APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
·
2020
·Vol. 104
·期 16
·7079-7091
·
IF 3.53
Wang Shengli,Han Jing,Xia Jiale,Hu Yanru,Shi Liang,Ren Ang,Zhu Jing,Zhao Mingwen
Abstract Identifying new and economical means to utilize diverse lignocellulosic biomass is an urgent task. Ganoderma lucidum is a well-known edible and medicinal basidiomycete with an excellent ability to degrade a wide range of cellulosic biomass, and its nutrient use efficiency is closely related to the activity of extracellular cellulase. Intracellular nicotinamide adenine dinucleotide (NAD + ) biosynthesis is controlled in response to nutritional status, and NAD + is an essential metabolite involved in diverse cellular processes. Nicotinamide mononucleotide adenylyltransferase (NMNAT) is a common enzyme in three NAD + synthesis pathways. In this study, a homologous gene of nmnat was cloned from G. lucidum and two G. lucidum overexpression strains, OE::nmnat4 and OE::nmnat19, were constructed using an Agrobacterium tumefaciens -mediated transformation method. The G. lucidum nmnat overexpression strains showed obviously increased colony growth on different carbon sources, and intracellular Ca 2+ concentrations in the G. lucidum OE::nmnat4 and OE::nmnat19 strains were increased by 2.04- and 2.30-fold, respectively, compared with those in the wild-type (WT) strains. In the G. lucidum OE::nmnat4 and OE::nmnat19 strains, endo- β -glucanase (CMCase) activity increased by approximately 2.8- and 3-fold, while β -glucosidase ( p NPGase) activity increased by approximately 1.9- and 2.1-fold, respectively, compared with the activity in the WT strains. Furthermore, overexpression of NAD + synthesis pathways was found to elicit cellulase production by increasing the intracellular Ca 2+ concentration. In summary, this study is the first to demonstrate that increased intracellular NAD + contents through overexpression of the nmnat gene of NAD + synthesis pathways may increase cellulase production by increasing intracellular Ca 2+ concentrations in G. lucidum . Key points
The concentration of NAD
+
influences cellulase production in G. lucidum.
The concentration of NAD
+
influences the intracellular Ca
2+
concentration in G. lucidum.
The concentration of NAD
+
influences cellulase production by eliciting a change in intracellular Ca
2+
in G. lucidum.
研究领域:
微生物学
·
生物技术
·
酶学
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DOI
PubMed
高影响因子
Plant Signaling & Behavior
·
2022
·
IF 2.734
Rui Zhou,Menglong Fan,Mei Zhao,Xinqiang Jiang,Qinghua Liu
Leaves are the main vegetative organs of the aboveground part of plants and play an important role in plant morphogenesis. KNOTTED-LIKE HOMEOBOX (KNOX) plays a crucial role in regulating leaf cell fate and maintaining leaf development. In this study, we analyzed LtKNOX1 from Lilium tsingtauense and illustrated its function in transgenic plants. Tissue-specific expression analysis indicated that LtKNOX1 was highly expressed in stems, young flower buds, and shoot apical meristems (SAMs). Ectopic overexpression of LtKNOX1 in Nicotiana benthamiana suggested that transformants with mild phenotypes were characterized by foliar wrinkles and mildly curled leaves; transformants with intermediate phenotypes showed severely crimped blades and narrow leaf angles, and the most severe phenotypes lacked normal SAMs and leaves. Moreover, the expression levels of genes involved in the regulation of KNOX in transgenic plants were detected, including ASYMMETRIC LEAVES1, PIN-FORMED 1, GA20-oxidase, CUP-SHAPED COTYLEDON 2, CLAVATA 1 and WUSCHEL(WUS), and the expression of other genes were down-regulated except WUS. This study contributes to our understanding of the LtKNOX1 function.
