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Tropomyosin 3 (TPM3) plays a significant role as a regulatory protein in muscle contraction, affecting the growth and development of skeletal muscles. Despite its importance, limited research has been conducted to investigate the influence of TPM3 on bovine skeletal muscle development. Therefore, this study revealed the role of TPM3 in bovine myoblast growth and development. This research involved conducting a thorough examination of the Qinchuan cattle TPM3 gene using bioinformatics tools to examine its sequence and structural characteristics. Furthermore, TPM3 expression was evaluated in various bovine tissues and cells using quantitative real-time polymerase chain reaction (qRT-PCR). The results showed that the coding region of TPM3 spans 855 bp, with the 161st base being the T base, encoding a protein with 284 amino acids and 19 phosphorylation sites. This protein demonstrated high conservation across species while displaying a predominant α-helix secondary structure despite being an unstable acidic protein. Notably, a noticeable increase in TPM3 expression was observed in the longissimus dorsi muscle and myocardium of calves and adult cattle. Expression patterns varied during different stages of myoblast differentiation. Functional studies that involved interference with TPM3 in Qinchuan cattle myoblasts revealed a very significantly decrease in S-phase cell numbers and EdU-positive staining (P

ABSTRACTIntestinal microbiota dysbiosis and metabolic disruption are considered essential characteristics in inflammatory bowel disorders (IBD). Reasonable butyrate supplementation can help patients regulate intestinal flora structure and promote mucosal repair. Here, to restore microbiota homeostasis and butyrate levels in the patient’s intestines, we modified the genome of Saccharomyces cerevisiae to produce butyrate. We precisely regulated the relevant metabolic pathways to enable the yeast to produce sufficient butyrate in the intestine with uneven oxygen distribution. A series of engineered strains with different butyrate synthesis abilities was constructed to meet the needs of different patients, and the strongest can reach 1.8 g/L title of butyrate. Next, this series of strains was used to co-cultivate with gut microbiota collected from patients with mild-to-moderate ulcerative colitis. After receiving treatment with engineered strains, the gut microbiota and the butyrate content have been regulated to varying degrees depending on the synthetic ability of the strain. The abundance of probiotics such as Bifidobacterium and Lactobacillus increased, while the abundance of harmful bacteria like Candidatus Bacilloplasma decreased. Meanwhile, the series of butyrate-producing yeast significantly improved trinitrobenzene sulfonic acid (TNBS)-induced colitis in mice by restoring butyrate content. Among the series of engineered yeasts, the strain with the second-highest butyrate synthesis ability showed the most significant regulatory and the best therapeutic effect on the gut microbiota from IBD patients and the colitis mouse model. This study confirmed the existence of a therapeutic window for IBD treatment by supplementing butyrate, and it is necessary to restore butyrate levels according to the actual situation of patients to restore intestinal flora.

Recently, the activation of chlorine dioxide (ClO2) by metal(oxide) for soil remediation has gained notable attention. However, the related activation mechanisms are still not clear. Herein, the variation of iron species and ClO2, the generated reactive oxygen species, and the toxicity of the degradation intermediates were explored and evaluated with nanoscale zero-valent iron (nFe0) being employed to activate ClO2 for soil polycyclic aromatic hydrocarbon (PAH) removal. With an optimized ClO2/nFe0 molar ratio of 15:1 and a soil/water ratio of 3:1, the degradation efficiency of phenanthrene improved 12% in comparison with that of a ClO2-alone system. The presence of nFe0 significantly promoted ClO2 consumption (improved 85.4%) but restrained ClO2− generation (reduced 22.5%). The surface Fe(II) and soluble Fe(II) in the ClO2/nFe0 system was 2.0-fold and 2.8-fold that in the nFe0 system after 2 min. Electron paramagnetic resonance analysis, along with quenching experiments, revealed that Fe(IV), HOCl, and •OH dominated phenanthrene degradation in a ClO2/nFe0 system, with oxidation contributions, respectively, of 34.3%, 52.8% and 12.9%. The degradation intermediates of PAHs in the ClO2/nFe0 system had lower estimated toxicity than those of the ClO2 system. The lettuces grown in ClO2/nFe0-treated soil displayed better results in bioassay indexes than those grown in ClO2-treated soil. This study offers new perspectives for the remediation of organic-pollutant-contaminated soil by using metal-activated ClO2 technology.

