Molecular basis of Wnt biogenesis, secretion, and Wnt7-specific signaling.

Wnt proteins are enzymatically lipidated by Porcupine (PORCN) in the ER and bind to Wntless (WLS) for intracellular transport and secretion. Mechanisms governing the transfer of these low-solubility Wnts from the ER to the extracellular space remain unclear. Through structural and functional analyses of Wnt7a, a crucial Wnt involved in central nervous system angiogenesis and blood-brain barrier maintenance, we have elucidated the principles of Wnt biogenesis and Wnt7-specific signaling. The Wnt7a-WLS complex binds to calreticulin (CALR), revealing that CALR functions as a chaperone to facilitate Wnt transfer from PORCN to WLS during Wnt biogenesis. Our structures, functional analyses, and molecular dynamics simulations demonstrate that a phospholipid in the core of Wnt-bound WLS regulates the association and dissociation between Wnt and WLS, suggesting a lipid-mediated Wnt secretion mechanism. Finally, the structure of Wnt7a bound to RECK, a cell-surface Wnt7 co-receptor, reveals how RECKCC4 engages the N-terminal domain of Wnt7a to activate Wnt7-specific signaling.

Structural and functional insights into Spns2-mediated transport of sphingosine-1-phosphate.

Sphingosine-1-phosphate (S1P) is an important signaling sphingolipid that regulates the immune system, angiogenesis, auditory function, and epithelial and endothelial barrier integrity. Spinster homolog 2 (Spns2) is an S1P transporter that exports S1P to initiate lipid signaling cascades. Modulating Spns2 activity can be beneficial in treatments of cancer, inflammation, and immune diseases. However, the transport mechanism of Spns2 and its inhibition remain unclear. Here, we present six cryo-EM structures of human Spns2 in lipid nanodiscs, including two functionally relevant intermediate conformations that link the inward- and outward-facing states, to reveal the structural basis of the S1P transport cycle. Functional analyses suggest that Spns2 exports S1P via facilitated diffusion, a mechanism distinct from other MFS lipid transporters. Finally, we show that the Spns2 inhibitor 16d attenuates the transport activity by locking Spns2 in the inward-facing state. Our work sheds light on Spns2-mediated S1P transport and aids the development of advanced Spns2 inhibitors.

Structural Insights into Yeast Telomerase Recruitment to Telomeres.

Telomerase maintains chromosome ends from humans to yeasts. Recruitment of yeast telomerase to telomeres occurs through its Ku and Est1 subunits via independent interactions with telomerase RNA (TLC1) and telomeric proteins Sir4 and Cdc13, respectively. However, the structures of the molecules comprising these telomerase-recruiting pathways remain unknown. Here, we report crystal structures of the Ku heterodimer and Est1 complexed with their key binding partners. Two major findings are as follows: (1) Ku specifically binds to telomerase RNA in a distinct, yet related, manner to how it binds DNA; and (2) Est1 employs two separate pockets to bind distinct motifs of Cdc13. The N-terminal Cdc13-binding site of Est1 cooperates with the TLC1-Ku-Sir4 pathway for telomerase recruitment, whereas the C-terminal interface is dispensable for binding Est1 in vitro yet is nevertheless essential for telomere maintenance in vivo. Overall, our results integrate previous models and provide fundamentally valuable structural information regarding telomere biology.

GPR35 and mediators from platelets and mast cells in neutrophil migration and inflammation.

Neutrophil recruitment from circulation to sites of inflammation is guided by multiple chemoattractant cues emanating from tissue cells, immune cells, and platelets. Here, we focus on the function of one G-protein coupled receptor, GPR35, in neutrophil recruitment. GPR35 has been challenging to study due the description of multiple ligands and G-protein couplings. Recently, we found that GPR35-expressing hematopoietic cells respond to the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA). We discuss distinct response profiles of GPR35 to 5-HIAA compared to other ligands. To place the functions of 5-HIAA in context, we summarize the actions of serotonin in vascular biology and leukocyte recruitment. Important sources of serotonin and 5-HIAA are platelets and mast cells. We discuss the dynamics of cell migration into inflamed tissues and how multiple platelet and mast cell-derived mediators, including 5-HIAA, cooperate to promote neutrophil recruitment. Additional actions of GPR35 in tissue physiology are reviewed. Finally, we discuss how clinically approved drugs that modulate serotonin uptake and metabolism may influence 5-HIAA-GPR35 function, and we speculate about broader influences of the GPR35 ligand-receptor system in immunity and disease.