研究领域:
植物学
·
遗传学
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DOI
PubMed
高影响因子
CATALYSIS LETTERS
·
2018
·Vol. 148
·期 6
·1714-1722
·
IF 2.911
Luo Wei,Deng Xin-Xing,Huo Jiao,Ruan Tao,Gong Zhi-Wei,Yan Jia-Bao,Yang Zhong-Hua,Quan Can,Cui Zhi-Fang
Abstract Microalgae are excellent biocatalyst candidates for photo-biocatalytic-asymmetric reduction of prochiral ketones to produce enantiomer alcohols. In the biocatalysis asymmetric reduction of carbonyl group process, the cofactor, NADPH, plays a key role. So the current study focused on the effect of NADPH availability on the reduction. Photosynthesis is a main pathway for NADPH regeneration in microalgae, and the ferredoxin-NADP + oxidoreductase (FNR) is the key enzyme in this process. In this work, we constructed an engineered cyanobacterium Synechocystis sp. PCC6803:: Ω- P petE-petH to overexpress FNR gene petH to strengthen NADPH regeneration. The results show that the intracellular NADPH content was increased 80.8% compared to the wild type. Applying the Synechocystis sp. PCC6803:: Ω- P petE-petH to catalyze the model asymmetric reduction reaction (reduction of acetophenone to S -phenylethanol), the yield was improved 61.2% compared to the wild-type. This work makes the microalgal biocatalysis asymmetric reduction process become more preponderant via providing a new route to enhance the regeneration of cofactor NADPH. Graphical Abstract Putative metabolic process of nicotinamide adenine dinucleotide phosphate (NADPH) regeneration and the photo-biocatalytic-asymmetric reduction reaction of prochiral ketones in microalgal cells.
研究领域:
生物化学
·
合成生物学
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DOI
高影响因子
PROCESS BIOCHEMISTRY
·
2024
·
IF 3.7
Long Zhang,Jinna Cui,Wei Liu,Zhanying Liu
Various protein secretion systems of gram-negative Escherichia coli ( E. coli ) are well known, but the secretion pathway of endoglucanase (BcsZ) is unknown. To further elucidate the secretion mechanism, we analyzed the transcriptome data of five ruminal cellulolytic E. coli strains. Gene Ontology (GO) enrichment analysis revealed that the differentially expressed genes (DEGs) upregulated in all five strains were significantly enriched in the membrane. Nine DEGs related to membrane transporters were overexpressed in E. coli JBZ-DH5α (overexpressing BcsZ in the genome of E. coli DH5α). The results revealed that the overexpression of the b4332 ( yjiJ ), b2142 ( yohK ), and b2377 ( yfdY ) genes significantly reduced the growth and BcsZ concentration of the strain. The overexpression of the genes b3533 ( bcsA ), b3009 ( yghB ), and b3522 ( yhjD ) did not significantly affect strain growth or the BcsZ concentration. The overexpression of the b1859 ( znuB ), b0458 ( ylaC ), and b1195 ( ymgE ) genes increased the secretion of BcsZ but had no significant effect on strain growth. In conclusion, the yjiJ , yohK , yfdY , znuB , ylaC , and ymgE genes are involved in the secretion of E. coli BcsZ. These findings provide insight into the secretion mechanism of E. coli BcsZ.
研究领域:
微生物学
·
分子生物学
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高影响因子
JOURNAL OF OBSTETRICS AND GYNAECOLOGY RESEARCH
·
2021
·Vol. 47
·期 6
·2196-2203
·
IF 1.73
Lianhua Wang,Qiming Huang,Qingqing Lin,Lvxuan Chen,Qiyang Shi
Aim Endometrial cancer (EC) is a common type of malignant gynecological cancer. Small nucleolar RNA host gene 9 (SNHG9) has been discovered to serve a role in several types of cancer; however, the role of SNHG9 in EC remains unclear. The present study aimed to investigate the effects of lncRNA SNHG9 on cell proliferation and glycolysis in EC cells. Methods SNHG9 and hexokinase 2 (HK2) mRNA expression levels were measured by reverse transcription-quantitative PCR. Glucose consumption and lactate production were detected by the glycolysis cell-based assay kit. Cell Counting Kit-8 and colony formation assays were conducted to detect cell proliferation. The knockdown experiments of SNHG9 and HK2 were carried out by transfection of corresponding small interference RNAs (siRNA). The SNHG9-overexpressed plasmid was transfected into the cells to upregulate SNHG9. HK2 protein levels were analyzed by western blotting assay. Results SNHG9 expression levels were significantly upregulated in EC tissues and cells. The knockdown of SNHG9 subsequently effectively attenuated cell proliferation and glycolysis in vitro, while SNHG9 overexpression reported the opposite effects. Notably, the transfection of 2-DG partially reversed the promoting effect of SNHG9 on glycolysis. Downregulation of HK2 markedly decreased cell proliferation and glycolysis in EC cells antagonized SNHG9. Conclusion Either downregulation of SNHG9 or HK2 inhibits EC cell proliferation and glycolysis via repressing EC cell proliferation and glycolysis.
