD pathway was not. The upregulation of dorsal switch protein one supported its physiological role in recognizing TSWV infection and triggering the eicosanoid biosynthetic pathway, which mediates melanization and apoptosis in thrips. In addition, the signal components of the RNAi pathways fluctuated highly after TSWV infection. Individual RNAi treatments specific to the antiviral signalling and response components led to significant increases in the TSWV amount in the thrips, causing virus-induced mortality. These findings suggest that immune signalling pathways leading to antiviral responses are operating in the thrips to regulate TSWV litres to prevent a fatal viral overload. This study also indicates the differential antiviral responses between the TSWV-transmitting F. occidentalis and F. intonsa.

Mammalian IRE1α dynamically and functionally coalesces with stress granules.

Upon endoplasmic reticulum (ER) stress, activation of the ER-resident transmembrane protein kinase/endoribonuclease inositol-requiring enzyme 1 (IRE1) initiates a key branch of the unfolded protein response (UPR) through unconventional splicing generation of the transcription factor X-box-binding protein 1 (XBP1s). Activated IRE1 can form large clusters/foci, whose exact dynamic architectures and functional properties remain largely elusive. Here we report that, in mammalian cells, formation of IRE1α clusters is an ER membrane-bound phase separation event that is coupled to the assembly of stress granules (SGs). In response to different stressors, IRE1α clusters are dynamically tethered to SGs at the ER. The cytosolic linker portion of IRE1α possesses intrinsically disordered regions and is essential for its condensation with SGs. Furthermore, disruption of SG assembly abolishes IRE1α clustering and compromises XBP1 mRNA splicing, and such IRE1α-SG coalescence engenders enrichment of the biochemical components of the pro-survival IRE1α-XBP1 pathway during ER stress. Our findings unravel a phase transition mechanism for the spatiotemporal assembly of IRE1α-SG condensates to establish a more efficient IRE1α machinery, thus enabling higher stress-handling capacity.

178 Effects of L-glutamate and L-aspartate supplementation on systemic and intestinal immune responses of weaned piglets challenged with F18 enterotoxigenic Escherichia coli challenge

Abstract L-glutamate (Glu) and L-aspartate (Asp) are pivotal components in the cellular metabolic pathways and immune regulation of pigs. This study aimed to investigate the effects of Glu and Asp on systemic and intestinal immune responses of weaned piglets challenged with an enterotoxigenic Escherichia coli (ETEC) F18. A total of 49 weaned pigs [initial body weight (BW) = 8.18 ± 1.54 kg] were randomly assigned to one of seven treatments, with seven piglets per treatment. The treatments included negative control (NC) and positive control (PC) that were fed with control diet without or with F18 enterotoxigenic Escherichia coli (ETEC) challenge, respectively. The other five dietary treatments were supplemented with either 1% or 2% Glu or Asp, or 50 mg/kg of Carbadox, and were challenged with ETEC. All pigs except for NC were challenged with F18 ETEC orally for three consecutive days at the dose of 1010 CFU•dose-1•d-1. The study period lasted for 3 wk, with a 7-d adaptation period and 14 d post-inoculation (PI). Blood samples were collected on d 0 before ETEC inoculation, and on d 2, 5, and 14 PI to analyze systemic inflammatory markers by the enzyme-linked immunosorbent assays. On d 14 PI, intestinal mucosa from jejunum and ileum were collected to analyze the mRNA expression of inflammatory cytokines by quantitative polymerase chain reaction. All data were analyzed with ANOVA using the PROC MIXED of SAS with pig as the experimental unit. Among all treatments, pigs fed with carbadox had the least (P < 0.05) serum TNF-α, haptoglobin, and C-reactive protein throughout the experiment. Pigs supplemented with 1% Glu had similar concentrations of TNF-α, haptoglobin, and C-reactive protein, while pigs fed with 2% Asp had similar serum haptoglobin compared with pigs fed with carbadox. Compared with PC, supplementing Glu and Asp reduced (P < 0.05) the mRNA expression of IL12 in jejunal mucosa. Pigs fed with carbadox tended to have greater (P < 0.10) expression of IFNG in ileal mucosa than pigs in PC, while no difference was observed in IFNG expression among 1% Glu, 1% or 2% Asp, and carbadox groups. In addition, pigs fed with 1% Glu or 2% Asp had greater (P < 0.05) expression of IL12 in ileal mucosa than pigs in PC. However, dietary treatments did not affect the gene expression of TNFA, IL1B and TGFB in jejunal and ileal mucosa of pigs. In conclusion, results of the present study indicate that dietary supplementation of 1% Glu or 2% Asp affected both systemic and intestinal immune responses of ETEC challenged weaned pigs with similar trend as the supplementation of carbadox.

