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INTRODUCTION: Understanding genetic contributors to sarcopenia (age-related loss of muscle strength and mass) is key to finding effective therapies. Variants of the bradykinin receptor 2 (BDKRB2) have been linked to athletic and muscle performance. The rs1799722-9 and rs5810761 T alleles have been shown to be overrepresented in endurance athletes, possibly due to increased transcriptional rates of the receptor. These variants have been rarely studied in older people or people with sarcopenia. METHODS: We performed a post hoc sub-study of the Leucine and ACE (LACE) inhibitor trial, which enrolled 145 participants aged ≥70 years with low grip strength and low gait speed. Participants' blood samples were genotyped for rs179972 using TaqMan and rs5810761 by amplification through Hotstar Taq. Genotypes were compared with outcomes of physical performance and body composition measures. RESULTS: Data from 136 individuals were included in the analysis. For rs1799722 the genotype frequency (TT: 17, CC: 48, CT: 71) remained in Hardy-Weinberg Equilibrium (HWE p = 0.248). There was no difference between the genotypes for six-Minute Walk Distance (6MWD) or Short Physical Performance Battery (SPPB). Men with the TT genotype had a significantly greater 6MWD than other genotypes (TT 400m vs CT 310m vs CC 314m, p = 0.027), and greater leg muscle mass (TT 17.59kg vs CT 15.04kg vs CC 15.65kg, p = 0.007). For rs5810761, the genotype frequency (-9-9: 31, +9+9: 43, -9+9: 60) remained in HWE (p = 0.269). The +9+9 genotype was associated with a significant change in SPPB score at 12 months (-9-9 0 vs -9+9 0 vs +9+9-1, p<0.001), suggesting an improvement. In men, the -9-9 genotype was associated with lower arm fat (-9-9 2.39kg vs -9+9 2.72kg vs +9+9 2.76kg, p = 0.019). CONCLUSION: In men, the rs1799722 TT genotype was associated with longer 6MWD and greater leg muscle mass, while the rs5810761 -9-9 genotype was associated with lower arm fat mass.
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Rendimiento Físico Funcional , Receptor de Bradiquinina B2 , Sarcopenia , Humanos , Masculino , Anciano , Femenino , Receptor de Bradiquinina B2/genética , Sarcopenia/genética , Músculo Esquelético/patología , Músculo Esquelético/fisiopatología , Genotipo , Alelos , Polimorfismo de Nucleótido Simple , Composición Corporal , Leucina/genética , Anciano de 80 o más Años , Fuerza de la Mano , Fuerza Muscular/genéticaRESUMEN
In response to excessive DNA damage, human cells can activate p53 to induce apoptosis. Cells lacking p53 can still undergo apoptosis upon DNA damage, yet the responsible pathways are unknown. We observed that p53-independent apoptosis in response to DNA damage coincided with translation inhibition, which was characterized by ribosome stalling on rare leucine-encoding UUA codons and globally curtailed translation initiation. A genetic screen identified the transfer RNAse SLFN11 and the kinase GCN2 as factors required for UUA stalling and global translation inhibition, respectively. Stalled ribosomes activated a ribotoxic stress signal conveyed by the ribosome sensor ZAKα to the apoptosis machinery. These results provide an explanation for the frequent inactivation of SLFN11 in chemotherapy-unresponsive tumors and highlight ribosome stalling as a signaling event affecting cell fate in response to DNA damage.
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Apoptosis , Daño del ADN , Biosíntesis de Proteínas , Ribosomas , Proteína p53 Supresora de Tumor , Humanos , Línea Celular Tumoral , Codón/genética , Leucina/genética , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Ribosomas/metabolismo , Transducción de Señal , Proteína p53 Supresora de Tumor/metabolismo , Proteína p53 Supresora de Tumor/genética , Quinasas Quinasa Quinasa PAM/genética , Quinasas Quinasa Quinasa PAM/metabolismoRESUMEN
Single nucleotide polymorphisms (SNPs) account for significant genomic variability in microbes, including the highly diverse gastric pathogen Helicobacter pylori. However, data on the effects of specific SNPs in pathogen-host interactions are scarce. Recent functional studies unravelled how a serine/leucine polymorphism in serine protease HtrA affects the formation of proteolytically active trimers and modulates cleavage of host cell-to-cell junction proteins during infection. A similar serine/leucine mutation in the carbohydrate binding domain of the adhesin BabA controls binding of ABO blood group antigens, enabling binding of either only the short Lewis b/H antigens of blood group O or also the larger antigens of blood groups A and B. Here we summarize the functional importance of these two remarkable bacterial SNPs and their effect on the outcome of pathogen-host interactions.
