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The polyene macrolide rimocidin, produced by Streptomyces rimosus M527, is highly effective against a broad range of fungal plant pathogens, but at low yields. Elicitation is an effective method of stimulating the yield of bioactive secondary metabolites. In this study, the biomass and filtrate of a culture broth of Escherichia coli JM109, Bacillus subtilis WB600, Saccharomyces cerevisiae, and Fusarium oxysporum f. sp. cucumerinum were employed as elicitors to promote rimocidin production in S. rimosus M527. Adding culture broth and biomass of S. cerevisiae (A3) and F. oxysporum f. sp. cucumerinum (B4) resulted in an increase of rimocidin production by 51.2% and 68.3% respectively compared with the production under normal conditions in 5-l fermentor. In addition, quantitative RT-PCR analysis revealed that the transcriptions of ten genes (rimA to rimK) located in the gene cluster involved in rimocidin biosynthesis in A3 or B4 elicitation experimental group were all higher than those of a control group. Using a β-glucuronidase (GUS) reporter system, GUS enzyme activity assay, and Western blot analysis, we discovered that elicitation of A3 or B4 increased protein synthesis in S. rimosus M527. These results demonstrate that the addition of elicitors is a useful approach to improve rimocidin production.Key Points• An effective strategy for enhancing rimocidin production in S. rimosus M527 is demonstrated.• Overproduction of rimocidin is a result of higher expressed structural genes followed by an increase in protein synthesis.

The nucleoside antibiotic toyocamycin (TM), which was produced by Streptomyces diastatochromogenes 1628, was found to be highly efficient against a broad range of plant pathogenic fungi. Despite its importance, little is known about the regulation TM biosynthesis. In this study, toyA , located in the TM biosynthetic gene cluster, was identified as a regulatory gene encoding a large ATP-binding regulator of the LuxR family (LAL-family). The role of toyA in TM biosynthesis in S. diastatochromogenes 1628 was investigated by gene deletion, complementation, and over-expression. Gene disruption of toyA resulted in almost loss of TM production. TM production in complemented strain was restored to the level comparable to that in the wild-type strain S. diastatochromogenes 1628. Over-expression of toyA separately controlled by promoter SPL57, SPL21, and p erm E * in wild-type strain S. diastatochromogenes 1628 led to a 2-fold, 1-fold, and 80% increase in TM production compared with wild-type strain S. diastatochromogenes 1628, respectively. Quantitative RT-PCR analysis revealed that the transcriptional level of toy structural genes was downregulated in the ΔtoyA mutant but restored in complemented strain and further upregulated in the toyA over-expression strain. The detection results from GFP reporter system in Escherichia coli and GUS reporter system and GUS activities in S. albus J1074 and S. diastatochromogenes 1628 showed that ToyA activated the expression of toyB and toyE operon directly and activated the expression of other toy structural genes indirectly. These results indicate that ToyA is essential for TM biosynthesis controlling the expression of structural genes.

Abstract Rimocidin is a polyene macrolide that exhibits a strong inhibitory activity against a broad range of plant-pathogenic fungi. In this study, fermentation optimization and ribosome engineering technology were employed to enhance rimocidin production in Streptomyces rimosus M527. After the optimization of fermentation, rimocidin production in S. rimosus M527 increased from 0.11 ± 0.01 to 0.23 ± 0.02 g/L during shake-flask experiments and reached 0.41 ± 0.05 g/L using 5-L fermentor. Fermentation optimization was followed by the generation of mutants of S. rimosus M527 through treatment of the strain with different concentrations of gentamycin (Gen) or rifamycin. One Genr mutant named S. rimosus M527-G37 and one Rifr mutant named S. rimosus M527-R5 showed increased rimocidin production. Double-resistant (Genr and Rifr) mutants were selected using S. rimosus M527-G37 and S. rimosus M527-R5, and subsequently tested. One mutant, S. rimosus M527-GR7, which was derived from M527-G37, achieved the greatest cumulative improvement in rimocidin production. In the 5-L fermentor, the maximum rimocidin production achieved by S. rimosus M527-GR7 was 25.36% and 62.89% greater than those achieved by S. rimosus M527-G37 and the wild-type strain S. rimosus M527, respectively. Moreover, in the mutants S. rimosus M527-G37 and S. rimosus M527-GR7 the transcriptional levels of ten genes (rimA sr to rimK sr ) located in the gene cluster involved in rimocidin biosynthesis were all higher than those in the parental strain M527 to varying degrees. In addition, after expression of the single rimocidin biosynthetic genes in S. rimosus M527 a few recombinants showed an increase in rimocidin production. Expression of rimE led to the highest production.

