A self-regulated network for dynamically balancing multiple precursors in complex biosynthetic pathways.

Microbial synthesis has emerged as a promising and sustainable alternative to traditional chemical synthesis and plant extraction. However, the competition between synthetic pathways and central metabolic pathways for cellular resources may impair final production efficiency. Moreover, when the synthesis of target product requires multiple precursors from the same node, the conflicts of carbon flux have further negative impacts on yields. In this study, a self-regulated network was developed to relieve the competition of precursors in complex synthetic pathways. Using 4-hydroxycoumarin (4-HC) synthetic pathway as a proof of concept, we employed an intermediate as a trigger to dynamically rewire the metabolic flux of pyruvate and control the expression levels of genes in 4-HC synthetic pathway, achieving self-regulation of multiple precursors and enhanced titer. Transcriptomic analysis results additionally demonstrated that the gene transcriptional levels of both pyruvate kinase PykF and synthetic pathway enzyme SdgA dynamically changed according to the intermediate concentrations. Overall, our work established a self-regulated network to dynamically balance the metabolic flux of two precursors in 4-HC biosynthesis, providing insight into balancing biosynthetic pathways where multiple precursors compete and interfere with each other.

Investigating and Engineering an 1,2-Propanediol-Responsive Transcription Factor-Based Biosensor.

Transcription factor (TF)-based biosensors have arisen as powerful tools in the advancement of metabolic engineering. However, with the emergence of numerous bioproduction targets, the variety of applicable TF-based biosensors remains severely limited. In this study, we investigated and engineered an 1,2-propanediol (1,2-PD)-responsive transcription activator, PocR, from Salmonella typhimurium to enrich the current biosensor repertoire. Heterologous characterization of PocR in E. coli revealed a significantly limited operational range and dynamic range, primarily attributed to the leaky binding between PocR and its corresponding promoters in the absence of the 1,2-PD inducer. Promiscuity characterization uncovered the minor responsiveness of PocR toward glycerol and 1,2-butanediol (1,2-BD). Using AlphaFold-predicted structure and protein mutagenesis, we preliminarily explored the underlying mechanism of PocR. Based on the investigated mechanism, we engineered a PcoR-F46R/G105D variant with an altered inducer specificity to glycerol, as well as a PocR-ARE (Q107A/S192R/A203E) variant with nearly a 4-fold higher dynamic range (6.7-fold activation) and a 20-fold wider operational range (0-20 mM 1,2-PD). Finally, we successfully converted PocR to a repressor through promoter engineering. Integrating the activation and repression functions established a versatile 1,2-PD-induced bifunctional regulation system based on PocR-ARE. Our work showcases the exploration and exploitation of an underexplored type of transcriptional activator capable of recruiting RNA polymerase. It also expands the biosensor toolbox by providing a 1,2-PD-responsive bifunctional regulator and glycerol-responsive activator.

The construction of a novel supplementary diagnostic model for patients with indeterminate HIV infection.

INTRODUCTION: The window period, defined as HIV nucleic acid test (NAT) reactivity but Western blot (WB) test inconclusive, is garnering more attention. Improving the detection efficiency of HIV high-risk populations in the window period is critical to reducing the risk of unanticipated transmission. The purpose of this study was to create an additional strategy for distinguishing indeterminate HIV infection cases. METHODS: Based on WB follow-up results, the individuals in this study were divided into persons in the HIV window period and persons without HIV. Plasma was analyzed using quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) to detect differentially expressed proteins (DEPs). The biological implications of these DEPs were investigated using enrichment analysis. Protein-protein interaction (PPI) analysis and LASSO regression were used to identify key proteins. The calibration curve, decision curve, and nomogram were utilized to create the model. RESULTS: Fifty-seven DEPs were screened out, with 33 up-regulated and 24 down-regulated in persons with HIV at window period. The most important Gene Ontology (GO) enrichment items are oxidoreductase activity and heme binding. Oxidoreductases account for half of the 10 main proteins identified from various DEPs. An auxiliary diagnostic model comprised of peroxiredoxin-2 (P32119), band 3 anion transport protein (P02730), and histone H2A type 1 (P0C0S8) was developed. The results of the confusion matrix parameters revealed that this diagnostic approach had strong practicability in distinguishing indeterminate HIV infection cases. CONCLUSIONS: The three DEPs identified and predicted by proteomics are useful for the supplemental identification of persons in the HIV window period.