Sensitivity and activation of endoplasmic reticulum stress response and apoptosis in the perinatal sheep heart.

We determined in vitro sensitivity to thapsigargin in sheep cardiomyocytes from four perinatal ages. In utero cardiac activation of endoplasmic reticulum (ER) stress and apoptotic pathways was determined at these same ages. Thapsigargin-induced phosphorylation of EIF2A decreased 72% between 135 and 143 dGA (P=0.0096) and remained low at 1 dPN (P=0.0080). Conversely, thapsigargin-induced caspase cleavage was highest around the time of birth: cleaved caspase 3 was highest at 1 dPN (3.8-fold vs. 135 dGA, P=0.0380; 7.8-fold vs. 5 dPN, P=0.0118), cleaved caspase 7 and cleaved caspase 12 both increased between 135 and 143 dGA (25-fold and 6.9-fold respectively, both P<0.0001), and remained elevated at 1 dPN. Induced apoptosis, measured by TUNEL assay, was highest around the time of birth (P<0.0001). There were changes in myocardial ER stress pathway components in utero. GRP78 protein levels were high in the fetus and declined after birth (P<0.0001). EIF2A phosphorylation was profoundly depressed at 1 dPN (vs. 143 dGA, P=0.0113). There is dynamic regulation of ER proteostasis, ER stress, and the apoptosis cascade in the perinatal heart. Apoptotic signaling is more readily activated in fetal cardiomyocytes near birth, leading to widespread caspase cleavage in the newborn heart. These pathways are important for regulation of normal maturation in the healthy perinatal heart.

Abstract PO5-24-04: Fibronectin, DHPS and SLC3A2 Signaling Cooperate to Control Tumor Spheroid Growth, Subcellular eIF5A1/2 Distribution and CDK4/6 Inhibitor Resistance

Abstract Extracellular matrix (ECM) protein expression/deposition within and stiffening of the breast cancer (BC) microenvironment, facilitates disease progression and correlates with poor patient survival. However, the mechanisms by which ECM components control tumorigenic behaviors and responses to therapeutic intervention remain poorly understood. Fibronectin (FN) is a major ECM protein controlling multiple processes related to cancer metastasis. In this regard, we previously reported that DHPS-dependent hypusination of eIF5A1/2 is necessary for fibronectin-mediated BC metastasis and epithelial to mesenchymal transition (EMT). Here, we explored the clinical significance of an interactome generated using hypusination pathway components and markers of intratumoral heterogeneity. Solute carrier 3A2 (SLC3A2 or CD98hc) stood out as an indicator of poor overall survival among patients with basal-like BCs that express elevated levels of DHPS. We subsequently discovered that blockade or knockdown (KD) of DHPS or SLC3A2 reduced BC spheroid growth. Interestingly, spheroids stimulated with exogenous fibronectin were less sensitive to inhibition of either DHPS or SLC3A2 – an effect that could be abrogated by dual DHPS/SLC3A2 blockade or KD. In this regard, we observed that inhibition of both SLC3A2 and DHPS reduced total and hypusinated eIF5A1/2 levels most effectively. We further discovered that a subset of BC cells responded to fibronectin by increasing cytoplasmic localization of eIF5A1/2. Notably, these fibronectin-induced subcellular localization phenotypes correlated with a G0/G1 cell cycle arrest. Fibronectin-treated BC cells responded to dual DHPS/SLC3A2 blockade by shifting eIF5A1/2 localization back to a nucleus-dominant state, suppressing proliferation and further arresting cells in the G2/M phase of the cell cycle. Finally, we observed that dual DHPS/SLC3A2 inhibition increased the sensitivity of both Rb-negative and -positive BC cells to the CDK4/6 inhibitor palbociclib. Taken together, these data identify an uncharacterized mechanism through which extracellular fibronectin controls cancer cell tumorigenicity by modulating subcellular eIF5A1/2 localization. These findings provide prognostic/therapeutic utility for targeting the cooperative DHPS/SLC3A2 signaling axis to improve BC treatment responses. Citation Format: Cameron Geller, Joanna Maddela, Gabriela Ortiz-Soto, Ranel Tuplano, Farhana Runa, Yvess Adamian, Robert Güth, Luke Tomaneng, Joseph Cantor, Jonathan A. Kelber. Fibronectin, DHPS and SLC3A2 Signaling Cooperate to Control Tumor Spheroid Growth, Subcellular eIF5A1/2 Distribution and CDK4/6 Inhibitor Resistance [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-24-04.