The effect of cathodic polarization on stress corrosion cracking (SCC) of 35CrMo steel for high strength fasteners in seawater was investigated by electrochemical measurement, slow strain rate tensile tests and fracture analyses. The results demonstrated that the 35CrMo steel with a typical tempered martensite microstructure has some SCC susceptibility at the open circuit potential (OCP), and the susceptibility decreases as the potential is shifted negatively until −850 mV vs. SCE, at which the steel presents the lowest SCC susceptibility due to the inhibition of anodic dissolution (AD) and little hydrogen evolution. The SCC sensitivity increases significantly as the applied potential further shifts negatively because of intensive hydrogen evolution. The fractures are transgranular at all potentials. AD is considered to be the main cracking mechanism at OCP, while hydrogen embrittlement becomes dominant at the potential below −850 mV vs. SCE.

Abstract Background In the process of cellulose hydrolysis, carbohydrate hydrolysates are transported into cells through membrane transporters, and then affect the expression of cellulase-encoding genes. Sugar transporters play a crucial role in cellulase production in lignocellulolytic fungi, of which relatively few have been functionally validated to date and are all reported to be on cell membrane. Result Through transcriptome analysis and qRT-PCR, three putative MFS sugar transporters GST, MFS, and LAC1 were found to display significantly higher mRNA levels in T. reesei grown on cellulose than on glucose. The individual deletion of these three genes compromised cellulase production and delayed sugar absorption by 24 h in T. reesei. Nevertheless, they transported pretty low level of sugars, including galactose, lactose, and mannose, and did not transport glucose, when expressed in yeast system. Meanwhile, all three transporters were unexpectedly found to be intracellular, being located in endoplasmic reticulum (ER). Particularly, the knockout of lac1 almost abolished cellulase production, and significantly inhibited biomass generation regardless of sugar types, indicating that lac1 is essential for cellulase production and biomass formation. The absence of lac1 upregulated genes involved in ribosome biogenesis, while downregulated genes in cellulase production, protein processing in ER (particularly protein glycosylation), and lipid biosynthesis. The inhibition of lac1 deletion on the transcriptional levels of genes related to cellulase biosynthesis was restored after 72 h, but the cellulase production was still inhibited, indicating lac1 might pose a post-transcription regulation on cellulase production that are independent on the known cellulase regulation mediated by CRT1 and XYR1. Conclusion For the first time, intracellular sugar transporters (mfs, gst, and lac1) facilitating cellulase production were identified, which was distributed in ER. Their sugar transporting ability was very weak, indicating that they might be related to sugar utilization inside cells rather than the cellular sugar uptake. More importantly, sugar transporter lac1 is first found to be essential for cellulase production and biomass formation by affecting protein processing in ER (particularly protein glycosylation) and lipid biosynthesis. The effect of LAC1 on cellulase production seems to be post-transcriptional at late stage of cellulase production, independent on the well-known cellulase regulation mediated by CRT1 and XYR1. These findings improve the understanding of intracellular sugar transporters in fungi and their important role in cellulase synthesis.

Antibiotics are a category of chemical compounds used to treat bacterial infections and are widely applied in cultivation, animal husbandry, aquaculture, and pharmacy. Currently, residual antibiotics and their metabolites pose a potential risk of allergic reactions, bacterial resistance, and increased cancer incidence. Residual antibiotics and the resulting bacterial antibiotic resistance have been recognized as a global challenge that has attracted increasing attention. Therefore, monitoring antibiotics is a critical way to limit the ecological risks from antibiotic pollution. Accordingly, it is desirable to devise new analytical platforms to achieve efficient antibiotic detection with excellent sensitivity and specificity. Quantum dots (QDs) are regarded as an ideal material for use in the development of antibiotic detection biosensors. In this review, we characterize different types of QDs, such as silicon, chalcogenide, carbon, and other doped QDs, and summarize the trends in QD-based antibiotic detection. QD-based sensing applications are classified according to their recognition strategies, including molecularly imprinted polymers (MIPs), aptamers, and immunosensors. We discuss the advantages of QD-derived antibiotic sensors, including low cost, good sensitivity, excellent stability, and fast response, and illustrate the current challenges in this field.