Structural insights into Pot1-ssDNA, Pot1-Tpz1 and Tpz1-Ccq1 Interactions within fission yeast shelterin complex.

The conserved shelterin complex caps chromosome ends to protect telomeres and regulate telomere replication. In fission yeast Schizosaccharomyces pombe, shelterin consists of telomeric single- and double-stranded DNA-binding modules Pot1-Tpz1 and Taz1-Rap1 connected by Poz1, and a specific component Ccq1. While individual structures of the two DNA-binding OB folds of Pot1 (Pot1OB1-GGTTAC and Pot1OB2-GGTTACGGT) are available, structural insight into recognition of telomeric repeats with spacers by the complete DNA-binding domain (Pot1DBD) remains an open question. Moreover, structural information about the Tpz1-Ccq1 interaction requires to be revealed for understanding how the specific component Ccq1 of S. pombe shelterin is recruited to telomeres to function as an interacting hub. Here, we report the crystal structures of Pot1DBD-single-stranded-DNA, Pot1372-555-Tpz1185-212 and Tpz1425-470-Ccq1123-439 complexes and propose an integrated model depicting the assembly mechanism of the shelterin complex at telomeres. The structure of Pot1DBD-DNA unveils how Pot1 recognizes S. pombe degenerate telomeric sequences. Our analyses of Tpz1-Ccq1 reveal structural basis for the essential role of the Tpz1-Ccq1 interaction in telomere recruitment of Ccq1 that is required for telomere maintenance and telomeric heterochromatin formation. Overall, our findings provide valuable structural information regarding interactions within fission yeast shelterin complex at 3' ss telomeric overhang.

Mechanism of Electrocatalytic H2 Evolution, Carbonyl Hydrogenation, and Carbon-Carbon Coupling on Cu.

Aqueous-phase electrocatalytic hydrogenation of benzaldehyde on Cu leads not only to benzyl alcohol (the carbonyl hydrogenation product), but Cu also catalyzes carbon-carbon coupling to hydrobenzoin. In the absence of an organic substrate, H2 evolution proceeds via the Volmer-Tafel mechanism on Cu/C, with the Tafel step being rate-determining. In the presence of benzaldehyde, the catalyst surface is primarily covered with the organic substrate, while H* coverage is low. Mechanistically, the first H addition to the carbonyl O of an adsorbed benzaldehyde molecule leads to a surface-bound hydroxy intermediate. The hydroxy intermediate then undergoes a second and rate-determining H addition to its α-C to form benzyl alcohol. The H additions occur predominantly via the proton-coupled electron transfer mechanism. In a parallel reaction, the radical α-C of the hydroxy intermediate attacks the electrophilic carbonyl C of a physisorbed benzaldehyde molecule to form the C-C bond, which is rate-determining. The C-C coupling is accompanied by the protonation of the formed alkoxy radical intermediate, coupled with electron transfer from the surface of Cu, to form hydrobenzoin.

A comparative study on 3D printing-assisted arthroscopic IDEAL point femoral tunnel positioning for anterior cruciate ligament reconstruction versus conventional arthroscopic positioning.

BACKGROUND: This study aimed to investigate the feasibility and precision of using a 3D-printed template for femoral tunnel placement in guiding the optimal positioning of the Internal anatomical stop and Low tension maintenance (IDEAL) bone tunnel during single-bundle anterior cruciate ligament (ACL) reconstruction. METHODS: A retrospective analysis was conducted on 40 patients who underwent arthroscopic single-bundle ACL reconstruction at our hospital between April 2021 and November 2021. In the direct vision group, the IDEAL bone tunnel was positioned using radiofrequency localization directly visualized at the stump. In the 3D-printed positioning group, preoperative CT scans and Digital Imaging and Communications in Medicine (DICOM) data were employed. Following the Quadrant method by Bernard, the femoral tunnel's depth was set at 25% and its height at 29%. Postoperative plain CT scans enabled the reconstruction of 3D models for both groups. The accuracy of femoral tunnel placement was then compared. RESULTS: The central locations of the bone tunnels in the direct vision group were at a mean depth of 25.74 ± 1.84% and a height of 29.22 ± 2.97%. In the 3D printing localization group, these values were 25.39 ± 2.98% for depth and 28.89 ± 2.50% for height, respectively. No significant differences were found in tunnel positioning between the groups. Both groups demonstrated statistically significant improvements in International Knee Documentation Committee Subjective Knee Form (IKDC) and Lysholm scores postoperatively, with no significant differences observed 12 months post-surgery. CONCLUSION: The findings of this study suggest that 3D printing-assisted arthroscopic IDEAL point femoral tunnel positioning and conventional arthroscopic positioning are feasible and effective for ACL reconstruction. Using 3D printing technology to design femoral anchor points in ACL reconstruction allows for the customization of anterior fork reconstruction and precise bone tunnel positioning, supporting the goal of individualized and accurate reconstruction.