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Adhesinas Bacterianas , Helicobacter pylori , Leucina , Serina , Helicobacter pylori/genética , Adhesinas Bacterianas/genética , Adhesinas Bacterianas/metabolismo , Humanos , Serina/genética , Serina/metabolismo , Leucina/genética , Leucina/metabolismo , Polimorfismo de Nucleótido Simple/genética , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Infecciones por Helicobacter/microbiología , Infecciones por Helicobacter/genética , AnimalesRESUMEN
Receptor tyrosine kinases (RTKs) mediate the actions of growth factors in metazoans. In decapod crustaceans, RTKs are implicated in various physiological processes, such molting and growth, limb regeneration, reproduction and sexual differentiation, and innate immunity. RTKs are organized into two main types: insulin receptors (InsRs) and growth factor receptors, which include epidermal growth factor receptor (EGFR), fibroblast growth factor receptor (FGFR), vascular endothelial growth factor receptor (VEGFR), and platelet-derived growth factor receptor (PDGFR). The identities of crustacean RTK genes are incomplete. A phylogenetic analysis of the CrusTome transcriptome database, which included all major crustacean taxa, showed that RTK sequences segregated into receptor clades representing InsR (72 sequences), EGFR (228 sequences), FGFR (129 sequences), and PDGFR/VEGFR (PVR; 235 sequences). These four receptor families were distinguished by the domain organization of the extracellular N-terminal region and motif sequences in the protein kinase catalytic domain in the C-terminus or the ligand-binding domain in the N-terminus. EGFR1 formed a single monophyletic group, while the other RTK sequences were divided into subclades, designated InsR1-3, FGFR1-3, and PVR1-2. In decapods, isoforms within the RTK subclades were common. InsRs were characterized by leucine-rich repeat, furin-like cysteine-rich, and fibronectin type 3 domains in the N-terminus. EGFRs had leucine-rich repeat, furin-like cysteine-rich, and growth factor IV domains. N-terminal regions of FGFR1 had one to three immunoglobulin-like domains, whereas FGFR2 had a cadherin tandem repeat domain. PVRs had between two and five immunoglobulin-like domains. A classification nomenclature of the four RTK classes, based on phylogenetic analysis and multiple sequence alignments, is proposed.
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Furina , Insulina , Furina/genética , Filogenia , Insulina/genética , Transcriptoma , Cisteína , Leucina/genética , Factor A de Crecimiento Endotelial Vascular/genética , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptores ErbB/metabolismo , Receptores de Factores de Crecimiento de Fibroblastos/genética , Receptores de Factores de Crecimiento de Fibroblastos/metabolismo , Perfilación de la Expresión Génica , TirosinaRESUMEN
Translation fidelity relies on accurate aminoacylation of transfer RNAs (tRNAs) by aminoacyl-tRNA synthetases (AARSs). AARSs specific for alanine (Ala), leucine (Leu), serine, and pyrrolysine do not recognize the anticodon bases. Single nucleotide anticodon variants in their cognate tRNAs can lead to mistranslation. Human genomes include both rare and more common mistranslating tRNA variants. We investigated three rare human tRNALeu variants that mis-incorporate Leu at phenylalanine or tryptophan codons. Expression of each tRNALeu anticodon variant in neuroblastoma cells caused defects in fluorescent protein production without significantly increased cytotoxicity under normal conditions or in the context of proteasome inhibition. Using tRNA sequencing and mass spectrometry we confirmed that each tRNALeu variant was expressed and generated mistranslation with Leu. To probe the flexibility of the entire genetic code towards Leu mis-incorporation, we created 64 yeast strains to express all possible tRNALeu anticodon variants in a doxycycline-inducible system. While some variants showed mild or no growth defects, many anticodon variants, enriched with G/C at positions 35 and 36, including those replacing Leu for proline, arginine, alanine, or glycine, caused dramatic reductions in growth. Differential phenotypic defects were observed for tRNALeu mutants with synonymous anticodons and for different tRNALeu isoacceptors with the same anticodon. A comparison to tRNAAla anticodon variants demonstrates that Ala mis-incorporation is more tolerable than Leu at nearly every codon. The data show that the nature of the amino acid substitution, the tRNA gene, and the anticodon are each important factors that influence the ability of cells to tolerate mistranslating tRNAs.