研究领域:
肿瘤学
·
分子生物学
·
癌症遗传学
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DOI
PubMed
高影响因子
Revista Brasileira de Farmacognosia-Brazilian Journal of Pharmacognosy
·
2024
·
IF 1.4
Xie Huilong,Fu Yunrong,Lin Zhisheng,Wang Jiayi,Wu Jingyu,You Xiuhua,Xie Jieming
This study explores the synergistic effect of T-CUS 245C combined with cyclosporin A, and its possible mechanism. T-CUS 245C is an immunotoxin conjugated with tastuzumab and recombinant cucurmosin 245C (CUS 245C ). Cyclosporin A is a calcium antagonist mainly used as an immune-suppressive agent, which enhances the cytotoxicity of immunotoxins through an unknown mechanism. The sulforhodamine B assay, flow cytometry, DNA fragmentation apoptosis assay, and western blot analysis were used to evaluate the synergistic effects of cyclosporin A combined with T-CUS 245C on proliferation inhibition and apoptosis induction in SK-OV-3 ovarian cancer cells with HER-2-overexpressing. Meanwhile, using confocal microscopy and flow cytometry, the effect of cyclosporin A on the intracellular distribution of T-CUS 245C was observed and quantified. The results of this study showed that the combination index of cyclosporin A combined with T-CUS 245C is less than 0.7, indicating that cyclosporin A significantly enhances the proliferation inhibition and apoptosis induction induced by T-CUS 245C in SK-OV-3 cells. The confocal microscopy and flow cytometry results show that cyclosporin A effectively increases the dispersion of T-CUS 245C in the cytoplasm by promoting the endolysosomal escape. This study demonstrates that cyclosporin A combined with T-CUS 245C has a synergistic anticancer effect, by facilitating the release of T-CUS 245C in the intracellular space. Graphical Abstract
研究领域:
肿瘤学
·
细胞生物学
·
药理学
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DOI
高影响因子
Materials Express
·
2021
·Vol. 11
·期 10
·1619-1627
·
IF 0
Zhang Yue,Liu Yuanfen,Hu Yumin,Pan Lei
In this study, we developed an active targeting nano-immunoplatform of doxorubicin (DOX)-loaded bovine serum albumin (BSA) modified with the humanized antibody cetuximab, which can target the epidermal growth factor receptor (EGFR) of EGFR-overexpressing colon cancer, and subjected
it to a series of in vitro evaluation. The obtained cetuximab-modified nanoparticles (cetuximab-DOX-immuno-NPs) had an average particle size of 218.2±2.2 nm, a polydispersity index of 0.162±0.023, a zeta potential of −23.28±0.75 mV, and a cetuximabloading
efficiency of 33.6%±3.5%. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cytotoxicity experiments showed that the cytotoxic of cetuximab-DOX-immuno-NPs in EGFR-overexpressing RKO colon cancer cells was significantly higher than it was in EGFR-under-expressing
LS174 T cells. The results of flow cytometry showed that when DOX-immuno-NPs were used, DOX was eliminated soon after entering the cells, whereas when cetuximab-DOX-immuno-NPs were used, DOX efflux was reduced. Confocal laser-scanning microscopy was used to visualize the endocytosis of nanoparticles,
and revealed that the fluorescence intensity of cetuximab-DOX-immuno-NPs was significantly higher than that of non-specific IgG-DOX-immuno-NPs in RKO cells. Moreover, in EGFR-under-expressing LS174 T cells, both cetuximab-DOXimmuno-NPs and Immunoglobulin G-DOX-immuno-NPs showed weak DOX fluorescence
intensity, reflecting the specificity and selectivity of cetuximab-modified nanoparticles toward EGFR-overexpressing cancer cells. This study demonstrated that cetuximab modification of DOX-loaded BSA nanoparticles can increase their selectivity and reduce their toxicity. As this would decrease
the side effects of these treatments in patients, this study shows that cetuximab-modified doxorubicin-loaded bovine serum albumin nanoparticles are promising candidates for targeted colon cancer therapy.