Air pollution-induced health impacts and health economic losses in China driven by US demand exports.

Understanding how trade between regions or countries drives the transfer of air pollution has attracted considerable interest recently, but few studies have explored the various transfer pathways or evaluated economic losses due to the health impact of such air pollution. Here, we assess the air pollutant emissions and related health impacts and economic losses in China caused by export trade due to US demand by combining the linked multi-regional input-output (MRIO) model, GEOS-Chem model, integrated exposure-response model, and the willingness to pay method. We show that the air pollutant emissions embedded in China's export due to the US demand reached 5792.38 Kt in 2012 (2.48% of the total), which includes direct exports of intermediate (40.27%) and final (33.61%) products and indirect exports of intermediate products via domestic provinces (16.43%, domestic spillover) and other countries (9.69%, foreign spillover). The resulting increase in PM2.5 (<2.8 µg m-3) leads to additional 27,963 deaths in 30 provinces, with a higher death toll in coastal areas and the corresponding economic loss was higher in more developed regions and reached USD 2.08 billion. This study highlights the region-different air pollution and health impacts in China embedded in the US-demand trade, and provides a framework for the analysis of health and economic losses hidden in global trade, particularly between developing and developed countries.

[Status and Development of Intelligent Cardiopulmonary Resuscitation Equipment].

The current status of cardiopulmonary resuscitation in China were introduced. The function and working principle of cardiopulmonary resuscitation equipment were described. The research status of cardiopulmonary resuscitation equipment was summarized. The main problem existing in cardiopulmonary resuscitation equipment were analyzed. Finally, according to the main technical direction involved in the conception, the existing technologies were reviewed from four aspects: path planning, human-computer interaction, automatic defibrillation and intelligent compression.

Design of highly sensitive and selective ethanol sensor based on α-Fe2O3/Nb2O5 heterostructure.

The introduction of heterostructures is a new approach in gas sensing due to their easy and quick transport of charges. Herein, facile hydrothermal and solid-state techniques are employed to synthesize an α-Fe2O3/Nb2O5 heterostructure. The morphology, microstructure, crystallinity and surface composition of the synthesized heterostructures are investigated by scanning electron microscope, transmission electron microscope, x-ray diffraction, x-ray photoelectron spectroscopy and Brunauer-Emmett-Teller analyses. The successful fabrication of the heterostructures was achieved via the mutual incorporation of α-Fe2O3 nanorods with Nb2O5 interconnected nanoparticles (INPs). A sensor based on the α-Fe2O3(0.09)/Nb2O5 heterostructure with a high surface area exhibited enhanced gas-sensing features, maintaining high selectivity and sensitivity, and a considerable recovery percentage towards ethanol gas. The sensing response of the α-Fe2O3(0.09)/Nb2O5 heterostructure at lower operating temperature (160 °C) is around nine times higher than a pure Nb2O5 (INP) sensor at 180 °C with the flow of 100 ppm ethanol gas. The sensors also show excellent selectivity, good long-term stability and a rapid response/recovery time (8s/2s, respectively) to ethanol. The superior electronic conductivity and upgraded sensitivity performance of gas sensors based on the α-Fe2O3(0.09)/Nb2O5 heterostructure are attributed due to its unique structural features, high specific surface area and the synergic effect of the n-n heterojunction. The promising results demonstrate the potential application of the α-Fe2O3(0.09)/Nb2O5 heterostructure as a good sensing material for the fabrication of ethanol sensors.

Interdomain communication in the phosphatidylcholine regulatory enzyme, CCTα, relies on a modular αE helix.