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Aminoacil-ARNt Sintetasas , Saccharomyces cerevisiae , Animales , Humanos , Saccharomyces cerevisiae/genética , Anticodón/genética , Leucina/genética , ARN de Transferencia de Leucina/genética , Código Genético , Codón , ARN de Transferencia/genética , Aminoacil-ARNt Sintetasas/genética , Aminoacil-ARNt Sintetasas/metabolismo , Alanina/genética , Mamíferos/genéticaRESUMEN
Chicken toll-like receptor 7 (chTLR7) is a viral sensing pattern recognition receptor and detects ssRNA. The ligand binding site comprises leucine-rich repeats (LRRs) located in the ectodomain of chTLR7. Hence, any polymorphism in the binding site would modify its functional interaction with the ligand, resulting in varied strength of immune response. This study first aimed to compare the single nucleotide polymorphisms (SNPs) associated with the ligand binding site of TLR7 in three indigenous chicken breeds namely Aseel, Kadaknath, Nicobari along with an exotic breed White Leghorn. Four synonymous SNPs (P123P, I171I, N339N and L421L) and four non-synonymous SNPs (I121V, S135T, F356S and S447G) were identified among various breeds. We employed in silico tools to screen the pathogenic nsSNPs and one nsSNP was identified as having potential impact on chTLR7 protein. Moreover, sequence and structure-based methods were used to determine the effect of nsSNPs on protein stability. It revealed I121V, F356S, and S447G as decreasing the stability while S135T increasing the stability of chTLR7. Additionally, docking analysis confirmed that I121V and F356S reduced the binding affinity of ligands (R-848 and polyU) to chTLR7 protein. The results suggest that the nsSNPs found in this study could alter the ligand binding of chTLR7 and modify the immune response between different breeds further contributing to disease susceptibility or resistance. Further, in vitro and in vivo studies are needed to analyze the effect of these SNPs on susceptibility or resistance against various viral diseases in poultry.
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Pollos , Receptor Toll-Like 7 , Animales , Pollos/genética , Receptor Toll-Like 7/genética , Leucina/genética , Ligandos , Polimorfismo de Nucleótido SimpleRESUMEN
Yeast cells are extensively used as a key model organism owing to their highly conserved genome, metabolic pathways, and cell biology processes. To assist in genetic engineering and analysis, laboratory yeast strains typically harbor auxotrophic selection markers. When uncompensated, auxotrophic markers cause significant phenotypic bias compared to prototrophic strains and have different combinatorial influences on the metabolic network. Here, we used BY4741, a laboratory strain commonly used as a "wild type" strain in yeast studies, to generate a set of revertant strains, containing all possible combinations of four common auxotrophic markers (leu2∆, ura3∆, his3∆1, met15∆). We examined the effect of the auxotrophic combinations on complex phenotypes such as resistance to rapamycin, acetic acid, and ethanol. Among the markers, we found that leucine auxotrophy most significantly affected the phenotype. We analyzed the phenotypic bias caused by auxotrophy at the genomic level using a prototrophic version of a genome-wide deletion library and a decreased mRNA perturbation (DAmP) library. Prototrophy was found to suppress rapamycin sensitivity in many mutants previously annotated for the phenotype, raising a possible need for reevaluation of the findings in a native metabolic context. These results reveal a significant phenotypic bias caused by common auxotrophic markers and support the use of prototrophic wild-type strains in yeast research.
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Fenotipo , Saccharomyces cerevisiae , Sirolimus , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/crecimiento & desarrollo , Saccharomyces cerevisiae/efectos de los fármacos , Sirolimus/farmacología , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Etanol/farmacología , Etanol/metabolismo , Ácido Acético/metabolismo , Ácido Acético/farmacología , Marcadores Genéticos , Leucina/metabolismo , Leucina/farmacología , Leucina/genéticaRESUMEN
Background: Previous reports have demonstrated post-operative dementia and Alzheimer's disease (AD), and increased amyloid-ß levels and tau hyperphosphorylation have been observed in animal models post-anesthesia. Objective: After surgical interventions, loss in memory has been observed that has been found linked with genes modulated after anesthesia. Present study aimed to study molecular pattern present in genes modulated post anesthesia and involved in characters progressing towards AD. Methods: In the present study, 17 transcript variants belonging to eight genes, which have been found to modulate post-anesthesia and contribute to AD progression, were envisaged for their compositional features, molecular patterns, and codon and codon context-associated studies. Results: The sequences' composition was G/C rich, influencing dinucleotide preference, codon preference, codon usage, and codon context. The G/C nucleotides being highly occurring nucleotides, CpGdinucleotides were also preferred; however, CpG was highly disfavored at p3-1 at the codon junction. The nucleotide composition of Cytosine exhibited a unique feature, and unlike other nucleotides, it did not correlate with codon bias. Contrarily, it correlated with the sequence lengths. The sequences were leucine-rich, and multiple leucine repeats were present, exhibiting the functional role of neuroprotection from neuroinflammation post-anesthesia. Conclusions: The analysis pave the way to elucidate unique molecular patterns in genes modulated during anesthetic treatment and might help ameliorate the ill effects of anesthetics in the future.