Characteristics of Hepatitis B Virus, Hepatitis C Virus, and Syphilis Coinfection in People With HIV/AIDS Contracted Through Different Sources: Retrospective Study.

BACKGROUND: The burden of hepatitis B virus (HBV), hepatitis C virus (HCV), and syphilis coinfections remains disproportionately high among people living with HIV/AIDS. Hubei province is located in central China, where there are distinct regional characteristics of the distribution of people living with HIV/AIDS acquired via diverse transmission routes and the AIDS epidemic itself. OBJECTIVE: We aimed to estimate the magnitude of HBV, HCV, or syphilis coinfections among people living with HIV/AIDS with blood-borne transmission, which includes former paid blood donors, contaminated blood recipients, and intravenous drug users, as well as among people with sex-borne HIV transmission (including heterosexual people and men who have sex with men) and people with mother-to-child HIV transmission. METHODS: From January 2010 to December 2020, people living with HIV/AIDS were tested for hepatitis B surface antigen (HBsAg), HCV antibodies, and syphilis-specific antibodies. The positive patients were further tested for HBV markers, HBV DNA, and HCV RNA, and received a rapid plasma reagin circle card test. All people living with HIV/AIDS were first divided into transmission groups (blood, sex, and mother-to-child); then, people with blood-borne HIV transmission were divided into former paid blood donors, contaminated blood recipients, and intravenous drug users, while people with sex-borne HIV transmission were divided into heterosexual people and men who have sex with men. RESULTS: Among 6623 people living with HIV/AIDS, rates of chronic HCV infection were 80.3% (590/735) in former paid blood donors, 73.3% (247/337) in intravenous drug users, 57.1% (444/777) in contaminated blood recipients, 19.4% (21/108) in people with mother-to-child HIV transmission, 8.1% (240/2975) in heterosexual people, and 1.2% (21/1691) in men who have sex with men. Chronic HBV infection rates were similar among all people with blood-borne HIV transmission. However, compared to heterosexual people, the chronic HBV infection rate was greater in men who have sex with men (213/1691, 12.6% vs 308/2975, 10.4%; χ21=5.469; P=.02), although HBV exposure was less common (827/1691, 48.9% vs 1662/2975, 55.9%; χ21=20.982; P<.001). Interestingly, the combination of HBsAg and hepatitis B e antigen (HBeAg) was found in 11 patients with sex-borne HIV transmission, but in 0 people with blood-borne HIV transmission (11/196, 5.6% vs 0/521, 0%; χ21=29.695, P<.001). In people with sex-borne HIV transmission, the proportions of patients with a syphilis titer ≥1:16 and neurosyphilis were 8.6% (105/1227) and 7.8% (37/473), respectively, whereas these values were 0 in people with blood-borne HIV transmission. CONCLUSIONS: In people living with HIV/AIDS, HCV transmission intensity was significantly associated with specific exposure modes of blood or sexual contact. The rate of chronic HBV infection among men who have sex with men was higher than in any other population. Attention should be paid to the high prevalence of neurosyphilis in people living with HIV/AIDS who contract HIV by sexual intercourse.

JCAD deficiency attenuates activation of hepatic stellate cells and cholestatic fibrosis.