CTP:phosphocholine cytidylyltransferase (CCT), the rate-limiting enzyme in phosphatidylcholine (PC) synthesis, is an amphitropic enzyme that regulates PC homeostasis. Recent work has suggested that CCTα activation by binding to a PC-deficient membrane involves conformational transitions in a helix pair (αE) that, along with a short linker of unknown structure (J segment), bridges the catalytic domains of the CCTα dimer to the membrane-binding (M) domains. In the soluble, inactive form, the αE helices are constrained into unbroken helices by contacts with two auto-inhibitory (AI) helices from domain M. In the active, membrane-bound form, the AI helices are displaced and engage the membrane. Molecular dynamics simulations have suggested that AI displacement is associated with hinge-like bending in the middle of the αE, positioning its C terminus closer to the active site. Here, we show that CCTα activation by membrane binding is sensitive to mutations in the αE and J segments, especially within or proximal to the αE hinge. Substituting Tyr-213 within this hinge with smaller uncharged amino acids that could destabilize interactions between the αE helices increased both constitutive and lipid-dependent activities, supporting a link between αE helix bending and stimulation of CCT activity. The solvent accessibilities of Tyr-213 and Tyr-216 suggested that these tyrosines move to new partially buried environments upon membrane binding of CCT, consistent with a folded αE/J structure. These data suggest that signal transduction through the modular αE helix pair relies on shifts in its conformational ensemble that are controlled by the AI helices and their displacement upon membrane binding.

Two-Dimensional SnSe2 /CNTs Hybrid Nanostructures as Anode Materials for High-Performance Lithium-Ion Batteries.

Tin diselenide (SnSe2 ), as an anode material, has outstanding potential for use in advanced lithium-ion batteries. However, like other tin-based anodes, SnSe2 suffers from poor cycle life and low rate capability due to large volume expansion during the repeated Li+ insertion/de-insertion process. This work reports an effective and easy strategy to combine SnSe2 and carbon nanotubes (CNTs) to form a SnSe2 /CNTs hybrid nanostructure. The synthesized SnSe2 has a regular hexagonal shape with a typical 2D nanostructure and the carbon nanotubes combine well with the SnSe2 nanosheets. The hybrid nanostructure can significantly reduce the serious damage to electrodes that occurs during electrochemical cycling processes. Remarkably, the SnSe2 /CNTs electrode exhibits a high reversible specific capacity of 457.6 mA h g-1 at 0.1 C and 210.3 mA h g-1 after 100 cycles. At a cycling rate of 0.5 C, the SnSe2 /CNTs electrode can still achieve a high value of 176.5 mA h g-1 , whereas a value of 45.8 mA h g-1 is achieved for the pure SnSe2 electrode. The enhanced electrochemical performance of the SnSe2 /CNTs electrode demonstrates its great potential for use in lithium-ion batteries. Thus, this work reports a facile approach to the synthesis of SnSe2 /CNTs as a promising anode material for lithium-ion batteries.

Fungi short-chain carboxylate transporter: shift from microbe hereditary functional component to metabolic engineering target.

Short-chain carboxylic acids and their derivatives are widely utilized in all aspects of our daily life. Given their specific functional groups, these molecules are also utilized in fine chemical synthesis. The traditional petroleum-based carboxylate production methods are restricted by petrol shortage and environmental pollution. Renowned for their more sustainable processes than traditional methods, biotechnological methods are preferred alternatives and have attracted increasing attention. However, the industrial application of biotechnological methods is currently limited by low factors: low productivity and low yield. Therefore, understanding the regulation of carboxylate accumulation will greatly enhance the industrial biotechnological production of short-chain carboxylate acids. The carboxylate transporter plays a crucial role in transmembrane uptake and secretion of carboxylate; therefore, regulating these transporters is of high academic and application relevance. This review concentrates on the physiological roles, regulation mechanisms, and harnessing strategies of Jen and AcpA orthologs in fungi, which provide potential clues for the biotechnological production of short-chain carboxylic acids with high efficiency.

[Application of Quality Control Circle in Reducing Incidence of Leakage in Digestive Endoscopy].

To study the role of the activity in reducing the incidence of water leakage in digestive endoscopy, through the activity of the quality control circle (QCC), the reasons for water leakage and high maintenance cost are analyzed deeply. Then, the author formulated and implemented targeted measures to intervene in the non-standard aspects of daily management of endoscopy. Then targeted measures has been taken to improve the nonstandard daily management.Results demonstrate that the application of the quality control circle can significantly reduce the incidence of water leakage in the digestive endoscope and control the other faults caused by the leakage. This not only reduces the high maintenance cost of endoscopy, improves the utilization rate and turnover rate of endoscopy, but also fully explores personal potential, enhances team cohesion, and improves quality management awareness.