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Enfermedad de Alzheimer , Anestesia , Anestésicos , Animales , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Agregado de Proteínas , Leucina/genética , Proteínas tau/genética , Proteínas tau/metabolismo , Codón/genética , Nucleótidos/genéticaRESUMEN
OBJECTIVE: Genetic studies have suggested that the branched-chain amino acids (BCAAs) valine, leucine, and isoleucine have a causal association with type 2 diabetes (T2D). However, inferences are based on a limited number of genetic loci associated with BCAAs. METHODS: Instrumental variables (IVs) for each BCAA were constructed and validated using large well-powered data sets and their association with T2D was tested using a two-sample inverse-variance weighted Mendelian randomization approach. Sensitivity analyses were performed to ensure the accuracy of the findings. A reverse association was assessed using instrumental variables for T2D. RESULTS: Estimated effect sizes between BCAA IVs and T2D, excluding outliers, were as follows: valine (ß = 0.14 change in log-odds per SD change in valine, 95% CI: -0.06 to 0.33, p = 0.17), leucine (ß = 0.15, 95% CI: -0.02 to 0.32, p = 0.09), and isoleucine (ß = 0.13, 95% CI: -0.08 to 0.34, p = 0.24). In contrast, T2D IVs were positively associated with each BCAA, i.e., valine (ß = 0.08 per SD change in levels per log-odds change in T2D, 95% CI: 0.05 to 0.10, p = 1.8 × 10-9 ), leucine (ß = 0.06, 95% CI: 0.04 to 0.09, p = 4.5 × 10-8 ), and isoleucine (ß = 0.06, 95% CI: 0.04 to 0.08, p = 2.8 × 10-8 ). CONCLUSIONS: These data suggest that the BCAAs are not mediators of T2D risk but are biomarkers of diabetes.
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Aminoácidos de Cadena Ramificada , Diabetes Mellitus Tipo 2 , Humanos , Diabetes Mellitus Tipo 2/genética , Análisis de la Aleatorización Mendeliana , Isoleucina/genética , Leucina/genética , Valina/genéticaRESUMEN
As chronic wasting disease (CWD) continues to spread across North America, the relationship between CWD and host genetics has become of interest. In Rocky Mountain elk (Cervus elaphus nelsoni), one or two copies of a leucine allele at codon 132 of the prion protein gene (132L*) has been shown to prolong the incubation period of CWD. Our study examined the relationship between CWD epidemiology and codon 132 evolution in elk from Wyoming, USA, from 2011 to 2018. Using PCR and Sanger sequencing, we genotyped 997 elk and assessed the relationship between genotype and CWD prevalence estimated from surveillance data. Using logistic regression, we showed that each 1% increase in CWD prevalence is associated with a 9.6% increase in the odds that an elk would have at least one copy of leucine at codon 132. In some regions, however, 132L* variants were found in the absence of CWD, indicating that evolutionary and epidemiologic patterns can be heterogeneous across space and time. We also provide evidence that naturally occurring CWD is not rare in 132L* elk, which merits the study of shedding kinetics in 132L* elk and the influence of genotype on CWD strain diversity. The management implications of cervid adaptations to CWD are difficult to predict. Studies that investigate the degree to which evolutionary outcomes are shaped by host spatial structure can provide useful epidemiologic insight, which can in turn aid management by informing scale and extent of mitigation actions.