Abstract Background Knowledge on regulatory networks associated with cellulase biosynthesis is prerequisite for exploitation of such regulatory systems in enhancing cellulase production with low cost. The biological functions of intron retention (IR) and nonsense-mediated mRNA decay (NMD) in filamentous fungi is lack of study, let alone their roles in cellulase biosynthesis. Results We found that major cellulase genes (cel7a, cel7b, and cel3a) exhibited concomitant decrease in IR rates and increase in their gene expression in T. reesei under cellulase-producing condition (cellulose and lactose) that was accompanied with a more active NMD pathway, as compared to cellulase non-producing condition (glucose). In the presence of the NMD pathway inhibitor that successfully repressed the NMD pathway, the mRNA levels of cellulase genes were sharply down-regulated, but the rates of IR in these genes were significantly up-regulated. Consistently, the cellulase activities were severely inhibited. In addition, the NMD pathway inhibitor caused the downregulated mRNA levels of two important genes of the target of rapamycin (TOR) pathway, trfkbp12 and trTOR1. The absence of gene trfkbp12 made the cellulase production in T. reesei more sensitive to the NMD pathway inhibitor. Conclusions All these findings suggest that the IR of cellulase genes regulates their own gene expression by coupling with the NMD pathway, which might involve the TOR pathway. Our results provide better understanding on intron retention, the NMD pathway, and cellulase production mechanism in filamentous fungi.

Micro/nano topographic structures have shown great utility in many biomedical areas including cell therapies, tissue engineering, and implantable devices. Computer-assisted informatics methods hold great promise for the design of topographic structures with targeted properties for a specific medical application. To benefit from these methods, researchers and engineers require a highly reusable “one structural parameter – one set of cell responses” database. However, existing confounding factors in topographic cell culture devices seriously impede the acquisition of this kind of data. Through carefully dissecting the confounding factors and their possible reasons for emergence, we developed corresponding guideline requirements for topographic cell culture device development to remove or control the influence of such factors. Based on these requirements, we then suggested potential strategies to meet them. In this work, we also experimentally demonstrated a topographic cell culture device with controlled confounding factors based on these guideline requirements and corresponding strategies. A “guideline for the development of topographic cell culture devices” was summarized to instruct researchers to develop topographic cell culture devices with the confounding factors removed or well controlled. This guideline aims to promote the establishment of a highly reusable “one structural parameter – one set of cell responses” database that could facilitate the application of informatics methods, such as artificial intelligence, in the rational design of future biotopographic structures with high efficacy.

Taxadiene is an important precursor in taxol biosynthesis pathway, but its biosynthesis in eukaryotic cell factories is limited, which seriously hinders the biosynthesis of taxol. In this study, it is found that there was the catalysis compartmentalization between two key exogenous enzymes of geranylgeranyl pyrophosphate synthase and taxadiene synthase (TS) for taxadiene synthesis progress, due to their different subcellular localization. Firstly, the enzyme-catalysis compartmentalization was overcome by means of the intracellular relocation strategies of taxadiene synthase, including N-terminal truncation of taxadiene synthase and enzyme fusion of GGPPS-TS. With the help of two strategies for enzyme relocation, the taxadiene yield was increased by 21% and 54% respectively, among them the GGPPS-TS fusion enzyme is more effective. Further, the expression of GGPPS-TS fusion enzyme was improved via the multi-copy plasmid, resulting that the taxadiene titer was increased by 38% to 21.8 mg/L at shake-flask level. Finally, the maximum taxadiene titer of 184.2 mg/L was achieved by optimization of the fed-batch fermentation conditions in 3 L bioreactor, which is the highest reported titer of taxadiene biosynthesis accomplished in eukaryotic microbes. This study provides a successful example for improving biosynthesis of complex natural products by solving the critical problem of multistep enzymes catalysis compartmentalization.

