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In the context of the digital economy, enterprise digitalization represents a significant opportunity to enhance core competitiveness. Nevertheless, the digitalization of enterprises may have unanticipated effects on employee’s mental health, which could impede the digital advancement of the enterprise. Such consequences may be evidenced by employee role overload and burnout, which in turn impede enterprise digitalization. This study employed the JR-D model to investigate the empirical data obtained from 250 completed questionnaires by employees. The results indicated that (1) enterprise digitalization had a positive effect on young employees’ levels of role overload and burnout; (2) employees’ role overload served as a mediator between enterprise digitalization and burnout; (3) perceived organizational support moderated the direct predictive effect of enterprise digitalization on burnout and the indirect predictive effect of enterprise digitalization on burnout through role overload, respectively. The higher the level of perceived organizational support, the weaker the direct and indirect predictive effects would be. This study proposed measures and recommendations to mitigate the unintended consequences of digitalization and promote optimal advancement of enterprise digitalization.

Cytosine base editors (CBEs) enable programmable C-to-T conversion without DNA double-stranded breaks and homology-directed repair in a variety of organisms, which exhibit great potential for agricultural and biomedical applications. However, all reported cases only involved C-to-T substitution at a single targeted genomic site. Whether C-to-T substitution is effective in multiple sites/loci has not been verified in large animals. Here, by using pigs, an important animal for agriculture and biomedicine, as the subjective animal, we showed that CBEs could efficiently induce C-to-T conversions at multiple sites/loci with the combination of three genes, including DMD , TYR , and LMNA , or RAG1 , RAG2 , and IL2RG , simultaneously, at the embryonic and cellular levels. CBEs also could disrupt genes ( pol gene of porcine endogenous retrovirus) with dozens of copies by introducing multiple premature stop codons. With the CBEs, pigs carrying single gene or multiple gene point mutations were generated through embryo injection or nuclear transfer approach. Base editors can efficiently produce single nucleotide alterations without requiring a double-strand break. Here the authors show base editing at multiple sites simultaneously in pigs.

Developed buds of pears ( Pyrus pyrifolia ) remain in a dormant state and do not break dormancy until the following spring. However, climatic factors and leaf diseases trigger defoliation in late summer and early autumn, which typically results in the bud paradormancy release and subsequent sprouting. This not only depletes tree nutrients but also diminishes the quantity of flowers and the yield of fruit in the following summer. In this study, metabolic changes in ‘Cuiguan’ pear buds were investigated following premature leaf fall. A total of 1,533 metabolites were annotated, with the majority being downregulated. Sugar levels decreased during the release of paradormancy, likely to provide energy for subsequent growth. Concurrently, most amino acids were consumed post-defoliation, with only a few showing increased trends. Furthermore, the observed low levels of phenylpropanoid-related metabolites in flower buds may contribute to premature senescence. This metabolic profile contributes to our understanding of the biological mechanism of paradormancy release from defoliation, and clarifies the metabolic changes during this process.

Background Many favorable traits of crops and livestock and human genetic diseases arise from multiple single nucleotide polymorphisms or multiple point mutations with heterogeneous base substitutions at the same locus. Current cytosine or adenine base editors can only accomplish C-to-T (G-to-A) or A-to-G (T-to-C) substitutions in the windows of target genomic sites of organisms; therefore, there is a need to develop base editors that can simultaneously achieve C-to-T and A-to-G substitutions at the targeting site. Results In this study, a novel fusion adenine and cytosine base editor (ACBE) was generated by fusing a heterodimer of TadA (ecTadA WT/* ) and an activation-induced cytidine deaminase (AID) to the N- and C-terminals of Cas9 nickase (nCas9), respectively. ACBE could simultaneously induce C-to-T and A-to-G base editing at the same target site, which were verified in HEK293-EGFP reporter cell line and 45 endogenous gene loci of HEK293 cells. Moreover, the ACBE could accomplish simultaneous point mutations of C-to-T and A-to-G in primary somatic cells (mouse embryonic fibroblasts and porcine fetal fibroblasts) in an applicable efficiency. Furthermore, the spacer length of sgRNA and the length of linker could influence the dual base editing activity, which provided a direction to optimize the ACBE system. Conclusion The newly developed ACBE would expand base editor toolkits and should promote the generation of animals and the gene therapy of genetic diseases with heterogeneous point mutations.