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Ciervos , Priones , Enfermedad Debilitante Crónica , Animales , Enfermedad Debilitante Crónica/epidemiología , Enfermedad Debilitante Crónica/genética , Proteínas Priónicas/genética , Proteínas Priónicas/metabolismo , Leucina/genética , Leucina/metabolismo , Codón/metabolismo , Ciervos/metabolismoRESUMEN
BACKGROUND: Mutations in the glucocerebrosidase (GBA1) and leucine-rich repeat kinase 2 (LRRK2) genes, encoding lysosomal enzyme glucocerebrosidase (GCase) and leucine-rich repeat kinase 2 (LRRK2), respectively, are the most common related to Parkinson's disease (PD). Recent data suggest a possible functional interaction between GCase and LRRK2 and their involvement in sphingolipid metabolism. The aim of the present study was to describe the clinical course and evaluate the lysosomal enzyme activities and sphingolipid concentrations in blood of patients with PD associated with dual mutations p.N370S GBA1 and p.G2019S LRRK2 (p.N370S/GBA-p.G2019S/LRRK2-PD) as well as in blood of asymptomatic mutation carriers (p.N370S/GBA1-p.G2019S/LRRK2-carrier). METHODS: One patient with p.N370S/GBA1-p.G2019S/LRRK2-PD and one p.N370S/GBA1-p.G2019S/LRRK2-carrier were enrolled. GBA1-associated PD (GBA1-PD), LRRK2-associated PD (LRRK2-PD), sporadic PD (sPD) patients were described earlier by our research group. A neuropsychiatric examination of the p.N370S/GBA1-p.G2019S/LRRK2-PD patient was carried out using scales (Montreal Cognitive Assessment scale (MoCA), Mini-mental State Examination scale (MMSE), Frontal Assessment Batter scale (FAB), Hospital Anxiety, and Depression Scale (HADS), etc). Lysosomal enzyme activity (GCase, alpha-galactosidase [GLA], acid sphingomyelinase [ASMase], galactosylcerebrosidase [GALC]) and sphingolipid concentrations (hexasylsphingosine [HexSph], lysoglobotriaosylsphingosine [LysoGb3], lysosphingomyelin [LysoSM]) were assessed with high-performance liquid chromatography-tandem mass spectrometry in blood. The following comparison with the previously described groups of GBA1-PD and sPD patients were conducted. RESULTS: Clinical features of p.N370S/GBA1-p.G2019S/LRRK2-PD included an early age of onset of the disease (46 years) and mild cognitive and affective disorders (MMSE = 29, MoCA = 23), despite a long (24 years) course of the disease. Interestingly, no differences were found in hydrolase activity and lysosphingolipid concentrations between the p.N370S/GBA1-p.G2019S/LRRK2-PD patient and GBA1-PD patients. However, GCase activity was lower in these groups than in LRRK2-PD, sPD, and controls. Additionally, the p.N370S/GBA1-p.G2019S/LRRK2-PD patient was characterized by a pronounced decreased in ASMase activity and increased LysoSM concentration compared to the p.N370S/GBA1-p.G2019S/LRRK2-carrier (p = 0.023, p = 0.027, respectively). CONCLUSIONS: Based on one patient, our results indicate a protective effect of the p.G2019S mutation in the LRRK2 gene on clinical course of p.N370S/GBA1-PD. The identified pronounced alteration of ASMase activity and LysoSM concentration in p.N370S/GBA1-p.G2019S/LRRK2-PD provide the basis for the further research.
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Glucosilceramidasa , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina , Enfermedad de Parkinson , Humanos , Persona de Mediana Edad , Progresión de la Enfermedad , Glucosilceramidasa/genética , Glucosilceramidasa/metabolismo , Hidrolasas/genética , Leucina/genética , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/genética , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/metabolismo , Lisosomas/metabolismo , Mutación , Enfermedad de Parkinson/genética , EsfingolípidosRESUMEN
BACKGROUND: Studies have suggested a possible relevance between branched-chain amino acid (BCAA) catabolic enzymes and cancers. However, few studies have explored the variation in circulating concentrations of BCAAs. Our study used bi-directional, two-sample Mendelian randomization (MR) analysis for predicting the causality between the BCAA levels and 9 types of cancers. METHODS: The largest genome-wide association studies (GWAS) provided data for total BCAAs, valine, leucine, and isoleucine from the UK Biobank. Data on multiple cancer endpoints were collected from various sources, such as the International Lung Cancer Consortium (ILCCO), the Pancreatic Cancer Cohort Consortium 1 (PanScan1), the Breast Cancer Association Consortium (BCAC), the FinnGen Biobank, and the Ovarian Cancer National Alliance (OCAC). The mainly analysis method was the inverse-variance-weighted (IVW). For assessing horizontal pleiotropy, the researchers performed MR-Egger regression and MR-PRESSO global test. Finally, the Cochran's Q test served for evaluating the heterogeneity. RESULTS: Circulating total BCAAs levels (OR 1.708, 95%CI 1.168, 2.498; p = 0.006), valine levels (OR 1.747, 95%CI 1.217, 2.402; p < 0.001), leucine levels (OR 1.923, 95%CI 1.279, 2.890; p = 0.002) as well as isoleucine levels (OR 1.898, 95%CI 1.164, 3.094; p = 0.010) positively correlated with the squamous cell lung cancer risk. Nevertheless, no compelling evidence was found to support a causal link between BCAAs and any other examined cancers. CONCLUSIONS: Increased circulating total-BCAAs levels, leucine levels, isoleucine levels and valine levels had higher hazard of squamous cell lung cancer. No such associations were found for BCAAs with other cancers.