Abstract PO1-24-04: Gedatolisib, a pan-PI3K/mTOR inhibitor, shows superior potency and efficacy relative to other PI3K/AKT/mTOR pathway inhibitors in breast cancer models

Abstract Background: The PI3K, AKT, and mTOR (PAM) pathway is frequently activated in breast cancer (BC). Current standard-of-care therapy options for patients with advanced BC (ABC) include PAM inhibitors (PAMi), such as everolimus and alpelisib, and hormonal therapy, such as letrozole and fulvestrant. Most PAMi selectively inhibit one or only a few PAM pathway components, which can lead to drug resistance. To minimize drug resistance, a more comprehensive inhibition of the multiple PI3K isoforms and downstream mTOR complexes may be required. We hypothesized that gedatolisib, which potently inhibits all Class I PI3K isoforms, as well as mTORC1 and mTORC2, can be more effective in BC cells than a PAMi targeting single PAM pathway components. Methods: A panel of BC cell lines with mutated or non-mutated PAM pathway genes (PIK3CA, PTEN, AKT) were evaluated for their sensitivity to gedatolisib and other PAMi (PI3K-α: alpelisib; AKT: capivasertib; mTORC1: everolimus; pan-PI3K/mTOR: gedatolisib). Cell viability, growth rate inhibition, and cell death were evaluated by luciferase- and fluorescence-based assays, both in monolayer cultures and three-dimensional (3D) cultures on reconstituted basement membrane. Flow cytometry analytical assays were conducted to assess DNA synthesis (EdU incorporation), protein synthesis (OPP incorporation), and PAM pathway activity (RPS6 and 4EBP1 phosphorylation). BC xenograft studies evaluating gedatolisib in vivo were also performed. Results: Gedatolisib strongly inhibited PAM pathway activity and reduced cell viability and growth rate in the cell lines tested. Compared to the other PAMi, gedatolisib exhibited more potent and efficacious anti-proliferative and cytotoxic effects, regardless of the cell lines’ PI3K pathway mutational status. Gedatolisib effects were confirmed in 3D culture on reconstituted basement membrane, where gedatolisib inhibited tumor cell spheroid growth and induced regression. Mechanistically, gedatolisib decreased both DNA and protein synthesis more effectively than the other PAMi tested. Gedatolisib also exerted superior anti-proliferative and cytotoxic effects relative to the other PAMi tested in estrogen receptor-positive BC cell lines concomitantly treated with fulvestrant. BC xenografts confirmed growth inhibitory effects of gedatolisib in vivo. Conclusions: The pan-PI3K/mTOR inhibitor, gedatolisib, may more effectively address potential drug resistance mechanisms associated with narrowly targeted PAM inhibitors. Gedatolisib has previously demonstrated promising preliminary clinical efficacy and safety data in ABC in combination with hormonal therapy. A Phase 3 study (VIKTORIA-1) evaluating gedatolisib plus fulvestrant with and without palbociclib is underway in patients with ABC. Citation Format: Stefano Rossetti, Aaron Broege, Adrish Sen, Salmaan Khan, Ian MacNeil, Jhomary Molden, Igor Gorbatchevsky, Brian F. Sullivan, Lance Laing. Gedatolisib, a pan-PI3K/mTOR inhibitor, shows superior potency and efficacy relative to other PI3K/AKT/mTOR pathway inhibitors in breast cancer models [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-24-04.