BACKGROUND/AIMS: Cholestatic liver diseases including primary biliary cholangitis (PBC) are associated with active hepatic fibrogenesis, which ultimately progresses to cirrhosis. Activated hepatic stellate cells (HSCs) are the main fibrogenic effectors in response to cholangiocyte damage. JCAD regulates cell proliferation and malignant transformation in nonalcoholic steatoheaptitis-associated hepatocellular carcinoma (NASH-HCC). However, its participation in cholestatic fibrosis has not been explored yet. METHODS: Serial sections of liver tissue of PBC patients were stained with immunofluorescence. Hepatic fibrosis was induced by bile duct ligation (BDL) in wild-type (WT), global JCAD knockout mice (JCAD-KO) and HSC-specific JCAD knockout mice (HSC-JCAD-KO), and evaluated by histopathology and biochemical tests. In situ-activated HSCs isolated from BDL mice were used to determine effects of JCAD on HSC activation. RESULTS: In consistence with staining of liver sections from PBC patients, immunofluorescent staining revealed that JCAD expression was identified in smooth muscle α-actin (α-SMA)-positive fibroblast-like cells and was significantly up-regulated in WT mice with BDL. JCAD deficiency remarkably ameliorated BDL-induced hepatic injury and fibrosis, as documented by liver hydroxyproline content, when compared to WT mice with BDL. Histopathologically, collagen deposition was dramatically reduced in both JCAD-KO and HSC-JCAD-KO mice compared to WT mice, as visualized by Trichrome staining and semi-quantitative scores. Moreover, JCAD deprivation significantly attenuated in situ HSC activation and reduced expression of fibrotic genes after BDL. CONCLUSION: JCAD deficiency effectively suppressed hepatic fibrosis induced by BDL in mice, and the underlying mechanisms are largely through suppressed Hippo-YAP signaling activity in HSCs.

Advancing microbial production through artificial intelligence-aided biology.

Microbial cell factories (MCFs) have been leveraged to construct sustainable platforms for value-added compound production. To optimize metabolism and reach optimal productivity, synthetic biology has developed various genetic devices to engineer microbial systems by gene editing, high-throughput protein engineering, and dynamic regulation. However, current synthetic biology methodologies still rely heavily on manual design, laborious testing, and exhaustive analysis. The emerging interdisciplinary field of artificial intelligence (AI) and biology has become pivotal in addressing the remaining challenges. AI-aided microbial production harnesses the power of processing, learning, and predicting vast amounts of biological data within seconds, providing outputs with high probability. With well-trained AI models, the conventional Design-Build-Test (DBT) cycle has been transformed into a multidimensional Design-Build-Test-Learn-Predict (DBTLP) workflow, leading to significantly improved operational efficiency and reduced labor consumption. Here, we comprehensively review the main components and recent advances in AI-aided microbial production, focusing on genome annotation, AI-aided protein engineering, artificial functional protein design, and AI-enabled pathway prediction. Finally, we discuss the challenges of integrating novel AI techniques into biology and propose the potential of large language models (LLMs) in advancing microbial production.

Discovery of a long-ranged charge order with 1/4 Ge1-dimerization in an antiferromagnetic Kagome metal.

Exotic quantum states arise from the interplay of various degrees of freedom such as charge, spin, orbital, and lattice. Recently, a short-ranged charge order (CO) was discovered deep inside the antiferromagnetic phase of Kagome magnet FeGe, exhibiting close relationships with magnetism. Despite extensive investigations, the CO mechanism remains controversial, mainly because the short-ranged behavior hinders precise identification of CO superstructure. Here, combining multiple experimental techniques, we report the observation of a long-ranged CO in high-quality FeGe samples, which is accompanied with a first-order structural transition. With these high-quality samples, the distorted 2 × 2 × 2 CO superstructure is characterized by a strong dimerization along the c-axis of 1/4 of Ge1-sites in Fe3Ge layers, and in response to that, the 2 × 2 in-plane charge modulations are induced. Moreover, we show that the previously reported short-ranged CO might be related to large occupational disorders at Ge1-site, which upsets the equilibrium of the CO state and the ideal 1 × 1 × 1 structure with very close energies, inducing nanoscale coexistence of these two phases. Our study provides important clues for further understanding the CO properties in FeGe and helps to identify the CO mechanism.