As the main product of livestock, muscle itself plays an irreplaceable role in maintaining animal body movement and regulating metabolism. Therefore, it is of great significance to explore its growth, development and regeneration to improve the meat yield and quality of livestock. In this study, we attempted to use RNA-seq and ATAC-seq techniques to identify differentially expressed genes (DEGs) specifically expressed in bovine skeletal muscle as potential candidates for studying the regulatory mechanisms of muscle development. Microarray data from 8 tissue samples were selected from the GEO database for analysis. First, we obtained gene modules related to each tissue through WGCNA analysis. Through Gene Ontology (GO) functional annotation, the module of lightyellow (MElightyellow) was closely related to muscle development, and 213 hub genes were screened as follow-up research targets. Further, the difference analysis showed that, except for PREB, all other candidate hub genes were up-regulated (muscle group vs. other-group). ATAC-seq analysis showed that muscle-specific accessible chromatin regions were mainly located in promoter of genes related to muscle structure development (GO:0061061), muscle cell development (GO:0055001) and muscle system process (GO:0003012), which were involved in cAMP, CGMP-PKG, MAPK, and other signaling pathways. Next, we integrated the results of RNA-seq and ATAC-seq analysis, and 54 of the 212 candidate hub genes were identified as key regulatory genes in skeletal muscle development. Finally, through motif analysis, 22 of the 54 key genes were found to be potential target genes of transcription factor MEF2C. Including CAPN3, ACTN2, MB, MYOM3, SRL, CKM, ALPK3, MAP3K20, UBE2G1, NEURL2, CAND2, DOT1L, HRC, MAMSTR, FSD2, LRRC2, LSMEM1, SLC29A2, FHL3, KLHL41, ATXN7L2, and PDRG1. This provides a potential reference for studying the molecular mechanism of skeletal muscle development in mammals.

Preadipocyte proliferation is a critical and precisely orchestrated procedure in adipogenesis, which is highly regulated by microRNAs (miRNAs). A previous study identified that the expression of miR-33a is different in intramuscular fat (IMF) tissues from steers and bulls. In the present study, miR-33a was overexpressed in bovine preadipocytes, and a total of 781 differentialy expressed genes were found, including 348 upregulated and 433 downregulated genes. Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway analyses of the differentially expressed genes enriched cell division and cell cycle respectively. MiR-33a overexpression decreased the rate of preadipocyte proliferation. Synchronously, the mRNA and protein expression levels of proliferation-related marker genes, including cyclin B1 (CCNB1) and proliferating cell nuclear antigen (PCNA), were decreased. In contrast, inhibiting miR-33a increased the rate of preadipocyte proliferation, and expression levels of CCNB1 and PCNA. Furthermore, based on luciferase reporter assays, miR-33a targeted directly cyclin-dependent kinase 6 (CDK6)-3ʹUTR and inhibited CDK6 protein expression. Interestingly, the silencing of CDK6 inhibited bovine preadipocyte proliferation and proliferation-related genes. Therefore, miR-33a inhibits the proliferation of bovine preadipocytes. CDK6 is the target gene of miR-33a and may be involved in the effects of miR-33a on bovine preadipocyte proliferation.

The family with sequence similarity 13 member A (FAM13A) gene has been discovered in recent years and is related to metabolism. In this study, the function of FAM13A in precursor adipocyte proliferation in Qinchuan cattle was investigated using fluorescence quantitative polymerase chain reaction (PCR), western blotting, 5-ethynyl-2′-deoxyuridine staining, and other tests. FAM13A promoted precursor adipocyte proliferation. To determine the pathway FAM13A was involved in, transcriptome sequencing, fluorescence quantitative PCR, western blotting, and other tests were used, which identified the hypoxia inducible factor-1 (HIF-1) signalling pathway. Finally, cobalt chloride, a chemical mimic of hypoxia, was used to treat precursor adipocytes. mRNA and protein levels of FAM13A were significantly increased after hypoxia. Thus, FAM13A promoted bovine precursor adipocyte proliferation by inhibiting the HIF-1 signalling pathway, whereas chemically induced hypoxia negatively regulated FAM13A expression, regulating cell proliferation.