Applying pathway engineering to enhance production of alpha-ketoglutarate in Yarrowia lipolytica.

α-Ketoglutarate (α-KG), one of short-chain carboxylates of high commercial relevance, has been widely used in food, medicine, chemical, and cosmetic fields. Compared to other carboxylates, α-KG occupies key positions in the tricarboxylate cycle (TCA cycle) and amino acid metabolic pathway, the over-accumulation of α-KG is restricted both by tighter carbon and nitrogen regulation process. Biotechnology production of α-KG on large industrial level has been impeded by many obstacles. This review aims at highlighting and stating recent efforts toward improving the yield and titer of α-KG in the strains of Yarrowia lipolytica to reach industrial relevance. Fermentation process optimization concerning feedstock utilization, dissolved oxygen controlling, pH manipulation and establishment of fed-batch process, have been assessed and evaluated. Moreover, pathway engineering routes have been applied for enhancing carbon commitment to α-KG, blocking competing pathways, regenerating of co-factors and regulating of carboxylate transporters to facilitate production and accumulation of α-KG. Although no engineered strain can satisfy the requirements of industrial production relevance to date, these strategies provide many clues for accelerating strain development for α-KG production.

Proteomic analysis of the response of α-ketoglutarate-producer Yarrowia lipolytica WSH-Z06 to environmental pH stimuli.

During bioproduction of short-chain carboxylates, a shift in pH is a common strategy for enhancing the biosynthesis of target products. Based on two-dimensional gel electrophoresis, comparative proteomics analysis of general and mitochondrial protein samples was used to investigate the cellular responses to environmental pH stimuli in the α-ketoglutarate overproducer Yarrowia lipolytica WSH-Z06. The lower environmental pH stimuli tensioned intracellular acidification and increased the level of reactive oxygen species (ROS). A total of 54 differentially expressed protein spots were detected, and 11 main cellular processes were identified to be involved in the cellular response to environmental pH stimuli. Slight decrease in cytoplasmic pH enhanced the cellular acidogenicity by elevating expression level of key enzymes in tricarboxylic acid cycle (TCA cycle). Enhanced energy biosynthesis, ROS elimination, and membrane potential homeostasis processes were also employed as cellular defense strategies to compete with environmental pH stimuli. Owing to its antioxidant role of α-ketoglutarate, metabolic flux shifted to α-ketoglutarate under lower pH by Y. lipolytica in response to acidic pH stimuli. The identified differentially expressed proteins provide clues for understanding the mechanisms of the cellular responses and for enhancing short-chain carboxylate production through metabolic engineering or process optimization strategies in combination with manipulation of environmental conditions.

Automated interventricular septum segmentation for black-blood myocardial T2* measurement in thalassemia.

PURPOSE: To develop and validate an automated segmentation method that extracts the interventricular septum (IS) from myocardial black-blood images for the T2* measurement in thalassemia patients. MATERIALS AND METHODS: A total of 144 thalassemia major patients (age range, 11-51 years; 73 males) were scanned with a black-blood multi-echo gradient-echo sequence using a 1.5 Tesla Siemens Sonata system (flip angle 20°, sampling bandwidth 810 Hz/pixel, voxel size 1.56 × 1.56 × 10 mm(3) and variable fields of view (20-30) × 40 cm(2) depending on patient size). The improved Chan-Vese model with an automated initialization by the circular Hough transformation was implemented to segment the endocardial and epicardial margins of the left ventricle (LV). Consequently, the IS was extracted by analyzing the anatomical relation between the LV and the blood pool of the right ventricle, identified by intensity thresholding. The proposed automated IS segmentation (AISS) method was compared with the conventional manual method by using the Bland-Altman analysis and the coefficient of variation (CoV). RESULTS: The T2* measurements using the AISS method were in good agreement with those manually measured by experienced observers with a mean difference of 1.71% and a CoV of 4.15% (P < 0.001). CONCLUSION: Black-blood myocardial T2* measurement can be fully automated with the proposed AISS method.

Posttraumatic growth and demoralization after cancer: The effects of patients' meaning-making.