Evidence of electron interaction with an unidentified bosonic mode in superconductor CsCa2Fe4As4F2.

The kink structure in band dispersion usually refers to a certain electron-boson interaction, which is crucial in understanding the pairing in unconventional superconductors. Here we report the evidence of the observation of a kink structure in Fe-based superconductor CsCa2Fe4As4F2 using angle-resolved photoemission spectroscopy. The kink shows an orbital selective and momentum dependent behavior, which is located at 15 meV below Fermi level along the Γ - M direction at the band with dxz orbital character and vanishes when approaching the Γ - X direction, correlated with a slight decrease of the superconducting gap. Most importantly, this kink structure disappears when the superconducting gap closes, indicating that the corresponding bosonic mode (~ 9 ± 1 meV) is closely related to superconductivity. However, the origin of this mode remains unidentified, since it cannot be related to phonons or the spin resonance mode (~15 meV) observed by inelastic neutron scattering. The behavior of this mode is rather unique and challenges our present understanding of the superconducting paring mechanism of the bilayer FeAs-based superconductors.

Dynamic Metabolic Control: From the Perspective of Regulation Logic.

Establishing microbial cell factories has become a sustainable and increasingly promising approach for the synthesis of valuable chemicals. However, introducing heterologous pathways into these cell factories can disrupt the endogenous cellular metabolism, leading to suboptimal production performance. To address this challenge, dynamic pathway regulation has been developed and proven effective in improving microbial biosynthesis. In this review, we summarized typical dynamic regulation strategies based on their control logic. The applicable scenarios for each control logic were highlighted and perspectives for future research direction in this area were discussed.

Short- and long-term effects of antiretroviral therapy on peripheral regulatory CD4+/CD25hi/CD127low T lymphocytes in people living with HIV/AIDS

ABSTRACT The effect of antiretroviral therapy (ART) on CD4+/CD25hi/CD127low T lymphocyte changes in people living with HIV/AIDS (PLWHA) is still a matter of debate. From October 2015 to December 2019, peripheral blood from 70 cases of PLWHA were collected for the detection of CD4+/CD25hi/CD127low T lymphocytes by flow cytometry. Statistical analysis was performed to detect changes of CD4+/CD25hi/CD127low T lymphocytes in patients with different duration of ART and different treatment effects. We found that the number of CD4+/CD25hi/CD127low T lymphocytes in ART-naive PLWHA were lower than those in healthy volunteers (10.3±٦.٠ cells/uL vs 31.7±8.0 cells/uL, P < 0.05). CD4+/CD25hi/CD127low T lymphocyte counts increased to 17.8±٤.٠ cells/uL 6 months post-ART and 25.0±١١.٩ cells/uL 9 months post-ART, respectively (P < 0.05). There was no significant difference in CD4+/CD25hi/CD127low T lymphocyte counts between PLWHA who reached a complete immune reconstruction after ART and healthy volunteers. The growth of CD4+/CD25hi/CD127low T lymphocyte counts in patients who had baseline CD4 > 200 cells/uL was greater than those who had baseline CD4 ≤ 200 cells/uL (12.6±٤.٦ cells/uL vs 5.6±٥.٠ cells/uL, P = 0.027). CD4+/CD25hi/CD127low T lymphocyte counts were positively correlated with CD4+ T lymphocyte counts (r = 0.923, P < 0.001) and CD4+/CD8+ ratio (r = 0.741, P < 0.001), but were negatively correlated with HIV-VL (r = −0.648, P = 0.000). In conclusion, the results of the present study showed that changes in CD4+/CD25hi/CD127low T lymphocyte counts can be used to assess the effect of ART in PLWHA.