Seeing through the density of dense deposit disease

Little is known about what constitutes the dense deposits of dense deposit disease (DDD), apart from components of the complement pathway. This study presents the novel finding that large accumulations of apolipoprotein E are present in the deposits of DDD, as revealed by mass spectroscopy and confirmed by both confocal microscopy and immunohistochemistry. The findings suggest a new modality for diagnosis of DDD and introduce potential new mechanisms for understanding DDD pathophysiology.

Clinical significance of PNO1 as a novel biomarker and therapeutic target of hepatocellular carcinoma

AbstractThe RNA‐binding protein PNO1 plays an essential role in ribosome biogenesis. Recent studies have shown that it is involved in tumorigenesis; however, its role in hepatocellular carcinoma (HCC) is not well understood. The purpose of this study was to examine whether PNO1 can be used as a biomarker of HCC and also examine the therapeutic potential of PNO1 knockout for the treatment of HCC. PNO1 expression was upregulated in HCC and associated with poor prognosis. PNO1 expression was positively associated with tumour stage, lymph node metastasis and poor survival. PNO1 expression was significantly higher in HCC compared to that in fibrolamellar carcinoma or normal tissues. Furthermore, HCC tissues with mutant Tp53 expressed higher PNO1 than those with wild‐type Tp53. PNO1 knockout suppressed cell viability, colony formation and EMT of HCC cells. Since activation of Notch signalling pathway promotes HCC, we measured the effects of PNO1 knockout on the components of Notch pathway and its targets. PNO1 knockout suppressed Notch signalling by modulating the expression of Notch ligands and their receptors, and downstream targets. PNO1 knockout also inhibited genes involved in surface adhesion, cell cycle, inflammation and chemotaxis. PNO1 knockout also inhibited colony and spheroid formation, cell migration and invasion, and markers of stem cells, pluripotency and EMT in CSCs. Overall, our data suggest that PNO1 can be used as a diagnostic and prognostic biomarker of HCC, and knockout of PNO1 by CRISPR/Cas9 can be beneficial for the management of HCC by targeting CSCs.

Protective role of 17β-estradiol in alcohol-associated liver fibrosis is mediated by suppression of integrin signaling.

Alcohol-associated liver disease is a complex disease regulated by genetic and environmental factors such as diet and sex. The combination of high-fat diet and alcohol consumption has synergistic effects on liver disease progression. Female sex hormones are known to protect females from liver disease induced by high-fat diet. In contrast, they promote alcohol-mediated liver injury. We aimed to define the role of female sex hormones on liver disease induced by a combination of high-fat diet and alcohol. Wild-type and protein arginine methyltransferase (Prmt)6 knockout female mice were subjected to gonadectomy (ovariectomy, OVX) or sham surgeries and then fed western diet and alcohol in the drinking water. We found that female sex hormones protected mice from western diet/alcohol-induced weight gain, liver steatosis, injury, and fibrosis. Our data suggest that these changes are, in part, mediated by estrogen-mediated induction of arginine methyltransferase PRMT6. Liver proteome changes induced by OVX strongly correlated with changes induced by Prmt6 knockout. Using Prmt6 knockout mice, we confirmed that OVX-mediated weight gain, steatosis, and injury are PRMT6 dependent, while OVX-induced liver fibrosis is PRMT6 independent. Proteomic and gene expression analyses revealed that estrogen signaling suppressed the expression of several components of the integrin pathway, thus reducing integrin-mediated proinflammatory (Tnf, Il6) and profibrotic (Tgfb1, Col1a1) gene expression independent of PRMT6 levels. Integrin signaling inhibition using Arg-Gly-Asp peptides reduced proinflammatory and profibrotic gene expression in mice, suggesting that integrin suppression by estrogen is protective against fibrosis development. Taken together, estrogen signaling protects mice from liver disease induced by a combination of alcohol and high-fat diet through upregulation of Prmt6 and suppression of integrin signaling.