Mitochondrial diseases are maternally inherited heterogeneous disorders that are primarily caused by mitochondrial DNA (mtDNA) mutations. Depending on the ratio of mutant to wild-type mtDNA, known as heteroplasmy, mitochondrial defects can result in a wide spectrum of clinical manifestations. Mitochondria-targeted endonucleases provide an alternative avenue for treating mitochondrial disorders via targeted destruction of the mutant mtDNA and induction of heteroplasmic shifting. Here, we generated mitochondrial disease patient-specific induced pluripotent stem cells (MiPSCs) that harbored a high proportion of m.3243A>G mtDNA mutations and caused mitochondrial encephalomyopathy and stroke-like episodes (MELAS). We engineered mitochondrial-targeted transcription activator-like effector nucleases (mitoTALENs) and successfully eliminated the m.3243A>G mutation in MiPSCs. Off-target mutagenesis was not detected in the targeted MiPSC clones. Utilizing a dual fluorescence iPSC reporter cell line expressing a 3243G mutant mtDNA sequence in the nuclear genome, mitoTALENs displayed a significantly limited ability to target the nuclear genome compared with nuclear-localized TALENs. Moreover, genetically rescued MiPSCs displayed normal mitochondrial respiration and energy production. Moreover, neuronal progenitor cells differentiated from the rescued MiPSCs also demonstrated normal metabolic profiles. Furthermore, we successfully achieved reduction in the human m.3243A>G mtDNA mutation in porcine oocytes via injection of mitoTALEN mRNA. Our study shows the great potential for using mitoTALENs for specific targeting of mutant mtDNA both in iPSCs and mammalian oocytes, which not only provides a new avenue for studying mitochondrial biology and disease but also suggests a potential therapeutic approach for the treatment of mitochondrial disease, as well as the prevention of germline transmission of mutant mtDNA.

Human pluripotent stem cells (hPSCs) exhibit very limited contribution to interspecies chimeras. One explanation is that the conventional hPSCs are in a primed state and so unable  to form chimeras in pre-implantation embryos. Here, we show that the conventional hPSCs undergo rapid apoptosis when injected into mouse pre-implantation embryos. While, forced-expression of BMI1, a polycomb factor in hPSCs overcomes the apoptosis and enables hPSCs to integrate into mouse pre-implantation embryos and subsequently contribute to chimeras with both embryonic and extra-embryonic tissues. In addition, BMI1 also enables hPSCs to integrate into pre-implantation embryos of other species, such as rabbit and pig. Notably, BMI1 high expression and anti-apoptosis are also indicators for naïve hPSCs to form chimera in mouse embryos. Together, our findings reveal that the apoptosis is an initial barrier in interspecies chimerism using hPSCs and provide a rational to improve it. Conventional human pluripotent stem cells (hPSCs) fail to contribute to interspecies chimaeras when injected into mouse blastocysts. Here the authors show that forced expression of BMI1 overcomes apoptosis of hPSCs in blastocysts of mouse, rabbit and pig allowing them to contribute to chimaeras.