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Neoplasias de la Mama , Carcinoma de Pulmón de Células no Pequeñas , Carcinoma de Células Escamosas , Neoplasias Pulmonares , Humanos , Femenino , Isoleucina/genética , Análisis de la Aleatorización Mendeliana , Leucina/genética , Estudio de Asociación del Genoma Completo , Aminoácidos de Cadena Ramificada , Valina/genética , Neoplasias Pulmonares/genéticaRESUMEN
BACKGROUND: Filamentous fungi are efficient degraders of plant biomass and the primary producers of commercial cellulolytic enzymes. While the transcriptional regulation mechanisms of cellulases have been continuously explored in lignocellulolytic fungi, the induction of cellulase production remains a complex multifactorial system, with several aspects still largely elusive. RESULTS: In this study, we identified a Zn2Cys6 transcription factor, designated as Clr-5, which regulates the expression of cellulase genes by influencing amino acid metabolism in Neurospora crassa during growth on cellulose. The deletion of clr-5 caused a significant decrease in secreted protein and cellulolytic enzyme activity of N. crassa, which was partially alleviated by supplementing with yeast extract. Transcriptomic profiling revealed downregulation of not only the genes encoding main cellulases but also those related to nitrogen metabolism after disruption of Clr-5 under Avicel condition. Clr-5 played a crucial role in the utilization of multiple amino acids, especially leucine and histidine. When using leucine or histidine as the sole nitrogen source, the Δclr-5 mutant showed significant growth defects on both glucose and Avicel media. Comparative transcriptomic analysis revealed that the transcript levels of most genes encoding carbohydrate-active enzymes and those involved in the catabolism and uptake of histidine, branched-chain amino acids, and aromatic amino acids, were remarkably reduced in strain Δclr-5, compared with the wild-type N. crassa when grown in Avicel medium with leucine or histidine as the sole nitrogen source. These findings underscore the important role of amino acid metabolism in the regulation of cellulase production in N. crassa. Furthermore, the function of Clr-5 in regulating cellulose degradation is conserved among ascomycete fungi. CONCLUSIONS: These findings regarding the novel transcription factor Clr-5 enhance our comprehension of the regulatory connections between amino acid metabolism and cellulase production, offering fresh prospects for the development of fungal cell factories dedicated to cellulolytic enzyme production in bio-refineries.
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Celulasa , Celulasas , Neurospora crassa , Celulasa/metabolismo , Neurospora crassa/genética , Neurospora crassa/metabolismo , Histidina/genética , Histidina/metabolismo , Leucina/genética , Leucina/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Celulosa/metabolismo , Celulasas/genética , Nitrógeno/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Regulación Fúngica de la Expresión GénicaRESUMEN
The coding sequences of developmental genes are expected to be deeply conserved, with cis-regulatory change driving the modulation of gene function. In contrast, proteins with roles in defense are expected to evolve rapidly, in molecular arms races with pathogens. However, some gene families include both developmental and defense genes. In these families, does the tempo and mode of evolution differ between genes with divergent functions, despite shared ancestry and structure? The leucine-rich repeat receptor-like kinase (LRR-RLKs) protein family includes members with roles in plant development and defense, thus providing an ideal system for answering this question. LRR-RLKs are receptors that traverse plasma membranes. LRR domains bind extracellular ligands; RLK domains initiate intracellular signaling cascades in response to ligand binding. In LRR-RLKs with roles in defense, LRR domains evolve faster than RLK domains. To determine whether this asymmetry extends to LRR-RLKs that function primarily in development, we assessed evolutionary rates and tested for selection acting on 11 subfamilies of LRR-RLKs, using deeply sampled protein trees. To assess functional evolution, we performed heterologous complementation assays in Arabidopsis thaliana (Arabidopsis). We found that the LRR domains of all tested LRR-RLK proteins evolved faster than their cognate RLK domains. All tested subfamilies of LRR-RLKs had strikingly similar patterns of molecular evolution, despite divergent functions. Heterologous transformation experiments revealed that multiple mechanisms likely contribute to the evolution of LRR-RLK function, including escape from adaptive conflict. Our results indicate specific and distinct evolutionary pressures acting on LRR versus RLK domains, despite diverse organismal roles for LRR-RLK proteins.