The polyene macrolide rimocidin, produced by Streptomyces rimosus M527, was found to be highly effective against a broad range of fungal plant pathogens. Current understanding of the regulatory mechanism of rimocidin biosynthesis and morphological differentiation in S. rimosus M527 is limited. NsdA is considered a negative regulator involved in morphological differentiation and biosynthesis of secondary metabolites in some Streptomyces species. In this study, nsdAsr was cloned from S. rimosus M527. The role of nsdAsr in rimocidin biosynthesis and morphological differentiation was investigated by gene deletion, complementation, and over-expression. A ΔnsdAsr mutant was obtained using CRISPR/Cas9. The mutant produced more rimocidin (46%) and accelerated morphological differentiation than the wild-type strain. Over-expression of nsdAsr led to a decrease in rimocidin production and impairment of morphological differentiation. Quantitative RT-PCR analysis revealed that transcription of rim genes responsible for rimocidin biosynthesis was upregulated in the ΔnsdAsr mutant but downregulated in the nsdAsr over-expression strain. Similar effects have been described for Streptomyces coelicolor M145 and the industrial toyocamycin-producing strain Streptomyces diastatochromogenes 1628.Key points• A negative regulator for sporulation and rimocidin production was identified.• The CRISPR/Cas9 system was used for gene deletion in S. rimosus M527.

The extracellular matrix (ECM) is a significant factor in cancer progression. Collagens, as the main component of the ECM, are greatly remodeled alongside cancer development. More and more studies have confirmed that collagens changed from a barrier to providing assistance in cancer development. In this course, collagens cause remodeling alongside cancer progression, which in turn, promotes cancer development. The interaction between collagens and tumor cells is complex with biochemical and mechanical signals intervention through activating diverse signal pathways. As the mechanism gradually clears, it becomes a new target to find opportunities to diagnose and treat cancer. In this review, we investigated the process of collagen remodeling in cancer progression and discussed the interaction between collagens and cancer cells. Several typical effects associated with collagens were highlighted in the review, such as fibrillation in precancerous lesions, enhancing ECM stiffness, promoting angiogenesis, and guiding invasion. Then, the values of cancer diagnosis and prognosis were focused on. It is worth noting that several generated fragments in serum were reported to be able to be biomarkers for cancer diagnosis and prognosis, which is beneficial for clinic detection. At a glance, a variety of reported biomarkers were summarized. Many collagen-associated targets and drugs have been reported for cancer treatment in recent years. The new targets and related drugs were discussed in the review. The mass data were collected and classified by mechanism. Overall, the interaction of collagens and tumor cells is complicated, in which the mechanisms are not completely clear. A lot of collagen-associated biomarkers are excavated for cancer diagnosis. However, new therapeutic targets and related drugs are almost in clinical trials, with merely a few in clinical applications. So, more efforts are needed in collagens-associated studies and drug development for cancer research and treatment.

Drug-resistant bacterial strains seriously threaten human health. Rapid screening of antibiotics is urgently required to improve clinical treatment. Conventional methods of antimicrobial susceptibility testing rely on turbidimetry that is evident only after several days of incubation. The lengthy time of the assay can delay clinical treatment. Here, we proposed a single-cell level rapid system based on a microfluidic chip. The detection period of 30 min to 2 h was significantly shorter than the conventional turbidity-based method. To promote detection efficiency, 16 independent channels were designed, permitting the simultaneous screening of 16 drugs in the microfluidic chip. Prepositioning of drugs in the chip permitted prolonged transportation and storage. This may allow for the widespread use of the novel system, particularly in the regions where medical facilities are scarce. The growth curves were reported rapidly through a custom code in Matlab after tracking and photographing the bacteria during microscopy examination. The capability of the proposed system was validated by antimicrobial susceptibility testing trials with standard strains. The system provides a potentially useful detection tool for drug-resistant bacteria.

Gastric cancer (GC) has a poor prognosis and is a leading cause of cancer-related death. Optimal therapeutic targets have not been identified. AQP3 is capable of transporting glycerol across the cytomembrane. Previous studies have shown that AQP3 is involved in proliferation, invasion and migration by regulating glycerol and lipid metabolism in diverse cancer cell types. However, the potential roles of glycerol and lipid metabolism in AQP3-related cell apoptosis in GC remain unclear. In this study, we observed that AQP3 expression was upregulated in tumor tissues, and positively correlated with tumor size, lymph node metastasis and glycerol concentration in human GC samples. Silencing of AQP3 resulted in decreased glycerol intake and impaired lipid synthesis, which contributed to increased cell apoptosis. Furthermore, inhibition of autophagy induced by AQP3 knockdown promoted cell apoptosis. Administration of either glycerol or rapamycin restored cell viability, and overexpression of AQP3 increased cell viability by upregulating cellular glycerol metabolism and autophagy. Our study demonstrates that the increase in cell apoptosis of AQP3-deficient GC cells is a consequence of reduced glycerol uptake and lipogenesis and is associated with autophagy inhibition induced by AQP3 deficiency.

功能性步态障碍是功能性运动障碍的常见亚型,以失稳、跛行、帕金森样步态等为主征,与其他类型步态障碍性疾病的临床表现无异,存在误诊可能,但二者治疗原则存在显著差异,因此明确诊断、精准治疗对改善患者症状及预后至关重要.本文拟系统综述功能性步态障碍流行病学特点、临床特征、辅助检查、诊断与鉴别诊断、治疗与预后相关研究进展,以提高临床医师对疾病的认识.