Yttrium–magnesium alloys are commonly employed as processing additives in magnesium alloy materials. Incorporating yttrium into magnesium alloys via Y-Mg intermediate alloys not only minimizes oxidation and burn-off loss but also simplifies operational procedures. Utilizing yttrium–magnesium alloys ensures a stable composition and reliable quality of magnesium alloy products, while contributing to reduced production costs and minimized environmental pollution. In this study, a molten salt co-reduction method was developed for the preparation Y-Mg intermediate alloys. The electrochemical co-reduction behaviors of Y(III) and Mg(II), as well as the transient states of Y-Mg intermediate alloys, were systematically investigated by transient electrochemical techniques. Results indicated that the reduction of Y(III) at the molybdenum (Mo) cathode is a reversible electrochemical process, whereas the reduction of Mg(II) is irreversible and diffusion-controlled. The diffusion coefficient of Y(III) and Mg(II) in the fluoride salt at 1000 °C were determined to be 3.98 × 10−5 cm2/s and 1.16 × 10−3 cm2/s, respectively. Electrochemical calculations revealed that the reduction of Y(III) involves a single-step transfer of three electrons, while Mg(II) involves a single-step transfer of two electrons. The corresponding electrode reactions are Y(III) + 3e−→Y and Mg(II) + 2e−→Mg, respectively. A Y-Mg alloy sample prepared by constant-current molten salt electrolysis primarily consists of the MgY phase with a composition of 88.38 wt% yttrium and 11.62 wt% magnesium.

Dear Editor, The traditional approach for the generation of gene- targeted animals is homologous recombination （HR） conducted in embryonic stem （ES） cells followed by chimera technology, such as in mice and rats, or HR in somatic cells combined with nuclear transfer in those animals in which germline-competent ES cells are not available, such as pigs, sheep, goats, and cattle. How- ever, no germline-competent rabbit ES cells are yet available. Furthermore, the efficiency of rabbit cloning is so low that no gene-targeted rabbits have been produced thus far by somatic cell nuclear transfer aooroaches.

The porcine pluripotent cells that can generate germline chimeras have not been developed. The Oct4 promoter-based fluorescent reporter system, which can be used to monitor pluripotency, is an important tool to generate authentic porcine pluripotent cells. In this study, we established a porcine Oct4 reporter system, wherein the endogenous Oct4 promoter directly controls red fluorescent protein (RFP). 2A-tdTomato sequence was inserted to replace the stop codon of the porcine Oct4 gene by homogenous recombination (HR). Thus, the fluorescence can accurately show the activation of endogenous Oct4. Porcine fetal fibroblast (PFF) lines with knock-in (KI) of the tdTomato gene in the downstream of endogenous Oct4 promoter were achieved using the CRISPR/CAS9 system. Transgenic PFFs were used as donor cells for somatic cell nuclear transfer (SCNT). Strong RFP expression was detected in the blastocysts and genital ridges of SCNT fetuses but not in other tissues. Two viable transgenic piglets were also produced by SCNT. Reprogramming of fibroblasts from the fetuses and piglets by another round of SCNT resulted in tdTomato reactivation in reconstructed blastocysts. Result indicated that a KI porcine reporter system to monitor the pluripotent status of cells was successfully developed.

Cas12a can process multiple sgRNAs from a single transcript of CRISPR array, conferring advantages in multiplexed base editing when incorporated into base editor systems, which is extremely helpful given that phenotypes commonly involve multiple genes or single-nucleotide variants. However, multiplexed base editing through Cas12a-derived base editors has been barely reported, mainly due to the compromised efficiencies and restricted protospacer-adjacent motif (PAM) of TTTV for wild-type Cas12a. Here, we develop Cas12a-mediated cytosine base editor (CBE) and adenine base editor (ABE) systems with elevated efficiencies and expanded targeting scope, by combining highly active deaminases with Lachnospiraceae bacterium Cas12a (LbCas12a) variants. We confirm that these CBEs and ABEs can perform efficient C-to-T and A-to-G conversions, respectively, on targets with PAMs of NTTN, TYCN, and TRTN. Notably, multiplexed base editing can be conducted using the developed CBEs and ABEs in somatic cells and embryos. These Cas12a variant-mediated base editors will serve as versatile tools for multiplexed point mutation, which is notably important in genetic improvement, disease modeling, and gene therapy. Cas12a-mediated cytosine (CBE) and adenine base editor (ABE) systems with elevated efficiencies and expanded target scopes are developed, by combining highly active deaminases with Cas12a variants, enCas12a, RR and RVR, recognizing non-TTTV PAMs.