研究领域:
药学
·
肿瘤学
·
生物医学工程
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DOI
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2024
·
IF 3.1
Yuan Dongxu,Liu Bingbing,Jiang Lin,Chen Yuhuan,Xu Gang,Lin Jianping,Yang Lirong,Lian Jiazhang,Jiang Yiqi,Ye Lidan,Wu Mianbin
Xylitol is a polyol widely used in food, pharmaceuticals, and light industries. It is currently produced through the chemical catalytic hydrogenation of xylose and generates xylose mother liquor as a substantial byproduct in the procedure of xylose extraction. If xylose mother liquor could also be efficiently bioconverted to xylitol, the greenness and atom economy of xylitol production would be largely improved. However, xylose mother liquor contains a mixture of glucose, xylose, and arabinose, raising the issue of carbon catabolic repression in its utilization by microbial conversion. Targeting this challenge, the transcriptional activator XylR was overexpressed in a previously constructed xylitol-producing E. coli strain CPH. The resulting strain CPHR produced 16.61 g/L of xylitol in shake-flask cultures from the mixture of corn cob hydrolysate and xylose mother liquor (1:1, v/v) with a xylose conversion rate of 90.1%, which were 2.23 and 2.15 times higher than the starting strain, respectively. Furthermore, XylR overexpression upregulated the expression levels of xylE , xylF , xylG , and xylH genes by 2.08–2.72 times in arabinose-containing medium, suggesting the alleviation of transcriptional repression of xylose transport genes by arabinose. This work lays the foundation for xylitol bioproduction from xylose mother liquor.
研究领域:
微生物学
·
生物技术
·
代谢工程
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DOI
PubMed
高影响因子
Frontiers in Microbiology
·
2021
·
IF 5.64
Chen Yongcan,Liang Jun,Chen Zhicong,Wang Bo,Si Tong
Heavy metal contamination is an environmental issue on a global scale. Particularly, cadmium poses substantial threats to crop and human health. Saccharomyces cerevisiae is one of the model organisms to study cadmium toxicity and was recently engineered as a cadmium hyperaccumulator. Therefore, it is desirable to overcome the cadmium sensitivity of S. cerevisiae via genetic engineering for bioremediation applications. Here we performed genome-scale overexpression screening for gene targets conferring cadmium resistance in CEN.PK2-1c, an industrial S. cerevisiae strain. Seven targets were identified, including CAD1 and CUP1 that are known to improve cadmium tolerance, as well as CRS5, NRG1, PPH21, BMH1, and QCR6 that are less studied. In the wild-type strain, cadmium exposure activated gene transcription of CAD1, CRS5, CUP1, and NRG1 and repressed PPH21, as revealed by real-time quantitative PCR analyses. Furthermore, yeast strains that contained two overexpression mutations out of the seven gene targets were constructed. Synergistic improvement in cadmium tolerance was observed with episomal co-expression of CRS5 and CUP1. In the presence of 200 μM cadmium, the most resistant strain overexpressing both CAD1 and NRG1 exhibited a 3.6-fold improvement in biomass accumulation relative to wild type. This work provided a new approach to discover and optimize genetic engineering targets for increasing cadmium resistance in yeast.
研究领域:
环境科学
·
遗传学
·
微生物学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF BIOLOGICAL CHEMISTRY
·
2025
·
IF 3.9
Yijun Yang,Nengyi Wang,Yuankun Chen,Tianpeng Ma,Minhua Weng,Haifeng Wu,Qiuping Wu,Yiqiang Xie,Chuanwu Zhu,Wenting Li
Nonalcoholic fatty liver disease (NAFLD) has posed a huge threat to public health globally, but there are currently no approved drugs available. Growing evidence has proved the close association between increased Th17 cells and NAFLD progression. Interleukin-4 induced protein 1 (IL4I1), an amino acid oxidase secreted by immune cells, was reported to regulate the Th17 cells, but its exact role in NAFLD progression has not been fully explained yet. We found that IL4I1 was highly expressed in the liver of C57BL/6J mice with NAFLD induced by an 8-weeks western diet. To explore the IL4I1’s effect, mice were injected with AAV8 encoding IL4I1 1 week before western diet administration. The results showed that IL4I1 overexpression inhibited the NAFLD progression, demonstrated by relieved liver damage and lipid accumulation. The underlying mechanism in which IL4I1 acts on NAFLD might be attributed to the inactivated AKT/forkhead box protein O1 (FOXO1) signaling pathway–mediated decrease of Th17 cells in liver tissues. Subsequently, by culturing naive CD4 + T cells isolated from the spleen of mice in Th17 cell-polarizing conditions, we determined that IL4I1 overexpression inhibited Th17 cell differentiation by inactivating the AKT/FOXO1 pathway, whereas its knockdown exhibited opposite effects. Further, the AKT activator SC79 reversed the effect of IL4I1 overexpression on Th17 cell differentiation. Collectively, our study supports that compensatory upregulation of IL4I1 protects against liver damage and lipid accumulation in NAFLD progression, partially by inhibiting the activated AKT/FOXO1 signaling pathway-induced Th17 cell differentiation.