Polyethylene terephthalate (PET) biodegradation is regarded as an environmentally friendly degradation method. In this study, an artificial microbial consortium composed of Rhodococcus jostii, Pseudomonas putida and two metabolically engineered Bacillus subtilis was constructed to degrade PET. First, a two-species microbial consortium was constructed with two engineered B. subtilis that could secrete PET hydrolase (PETase) and monohydroxyethyl terephthalate hydrolase (MHETase), respectively; it could degrade 13.6% (weight loss) of the PET film within 7 days. A three-species microbial consortium was further obtained by adding R. jostii to reduce the inhibition caused by terephthalic acid (TPA), a breakdown product of PET. The weight of PET film was reduced by 31.2% within 3 days, achieving about 17.6% improvement compared with the two-species microbial consortium. Finally, P. putida was introduced to reduce the inhibition caused by ethylene glycol (EG), another breakdown product of PET, obtaining a four-species microbial consortium. With the four-species consortium, the weight loss of PET film reached 23.2% under ambient temperature. This study constructed and evaluated the artificial microbial consortia in PET degradation, which demonstrated the great potential of artificial microbial consortia in the utilization of complex substrates, providing new insights for biodegradation of complex polymers.

Sugar transporters play an important role in the cellulase production of lignocellulose-degrading fungi. Nevertheless, the role and function of these transporters are still unclear. Here we first report intracellular sugar transporters assisting cellulase production in Trichoderma reesei (T. reesei) using lactose. The mRNA levels of sugar transporter genes mfs, gst, and lac1 were substantially upregulated in T. reesei cultivated on lactose, with the most abundant mRNA levels at 24 h as compared to glucose. Moreover, the individual deletion of these sugar transporters significantly inhibited cellulase production, solid cell growth, and sporulation of T. reesei, suggesting they play a supporting role in cellulase production when grown in lactose. Surprisingly, MFS, GST, and LAC1 were mainly localized in the cytoplasm, with MFS and LAC1 in the endoplasmic reticulum (ER), representing the first discovery of intracellular sugar transporters involved in cellulase biosynthesis in lactose culture. The absence of the gene lac1 noticeably inhibited most of the crucial genes related to cellulase production, including cellulase-encoding genes, transcription factors, and sugar transporters, at 24 h, which was fully relieved at 48 h or 72 h, indicating that lac1 affects cellulase production more at the early step. This research advances the understanding of the function of intracellular sugar transporters in fungi, particularly for fungal cellulase production.

Lactic acid, a metabolic by-product of host and intestinal microbiota, has been recovered as an active signal molecule in the immune system. In this study, a lactic acid biosynthesis pathway that directly produces lactic acid from glucose rather than ethanol with high production was reconstructed in Saccharomyces cerevisiae. The engineered S. cerevisiae showed anti-inflammatory activity in dextran sulfate sodium (DSS)-induced mice with improved histological damage, increased mucosal barrier, and decreased intestinal immune response. Lactic acid regulated the macrophage polarization state and inhibited the expression of pro-inflammatory cytokines in vivo and in vitro. Increasing the macrophage monocarboxylic acid transporter-mediated active lactic acid uptake suppressed the excessive activation of the NLRP3 inflammasome and the downstream caspase-1 pathway in macrophages. Moreover, lactic acid promoted histone H3K9 acetylation and histone H3K18 lactylation. Meanwhile, the engineered S. cerevisiae altered the diversity and composition of the intestinal microbiota and changed the abundance of metabolic products in mice with colitis. In conclusion, this study shows that the application of engineered S. cerevisiae attenuated DSS-induced colitis in mice via suppressing macrophage pyroptosis and modulating the intestinal microbiota, which is an effective and safe treatment strategy for ulcerative colitis.