OBJECTIVE: It is common for patients to experience positive and negative psychological changes (e.g., posttraumatic growth or demoralization) after being diagnosed with cancer. Although demoralization and posttraumatic growth are both related to meaning-making, little attention has been paid to the associations among these concepts. The current study investigated the relationship between demoralization, posttraumatic growth, and meaning-making (focusing on sense-making and benefit-finding during the experience of illness) in cancer patients. METHOD: Some 200 cancer patients (with lung cancer, lymphoma, or leukemia) at the MacKay Memorial Hospital in New Taipei completed the Demoralization Scale-Mandarin Version (DS-MV), the Chinese Posttraumatic Growth Inventory (CPTGI), and a self-designed questionnaire for assessing sense-making and benefit-finding. RESULTS: Demoralization was negatively correlated with posttraumatic growth, sense-making, benefit-finding, and time-since-diagnosis. Multiple regression analysis showed that meaning-making had different effects on demoralization and posttraumatic growth. The interactions of sense-making with either benefit-finding or time-since-diagnosis significantly predicted demoralization. Individuals with relatively higher sense-making and benefit-finding or shorter time-since-diagnosis experienced less demoralization. SIGNIFICANCE OF RESULTS: The suffering of cancer may turn on the psychological process of demoralization, posttraumatic growth, and meaning-making in patients. Cancer patients who evidenced higher posttraumatic growth experienced less demoralization. Trying to identify positive changes in the experience of cancer may be a powerful way to increase posttraumatic growth. As time goes by, patients experienced less demoralization. Facilitating sense-making can have similar effects. Cancer patients with less benefit-finding experience higher demoralization, but sense-making buffers this effect.

A correlational study of suicidal ideation with psychological distress, depression, and demoralization in patients with cancer.

PURPOSE: This study aims to study the effects of depression and demoralization on suicidal ideation and to determine the feasibility of the Distress Thermometer as a screening tool for patients with cancer who experience depression and demoralization, and thus to establish a model screening process for suicide prevention. METHODS: Purposive sampling was used to invite inpatients and outpatients with lung cancer, leukemia, and lymphoma. Two hundred participants completed the questionnaire, which included the Distress Thermometer (DT), Patient Health Questionnaire-9 (PHQ-9), Demoralization Scale-Mandarin Version (DS-MV), and Beck Scale for Suicide Ideation. All data obtained were analyzed using SPSS 18.0 and SAS 9.3. RESULTS: Tobit regression analysis showed that demoralization influenced suicidal ideation more than depression did (t = 2.84, p < 0.01). When PHQ-9 ≥ 10 and DS-MV ≥42 were used as criteria for the DT, receiver operating characteristic analysis revealed that the AUC values were 0.77-0.79, with optimal cutoff points for both of DT ≥5; sensitivity 76.9 and 80.6 %, respectively; and specificity of 73.9 and 72.2 %, respectively. CONCLUSIONS: Demoralization had more influence on suicidal ideation than depression did. Therefore, attention should be paid to highly demoralized patients with cancer or high demoralization comorbid with depression for the purposes of suicide evaluation and prevention. The DT scale (with a cutoff of ≥5 points) has discriminative ability as a screening tool for demoralization or depression and can also be used in clinical settings for the preliminary screening of patients with cancer and high suicide risk.

Construction of brewing-wine Aspergillus oryzae pyrG- mutant by pyrG gene deletion and its application in homology transformation.

pyrG(-) host cells are indispensable for pyrG(-) based transformation system. Isolations of pyrG(-) host cells by random mutations are limited by time-consuming, unclear genetic background and potential interferences of homogenous recombination. The purpose of this study was to construct brewing-wine Aspergillus oryzae pyrG(-) mutant by site-directed mutation of pyrG gene deletion which would be used as a host for further transformation. pMD-pyrGAB, a vector carrying pyrG deletion cassette, was used to construct pyrG(-) mutant of A. oryzae. Three stable pyrG deletion mutants of A. oryzae were isolated by resistant to 5-fluoroorotic acid and confirmed by polymerase chain reaction analysis, indicating that pyrG was completely excised. The ΔpyrG mutants were applied as pyrG(-) host cells to disrupt xdh gene encoding xylitol dehydrogenase, which involves in xylitol production of A. oryzae. The xdh disruption mutants were efficiently constructed by transforming a pMD-pyrG-xdh disruption plasmid carrying pyrG, and the produced xylitol concentration of the Δxdh mutant was three times as much as that of the ΔpyrG recipient. Site-directed pyrG gene deletion is thus an effective way for the isolation of pyrG(-) host cells, and the established host-vector system could be applied in further functional genomics analysis and molecular breeding of A. oryzae.