Activation of stress-response genes by retrograde signaling-mediated destabilization of nuclear importin IMPα-9 and its interactor TPR2

Deciphering stress-induced retrograde signal transmission from plastids to the nucleus has long puzzled plant biologists. To address this, we performed a suppressor screen of the ceh1 mutant, known for elevated MEcPP levels, and identified the gain-of-function mutant impα-9, which reverses dwarfism and suppresses stress-response genes in the ceh1 background despite heightened MEcPP.Subsequent genetics and biochemical analyses established that the accumulation of MEcPP initiates an upsurge in ASK1 abundance, a pivotal component in the proteasome degradation pathway. This increase in ASK1 prompts the degradation of IMPα-9. Additionally, we uncovered a protein interaction between IMPα-9 and TPR2, a transcriptional co-suppressor. Reduction in IMPα-9 levels coincides with a decrease in TPR2 abundance. Significantly, these interactions were disrupted in impα-9 mutants, highlighting the critical role of a single amino acid alteration in maintaining these associations. Disruption of these interactions results in the reversal of MEcPP-associated phenotypes.ChIP-seq analyses unveiled TPR2's binding to stress response genes and suggested IMPα-9-DNA association. Together, these associations function to suppress stress genes under normal conditions, but this suppression is alleviated in response to stress through the degradation of the suppressing machinery. The biological relevance of these findings was emphasized during high light stress, characterized by MEcPP accumulation, elevated ASK1 levels, degradation of IMPα-9, reduced TPR2 abundance, and subsequent activation of a network of stress response genes.In essence, our study uncovers new insights into plant adaptive responses, revealing complex interactions among retrograde signaling, the proteasome, and nuclear transport machinery, and establishes plastids as a regulatory stress response hub.

Neurochemistry and circuit organization of the lateral spiriform nucleus of birds: A uniquely nonmammalian direct pathway component of the basal ganglia.

We used diverse methods to characterize the role of avian lateral spiriform nucleus (SpL) in basal ganglia motor function. Connectivity analysis showed that SpL receives input from globus pallidus (GP), and the intrapeduncular nucleus (INP) located ventromedial to GP, whose neurons express numerous striatal markers. SpL-projecting GP neurons were large and aspiny, while SpL-projecting INP neurons were medium sized and spiny. Connectivity analysis further showed that SpL receives inputs from subthalamic nucleus (STN) and substantia nigra pars reticulata (SNr), and that the SNr also receives inputs from GP, INP, and STN. Neurochemical analysis showed that SpL neurons express ENK, GAD, and a variety of pallidal neuron markers, and receive GABAergic terminals, some of which also contain DARPP32, consistent with GP pallidal and INP striatal inputs. Connectivity and neurochemical analysis showed that the SpL input to tectum prominently ends on GABAA receptor-enriched tectobulbar neurons. Behavioral studies showed that lesions of SpL impair visuomotor behaviors involving tracking and pecking moving targets. Our results suggest that SpL modulates brainstem-projecting tectobulbar neurons in a manner comparable to the demonstrated influence of GP internus on motor thalamus and of SNr on tectobulbar neurons in mammals. Given published data in amphibians and reptiles, it seems likely the SpL circuit represents a major direct pathway-type circuit by which the basal ganglia exerts its motor influence in nonmammalian tetrapods. The present studies also show that avian striatum is divided into three spatially segregated territories with differing connectivity, a medial striato-nigral territory, a dorsolateral striato-GP territory, and the ventrolateral INP motor territory.