Engineering the biosynthesis of plant-derived natural products in microbes presents several challenges, especially when the expression and activation of the plant cytochrome P450 enzyme is required. By recruiting two enzymes—HpaB and HpaC—from several bacteria, we constructed functional 4-hydroxyphenylacetate 3-hydroxylase (4HPA3H) in Saccharomyces cerevisiae to take on a role similar to that of the plant-derived cytochrome P450 enzyme and produce caffeic acid. Along with a common tyrosine ammonia lyase (TAL), the different combinations of HpaB and HpaC presented varied capabilities in producing the target product, caffeic acid, from the substrate, L-tyrosine. The highest production of caffeic acid was obtained with the enzyme combination of HpaB from Pseudomonas aeruginosa and HpaC from Salmonella enterica, which yielded up to (289.4 ± 4.6) mg·L−1 in shake-flask cultivation. The compatibility of heterologous enzymes within a yeast chassis was effectively improved, as the caffeic acid production was increased by 40 times from the initial yield. Six key amino acid residues around the flavin adenine dinucleotide (FAD) binding domain in HpaB from Pseudomonas aeruginosa were differentiate from those other HpaBs, and might play critical roles in affecting enzyme activity. We have thus established an effective approach to construct a highly efficient yeast system to synthesize non-native hydroxylated phenylpropanoids. Keywords: Saccharomyces cerevisiae, Caffeic acid, Heterologous enzyme, Cytochrome P450, Synthetic biology

Although β-glucosidases play an important role in fungal cellulase induction and production, our current understanding does not provide a global perspective on β-glucosidase function. This work comprehensively studies all the β-glucosidases regarding their effect on cellulase production and their cellular distribution and secretion.

In this study, an silver (Ag) plating with micro/nano-dendrite structures is prepared on the 304 stainless steel (SS304) surface by potentiostatic deposition (Ag/SS304). After being modified by n-dodecyl mercaptan (NDM) with the low surface energy, the obtained sample (NDM@Ag/SS304) exhibits stable superhydrophobicity and excellent hot-water repellency. The surface morphology and composition of NDM@Ag/SS304 are analyzed by scanning electron microscope (SEM), X-ray spectrometer (EDS), X-ray diffractometer (XRD), and X-ray photoelectron spectrometer (XPS) characterization. The electrochemical measurements, tests of water contact angle (WCA), and interfacial contact resistance (ICR) are employed to systematically study the performance of the NDM@Ag/SS304 in the simulated cathode environment of proton exchange membrane fuel cell (PEMFC). The results show that the NDM@Ag/SS304 has high corrosion potential (~0.25 V) and low corrosion current density (~4.04 μA/cm2); after potentiostatic polarization (0.6 V, 5 h), the NDM@Ag/SS304 also shows high superhydrophobic stability.

Poly(ethylene terephthalate) hydrolase (PETase) from Ideonella sakaiensis exhibits a strong ability to degrade poly(ethylene terephthalate) (PET) at room temperature, and is thus regarded as a potential tool to solve the issue of polyester plastic pollution. Therefore, we explored the interaction between PETase and the substrate (a dimer of the PET monomer ethylene terephthalate, 2PET), using a model of PETase and its substrate. In this study, we focused on six key residues around the substrate-binding groove in order to create novel high-efficiency PETase mutants through protein engineering. These PETase mutants were designed and tested. The enzymatic activities of the R61A, L88F, and I179F mutants, which were obtained with a rapid cell-free screening system, exhibited 1.4 fold, 2.1 fold, and 2.5 fold increases, respectively, in comparison with wild-type PETase. The I179F mutant showed the highest activity, with the degradation rate of a PET film reaching 22.5 mg per μmol·L−1 PETase per day. Thus, this study has created enhanced artificial PETase enzymes through the rational protein engineering of key hydrophobic sites, and has further illustrated the potential of biodegradable plastics. Keywords: Polyesterase, PET degradation, Cell-free protein synthesis, Polyester, PETase