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Arabidopsis , Proteínas de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Leucina/genética , Dominios Proteicos , Proteínas Quinasas/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas Tirosina Quinasas/genética , Evolución Molecular , FilogeniaRESUMEN
Resistance genes (R genes) are a class of genes that are immune to a wide range of diseases and pests. In planta, NLR genes are essential components of the innate immune system. Currently, genes belonging to NLR family have been found in a number of plant species, but little is known in peach. Here, 286 NLR genes were identified on peach genome by using their homologous genes in Arabidopsis thaliana as queries. These 286 NLR genes contained at least one NBS domain and LRR domain. Phylogenetic and N-terminal domain analysis showed that these NLRs could be separated into four subfamilies (I-IV) and their promoters contained many cis-elements in response to defense and phytohormones. In addition, transcriptome analysis showed that 22 NLR genes were up-regulated after infected by Green Peach Aphid (GPA), and showed different expression patterns. This study clarified the NLR gene family and their potential functions in aphid resistance process. The candidate NLR genes might be useful in illustrating the mechanism of aphid resistance in peach.
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Áfidos , Proteínas de Arabidopsis , Arabidopsis , Animales , Proteínas de Arabidopsis/genética , Áfidos/fisiología , Leucina/genética , Filogenia , Arabidopsis/genética , Nucleótidos/metabolismoRESUMEN
Amyloid aggregation is a key process in amyloidoses and neurodegenerative diseases. Hydrophobicity is one of the major driving forces for this type of aggregation, as an increase in hydrophobicity generally correlates with aggregation susceptibility and rate. However, most experimental systems in vitro and prediction tools in silico neglect the contribution of protective osmolytes present in the cellular environment. Here, we assessed the role of hydrophobic mutations in amyloid aggregation in the presence of osmolytes. To achieve this goal, we used the model protein human muscle acylphosphatase (mAcP) and mutations to leucine that increased its hydrophobicity without affecting its thermodynamic stability. Osmolytes significantly slowed down the aggregation kinetics of the hydrophobic mutants, with an effect larger than that observed on the wild-type protein. The effect increased as the mutation site was closer to the middle of the protein sequence. We propose that the preferential exclusion of osmolytes from mutation-introduced hydrophobic side-chains quenches the aggregation potential of the ensemble of partially unfolded states of the protein by inducing its compaction and inhibiting its self-assembly with other proteins. Our results suggest that including the effect of the cellular environment in experimental setups and predictive softwares, for both mechanistic studies and drug design, is essential in order to obtain a more complete combination of the driving forces of amyloid aggregation.
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Acilfosfatasa , Amiloide , Agregado de Proteínas , Humanos , Secuencia de Aminoácidos , Amiloide/química , Amiloide/genética , Leucina/química , Leucina/genética , Pliegue de Proteína , Agregado de Proteínas/genética , Acilfosfatasa/química , Acilfosfatasa/genética , Interacciones Hidrofóbicas e Hidrofílicas , Solubilidad , Presión Osmótica , Urea/químicaRESUMEN
The proliferation and differentiation of skeletal muscle satellite cells (SCs) are necessary for the development of mature skeletal muscle. Leucine (Leu) is both an essential amino acid (EAA) and a branched-chain amino acid (BCAA), which has attracted worldwide attention due to its ability to repair and become new fibers. We separated the equine SCs into the control group (CON) and the Leu-supplemented group (LEU), which the cells were cultured in Leu-deprived and Leu-supplemented media respectively. We combined the transcriptome (RNA-Seq) and quantitative proteome (TMT) profiling analyses on proliferation of equine SCs associated with Leu. 1839 up-regulated and 631 down-regulated genes made up the 2470 differentially expressed genes (DEGs), and the 253 differentially abundant proteins (DEPs) included 118 up-regulated and 135 down-regulated proteins. 110 overlapping genes were verified based on the mRNA and protein translation relationship. Moreover, by comparing overlapped pathways through enrichment analysis, we found 13 genes not only appeared among 110 key DEGs/DEPs but also enriched in the KEGG overlapping signaling pathway, including CCL26, STAT2, PCK2, ASNS, GPT2, SHMT2, PHGDH, PGAM2, PSAT1, FTL, HMOX1, STEAP1 and STEAP2. To our knowledge, this is the first report in the world to systematically show how Leu regulated the growth of equine SCs. Leu deficiency inhibits the proliferation of equine SCs and development of fresh muscle fibers was proved in this paper. The main genes in charge of the Leu-induced proliferation of horse SCs have been found. These genes will make it easier to understand the mechanism at work and offer new information for enhancing the performance of sport horses and alleviating the equine muscle damage during exercise in the future.