Chromosome-level Genome Assembly of the Yeast Lodderomyces beijingensis Reveals the Genetic Nature of Metabolic Adaptations and Identifies Subtelomeres as Hotspots for Amplification of Mating Type Loci.

Lodderomyces beijingensis is an ascosporic ascomycetous yeast. In contrast to related species Lodderomyces elongisporus, which is a recently emerging human pathogen, L. beijingensis is associated with insects. To provide an insight into its genetic makeup, we investigated the genome of its type strain, CBS 14171. We demonstrate that this yeast is diploid and describe the high contiguity nuclear genome assembly consisting of eight chromosome-sized contigs with a total size of about 15.1 Mbp. We find that the genome sequence contains multiple copies of the mating type loci and codes for essential components of the mating pheromone response pathway, however, the missing orthologs of several genes involved in the meiotic program raise questions about the mode of sexual reproduction. We also show that L. beijingensis genome codes for the 3-oxoadipate pathway enzymes, which allow the assimilation of protocatechuate. In contrast, the GAL gene cluster underwent a decay resulting in an inability of L. beijingensis to utilize galactose. Moreover, we find that the 56.5 kbp long mitochondrial DNA is structurally similar to known linear mitochondrial genomes terminating on both sides with covalently closed single-stranded hairpins. Finally, we discovered a new double-stranded RNA mycovirus from the Totiviridae family and characterize its genome sequence.

The Effect of Aspalathin on SMAD2, SMAD3, TGF-β - A Major Contributor of Inflammation – An In-silico Approach

Chronic inflammation plays a crucial role in the development of several illnesses, and the transforming growth factor-beta (TGF-β) pathway, including SMAD2 and SMAD3, regulates inflammatory responses. Aspalathin, a naturally-occurring compound extracted from several plant sources, has shown potential anti-inflammatory properties. This study uses computational methods to investigate how aspalathin affects SMAD2, SMAD3, and TGF-β in order to understand how it regulates inflammation. We use molecular docking to analyze how aspalathin binds with important components of the TGF-β pathway. Our research reveals promising information about the potential of aspalathin, which shows a strong ability to bind with the inflammatory regulator (SMAD2, SMAD3, and TGF-β). Aspalathin may offer therapeutic benefits for treating inflammatory diseases. Further testing in both controlled laboratory environments (in vitro) and inside live creatures (in vivo) are required to validate the computational findings and prove aspalathin's potential as a viable option for inflammation management.

An essential and highly selective protein import pathway encoded by nucleus-forming phage

Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate their genomes in a nucleus-like replication compartment composed of the protein chimallin (ChmA) that excludes ribosomes and decouples transcription from translation. These phages selectively partition proteins between the phage nucleus and the bacterial cytoplasm. Currently, the genes and signals that govern selective protein import into the phage nucleus are unknown. Here, we identify two components of this protein import pathway: a species-specific surface-exposed region of a phage intranuclear protein required for nuclear entry and a conserved protein, PicA (Protein importer of chimalliviruses A), that facilitates cargo protein trafficking across the phage nuclear shell. We also identify a defective cargo protein that is targeted to PicA on the nuclear periphery but fails to enter the nucleus, providing insight into the mechanism of nuclear protein trafficking. Using CRISPRi-ART protein expression knockdown of PicA, we show that PicA is essential early in the chimallivirus replication cycle. Together, our results allow us to propose a multistep model for the Protein Import Chimallivirus pathway, where proteins are targeted to PicA by amino acids on their surface and then licensed by PicA for nuclear entry. The divergence in the selectivity of this pathway between closely related chimalliviruses implicates its role as a key player in the evolutionary arms race between competing phages and their hosts.