Abstract Background The induction of cellulase production by insoluble carbon source cellulose was a common and efficient strategy, but has some drawbacks, such as difficult fermentation operation, substantial cellulase loss, long fermentation time, and high energy-consumption, resulting in high cost of cellulase production in industry. These drawbacks can be overcome if soluble carbon sources are utilized as the inducers for cellulase production. However, until now the induction efficiency of most soluble carbon sources, especially lactose and glucose, is still inferior to cellulose despite extensive efforts have been made by either optimizing the fermentation process or constructing the recombinant strains. Therefore, strain improvement by metabolic engineering for high induction efficiency of soluble carbon sources is of great interest. Results Trichoderma reesei mutant SEU-7 was constructed from T. reesei RUT-C30 with the overexpression of endogenous gene β-glucosidase (BGL1) by insertional mutagenesis via Agrobacterium tumefaciens-mediated transformation (AMT). Compared to RUT-C30, SEU-7 displays substantially enhanced activities of both cellulase and hemicellulase when grown on either lactose or cellulose. The induction efficiency with lactose was found to be higher than cellulose in strain SEU-7. To the best of our knowledge, we achieved the highest FPase activity in SEU-7 in both batch culture (13.0 IU/mL) and fed-batch culture (47.0 IU/mL) on lactose. Moreover, SEU-7 displayed unrivaled pNPGase activity on lactose in both batch culture (81.0 IU/mL) and fed-batch culture (144.0 IU/mL) as compared to the other reported T. reesei strains in the literature grown in batch or fed-batch experiments on cellulose or lactose. This superiority of SEU-7 over RUT-C30 improves markedly the saccharification ability of SEU-7 on pretreated corn stover. The overexpression of gene BGL1 was found either at the mRNA or at the protein level in the mutant strains with increased cellulase production in comparison with RUT-C30, but only SEU-7 displayed much higher expression of gene BGL1 on lactose than on cellulose. Two copies of gene BGL1 were inserted into the chromosome of T. reesei SEU-7 between KI911141.1:347357 and KI911141.1:347979, replacing the original 623-bp fragment that is not within any genes’ coding region. The qRT-PCR analysis revealed that the mRNA levels of both cellulase and hemicellulase were upregulated significantly in SEU-7, together with the MFS transporter CRT1 and the XYR1 nuclear importer KAP8. Conclusions Recombinant T. reesei SEU-7 displays hyper-production of both cellulase and hemicellulase on lactose with the highest FPase activity and pNPGase activity for T. reesei, enabling highly efficient saccharification of pretreated biomass. For the first time, the induction efficiency for cellulase production by lactose in T. reesei was reported to be higher than that by cellulose. This outperformance of T. reesei SEU-7, which is strain-specific, is attributed to both the overexpression of gene BGL and the collateral mutation. Moreover, the increased transcription levels of cellulase genes, the related transcription factors, and the MFS transporter CRT1 contribute to the outstanding cellulase production of SEU-7. Our research advances strain improvement to enhance the induction efficiency of soluble carbon sources to produce cost-effective cellulase and hemicellulase in industry.

Skeletal muscle is an important component of livestock and poultry organisms. The proliferation and differentiation of myoblasts are highly coordinated processes, which rely on the regulation of miRNA. MiRNAs are widely present in organisms and play roles in various biological processes, including cell proliferation, differentiation, and apoptosis. MiR-181d and miR-196a, identified as tumor suppressors, have been found to be involved in cell proliferation, apoptosis, directed differentiation, and cancer cell invasion. However, their role in beef cattle skeletal muscle metabolism remains unclear. In this study, we discovered that overexpression of bta-miR-181d and bta-miR-196a in Qinchuan cattle myoblasts inhibited proliferation and apoptosis while promoting myogenic differentiation through EDU staining, flow cytometry analysis, immunofluorescence staining, and Western blotting. RNA-seq analysis of differential gene expression revealed that after overexpression of bta-miR-181d and bta-miR-196a, the differentially expressed genes were mainly enriched in the PI3K-Akt and MAPK signaling pathways. Furthermore, the phosphorylation levels of key proteins p-AKT in the PI3K signaling pathway and p-MAPK in the MAPK signaling pathway were significantly decreased after overexpression of bta-miR-181d and bta-miR-196a. Overall, this study provides preliminary evidence that bta-miR-181d and bta-miR-196a may regulate proliferation, apoptosis, and differentiation processes in Qinchuan cattle myoblasts by affecting the phosphorylation status of key proteins in PI3K-Akt and MAPK-ERK signaling pathways [ABSTRACT FROM AUTHOR]