研究领域:
免疫学
·
肝病学
·
分子生物学
·
细胞生物学
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产品关联
DOI
PubMed
高影响因子
Microbial Cell Factories
·
2024
·Vol. 23
·期 1
·1-16
·
IF 4.3
Wang Yifan,Shu Dan,Li Zhemin,Luo Di,Yang Jie,Chen Dongbo,Li Tianfu,Hou Xiaonan,Yang Qi,Tan Hong
Background Currently, industrial fermentation of Botrytis cinerea is a significant source of abscisic acid (ABA). The crucial role of ABA in plants and its wide range of applications in agricultural production have resulted in the constant discovery of new derivatives and analogues. While modifying the ABA synthesis pathway of existing strains to produce ABA derivatives is a viable option, it is hindered by the limited synthesis capacity of these strains, which hinders further development and application. Results In this study, we knocked out the bcaba4 gene of B. cinerea TB-31 to obtain the 1′,4′- trans -ABA-diol producing strain ZX2. We then studied the fermentation broth of the batch-fed fermentation of the ZX2 strain using metabolomic analysis. The results showed significant accumulation of 3-hydroxy-3-methylglutaric acid, mevalonic acid, and mevalonolactone during the fermentation process, indicating potential rate-limiting steps in the 1′,4′- trans -ABA-diol synthesis pathway. This may be hindering the flow of the synthetic pathway. Additionally, analysis of the transcript levels of terpene synthesis pathway genes in this strain revealed a correlation between the bchmgr , bcerg12 , and bcaba1-3 genes and 1′,4′- trans -ABA-diol synthesis. To further increase the yield of 1′,4′- trans -ABA-diol, we constructed a pCBg418 plasmid suitable for the Agrobacterium tumefaciens -mediated transformation (ATMT) system and transformed it to obtain a single-gene overexpression strain. We found that overexpression of bchmgr , bcerg12 , bcaba1 , bcaba2 , and bcaba3 genes increased the yield of 1′,4′- trans -ABA-diol. The highest yielding ZX2 A3 strain was eventually screened, which produced a 1′,4′- trans -ABA-diol concentration of 7.96 mg/g DCW (54.4 mg/L) in 144 h of shake flask fermentation. This represents a 2.1-fold increase compared to the ZX2 strain. Conclusions We utilized metabolic engineering techniques to alter the ABA-synthesizing strain B. cinerea , resulting in the creation of the mutant strain ZX2, which has the ability to produce 1′,4′- trans -ABA-diol. By overexpressing the crucial genes involved in the 1′,4′- trans -ABA-diol synthesis pathway in ZX2, we observed a substantial increase in the production of 1′,4′- trans -ABA-diol.
研究领域:
生物技术
·
微生物学
·
代谢工程
✓ 已验证
DOI
PubMed
高影响因子
PLANT PHYSIOLOGY AND BIOCHEMISTRY
·
2025
·
IF 5.7
Feixiong Zheng,Zhangting Xu,Xiaoji Deng,Yiming Sun,Jaime A. Teixeira da Silva,Xiaoxia Shen,Zhenming Yu
Salt stress severely constrains plant growth and productivity, so understanding the underlying mechanisms of salt stress is crucial for the genetic improvement of plants. Aquaporins (AQPs), which facilitate the transport of water or nutrients, exert an indispensable role in plants' response to salt stress. The AQP family in Dendrobium officinale , a commercially important medicinal and edible orchid, has not been identified or functionally characterized, while an understanding of this plant's tolerance to salinity remains unexplored. In this study, a genome-wide identification of AQP genes in D. officinale was performed using a salt-induced transcriptome. The 28 AQP members were mined and classified into four subfamilies (NIPs, PIPs, SIPs, and TIPs), and their physicochemical and bioinformatic properties were analyzed. Overexpression of DoTIP1-1 in D. officinale significantly enhanced salt tolerance via the accumulation of proline and water-soluble polysaccharides that improved water retention. Furthermore, salt tolerance was fortified by the improved activity of antioxidant enzymes (peroxidase and superoxide dismutase), allowing reactive oxygen species to be scavenged, and augmenting photosynthetic efficiency due to the upregulated expression of stress-responsive genes. This study provides the first comprehensive analysis of the AQP family in D. officinale and demonstrates that DoTIP1-1 is a key genetic determinant for enhancing salt tolerance, offering significant potential for the genetic improvement of this plant to resist abiotic stress.
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高影响因子
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