Planar Cell Polarity Signaling: Coordinated Crosstalk for Cell Orientation

The planar cell polarity (PCP) system is essential for positioning cells in 3D networks to establish the proper morphogenesis, structure, and function of organs during embryonic development. The PCP system uses inter- and intracellular feedback interactions between components of the core PCP, characterized by coordinated planar polarization and asymmetric distribution of cell populations inside the cells. PCP signaling connects the anterior–posterior to left–right embryonic plane polarity through the polarization of cilia in the Kupffer’s vesicle/node in vertebrates. Experimental investigations on various genetic ablation-based models demonstrated the functions of PCP in planar polarization and associated genetic disorders. This review paper aims to provide a comprehensive overview of PCP signaling history, core components of the PCP signaling pathway, molecular mechanisms underlying PCP signaling, interactions with other signaling pathways, and the role of PCP in organ and embryonic development. Moreover, we will delve into the negative feedback regulation of PCP to maintain polarity, human genetic disorders associated with PCP defects, as well as challenges associated with PCP.

Gene Expression Profiling to Unfolded Proteins Response as a Risk Modulator of Patients with Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease. Despite new methods of diagnostics and treatment as well as extensive biological and immunosuppressive treatment, the etiology of RA is not fully understood. Moreover, the problem of diagnosis and treatment of RA patients is still current and affects a large group of patients. It is suggested that endoplasmic reticulum (ER)-related features may impair adaptation to chronic stress, inferring the risk of rheumatoid arthritis. The main goal in this study was evaluation of changes in mRNA translation to determine chronic ER stress conditions in rheumatoid arthritis patients. The study group consist of 86 individuals including a total of 56 rheumatoid arthritis patients and 30 healthy controls. The expression level of mRNA form blood samples of RA patients as well as controls of the unfolded protein response (UPR)-associated genes (p-eIF2, BCL-2, PERK, ATF4, and BAX) were investigated using real-time qPCR. GAPDH expression was used as a standard control. Considering the median, the expression levels of PERK, BCL-2, p-eIF2, ATF4, and BAX were found to be significantly increased in the blood of RA patients compared with the control group. The p-value for the PERK gene was 0.0000000036, the p-value for the BCL-2 gene was 0.000000014, the p-value for the p-eIF2 gene was 0.006948, the p-value for the ATF4 gene was 0.0000056, and the p-value for the BAX gene was 0.00019, respectively. Thus, it can be concluded that the targeting of the components of the PERK-dependent UPR signaling pathway via small-molecule PERK inhibitors may contribute to the development of novel, innovative treatment strategies against rheumatoid arthritis.

Prognostic and Immunotherapeutic Predictive Value of CAD Gene in Hepatocellular Carcinoma: Integrated Bioinformatics and Experimental Analysis.

Hepatocellular carcinoma (HCC) is a common type of cancer that ranks first in cancer-associated death worldwide. Carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) are the key components of the pyrimidine pathway, which promotes cancer development. However, the function of CAD in HCC needs to be clarified. In this study, the clinical and transcriptome data of 424 TCGA-derived HCC cases were analyzed. The results demonstrated that high CAD expression was associated with poor prognosis in HCC patients. The effect of CAD on HCC was then investigated comprehensively using GO annotation analysis, KEGG enrichment analysis, Gene Set Enrichment Analysis (GSEA), and CIBERSORT algorithm. The results showed that CAD expression was correlated with immune checkpoint inhibitors and immune cell infiltration. In addition, low CAD levels in HCC patients predicted increased sensitivity to anti-CTLA4 and PD1, while HCC patients with high CAD expression exhibited high sensitivity to chemotherapeutic and molecular-targeted agents, including gemcitabine, paclitaxel, and sorafenib. Finally, the results from clinical sample suggested that CAD expression increased remarkably in HCC compared with non-cancerous tissues. Loss of function experiments demonstrated that CAD knockdown could significantly inhibit HCC cell growth and migration both in vitro and in vivo. Collectively, the results indicated that CAD is a potential oncogene during HCC metastasis and progression. Therefore, CAD is recommended as a candidate marker and target for HCC prediction and treatment.