The protein methyltransferase TrSAM inhibits cellulase gene expression by interacting with the negative regulator ACE1 in Trichoderma reesei.

Protein methylation is a commonly posttranslational modification of transcriptional regulators to fine-tune protein function, however, whether this regulation strategy participates in the regulation of lignocellulase synthesis and secretion in Trichoderma reesei remains unexplored. Here, a putative protein methyltransferase (TrSAM) is screened from a T. reesei mutant with the ability to express heterologous ß-glucosidase efficiently even under glucose repression. The deletion of its encoding gene trsam causes a significant increase of cellulase activities in all tested T. reesei strains, including transformants of expressing heterologous genes using cbh1 promotor. Further investigation confirms that TrSAM interacts with the cellulase negative regulator ACE1 via its amino acid residue Arg383, which causes a decrease in the ACE1-DNA binding affinity. The enzyme activity of a T. reesei strain harboring ACE1R383Q increases by 85.8%, whereas that of the strains with trsam or ace1 deletion increases by more than 100%. By contrast, the strain with ACE1R383K shows no difference to the parent strain. Taken together, our results demonstrate that TrSAM plays an important role in regulating the expression of cellulase and heterologous proteins initiated by cbh1 promotor through interacting with ACE1R383. Elimination and mutation of TrSAM and its downstream ACE1 alleviate the carbon catabolite repression (CCR) in expressing cellulase and heterologous protein in varying degrees. This provides a new solution for the exquisite modification of T. reesei chassis.

Stigma and its influencing factors for seeking professional psychological help among pregnant women: A cross-sectional study.

BACKGROUND: Although the stigma associated with seeking professional psychological help has received significant attention, few studies have focused on the status and factors influencing this stigma among pregnant women, especially those with a Chinese cultural background. This study aimed to assess the status of stigma associated with seeking professional psychological help and its influencing factors. METHODS: A total of 1325 pregnant women were recruited from Hunan Province, China. The Stigma for Seeking Professional Psychological Help questionnaire, Patient Health Questionnaire-9, Perceived Social Support Scale, Simplified Coping Style Questionnaire, and questionnaires on sociodemographic characteristics and pregnancy-related information were used. Descriptive, univariate, correlation, and multivariate linear regression analyses were used to investigate the status of stigma and identify its influencing factors. FINDINGS: Pregnant Chinese women experienced a mild to moderate level of stigma associated with seeking professional psychological help, and self-stigma was more common than social stigma associated with seeking psychological help. The results of the multivariate analysis suggested that perceived social support, positive coping styles, negative coping styles, and depressive symptoms were the factors influencing self-stigma and social stigma associated with seeking professional psychological help. Advanced age and a low educational level were the only predictors associated with self-stigma, while a history of multiple pregnancies had an effect only on social stigma associated with seeking professional psychological help. CONCLUSION: Chinese pregnant women experienced a mild to moderate level of stigma associated with seeking professional psychological help. Destigmatizing interventions focusing on self-stigma and social stigma should be designed according to maternal characteristics, social support, coping style, and depressive symptoms.

Microstructure, Tensile Properties, and Fracture Toughness of an In Situ Rolling Hybrid with Wire Arc Additive Manufacturing AerMet100 Steel.

As a type of ultra-high strength steel, AerMet100 steel is used in the aerospace and military industries. Due to the fact that AerMet100 steel is difficult to machine, people have been exploring the process of additive manufacturing to fabricate AerMet100 steel. In this study, AerMet100 steel was produced using an in situ rolling hybrid with wire arc additive manufacturing. Microstructure, tensile properties, and fracture toughness of as-deposited and heat-treated AerMet100 steel were evaluated in different directions. The results reveal that the manufacturing process leads to grain fragmentation and obvious microstructural refinement of the AerMet100 steel, and weakens the anisotropy of the mechanical properties. After heat treatment, the microstructure of the AerMet100 steel is mainly composed of lath martensite and reversed austenite. Alloy carbides are precipitated within the martensitic matrix, and a high density of dislocations is the primary strengthening mechanism. The existence of film-like austenite among the martensite matrix enhances the toughness of AerMet100 steel, which coordinates stress distribution and restrains crack propagation, resulting in an excellent balance between strength and toughness. The AerMet100 steel with in situ rolling is isotropy and achieves the following values: an average ultimate strength of 1747.7 ± 16.3 MPa, yield strength of 1615 ± 40.6 MPa, elongation of 8.3 ± 0.2% in deposition direction, and corresponding values in the building direction are 1821.3 ± 22.1 MPa, 1624 ± 84.5 MPa, and 7.6 ± 1.7%, and the KIC value up to 70.6 MPa/m0.5.

An engineered Pichia pastoris platform for the biosynthesis of silk-based nanomaterials with therapeutic potential.

Silk fibroin (SF) from the cocoon of silkworm has exceptional mechanical properties and biocompatibility and is used as a biomaterial in a variety of fields. Sustainable, affordable, and scalable manufacturing of SF would enable its large-scale use. We report for the first time the high-level secretory production of recombinant SF peptides in engineered Pichia pastoris cell factories and the processing thereof to nanomaterials. Two SF peptides (BmSPR3 and BmSPR4) were synthesized and secreted by P. pastoris using signal peptides and appropriate spacing between hydrophilic sequences. By strain engineering to reduce protein degradation, increase glycyl-tRNA supply, and improve protein secretion, we created the optimized P. pastoris chassis PPGSP-8 to produce BmSPR3 and BmSPR4. The SF fed-batch fermentation titers of the resulting two P. pastoris cell factories were 11.39 and 9.48 g/L, respectively. Protein self-assembly was inhibited by adding Tween 80 to the medium. Recombinant SF peptides were processed to nanoparticles (NPs) and nanofibrils. The physicochemical properties of nanoparticles R3NPs and R4NPs from the recombinant SFs synthesized in P. pastoris cell factories were similar or superior to those of RSFNPs (Regenerated Silk Fibroin NanoParticles) originating from commercially available SF. Our work will facilitate the production by microbial fermentation of functional SF for use as a biomaterial.