Asunto(s)
Células Satélite del Músculo Esquelético , Transcriptoma , Caballos/genética , Animales , Leucina/genética , Leucina/metabolismo , Leucina/farmacología , Células Satélite del Músculo Esquelético/metabolismo , Proteoma/metabolismo , Proliferación CelularRESUMEN
Drought and salt are major abiotic stresses that severely restricts plant growth and development, leading to serious losses in agricultural production. Therefore, improving crop tolerance to drought and salt stresses is an urgent issue. A previous study showed that overexpression of Arabidopsis NLR gene AtRPS2 conferred broad-spectrum disease resistance in rice. In this study, we demonstrated that constitutive expression of AtRPS2 increased abscisic acid (ABA) sensitivity during seedling stage, the shoot length of transgenic plants were shorter than wild type plants. Exogenous application of ABA markedly induced the expression of stress-related genes and promoted stomatal close in transgenic plants. Overexpression of AtRPS2 also enhanced drought and salt tolerance in rice, transgenic plants exhibited higher survival rates under drought and salt conditions than wild type plants. The activities of catalase (CAT) and superoxide dismutase (SOD) were higher in AtRPS2 transgenic rice than wild type plants. In addition, the expression of stress-related genes and ABA-responsive genes were significantly upregulated in AtRPS2 transgenic plants than wild type plants under drought and salt treatments. Besides, exogenous application of ABA could facilitate drought and salt tolerance in AtRPS2 transgenic plants. Taken together, this study indicated that AtRPS2 could improve drought and salt tolerance in rice, and this phenomenon is likely to be regulated through ABA signaling pathways.
Asunto(s)
Arabidopsis , Oryza , Tolerancia a la Sal/genética , Oryza/genética , Oryza/metabolismo , Leucina/genética , Leucina/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sequías , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/metabolismo , Ácido Abscísico/farmacología , Ácido Abscísico/metabolismo , Arabidopsis/genética , Nucleótidos/metabolismo , Regulación de la Expresión Génica de las Plantas , Estrés Fisiológico/genéticaRESUMEN
L-leucine is an essential amino acid widely used in food and pharmaceutical industries. However, the relatively low production efficiency limits its large-scale application. In this study, we rationally developed an efficient L-leucine-producing Escherichia coli strain. Initially, the L-leucine synthesis pathway was enhanced by overexpressing feedback-resistant 2-isopropylmalate synthase and acetohydroxy acid synthase both derived from Corynebacterium glutamicum, along with two other native enzymes. Next, the pyruvate and acetyl-CoA pools were enriched by deleting competitive pathways, employing the nonoxidative glycolysis pathway, and dynamically modulating the citrate synthase activity, which significantly promoted the L-leucine production and yield to 40.69 g/L and 0.30 g/g glucose, respectively. Then, the redox flux was improved by substituting the native NADPH-dependent acetohydroxy acid isomeroreductase, branched chain amino acid transaminase, and glutamate dehydrogenase with their NADH-dependent equivalents. Finally, L-leucine efflux was accelerated by precise overexpression of the exporter and deletion of the transporter. Under fed-batch conditions, the final strain LXH-21 produced 63.29 g/L of L-leucine, with a yield and productivity of 0.37 g/g glucose and 2.64 g/(L h), respectively. To our knowledge, this study achieved the highest production efficiency of L-leucine to date. The strategies presented here will be useful for engineering E. coli strains for producing L-leucine and related products on an industrial scale.