Comparative Proteomic Analysis of saccharopolyspora spinosa SP06081 and PR2 strains reveals the differentially expressed proteins correlated with the increase of spinosad yield

<p>Abstract</p> <p>Background</p> <p><it>Saccharopolyspora spinosa </it>produces the environment-friendly biopesticide spinosad, a mixture of two polyketide-derived macrolide active ingredients called spinosyns A and D. Therefore considerable interest is in the improvement of spinosad production because of its low yield in wild-type <it>S. spinosa</it>. Recently, a spinosad-hyperproducing PR2 strain with stable heredity was obtained from protoplast regeneration of the wild-type <it>S. spinosa </it>SP06081 strain. A comparative proteomic analysis was performed on the two strains during the first rapid growth phase (RG1) in seed medium (SM) by using label-free quantitative proteomics to investigate the underlying mechanism leading to the enhancement of spinosad yield.</p> <p>Results</p> <p>In total, 224 proteins from the SP06081 strain and 204 proteins from the PR2 strain were unambiguously identified by liquid chromatography-tandem mass spectrometry analysis, sharing 140 proteins. A total of 12 proteins directly related to spinosad biosynthesis were identified from the two strains in RG1. Comparative analysis of the shared proteins revealed that approximately 31% of them changed their abundance significantly and fell in all of the functional groups, such as tricarboxylic acid cycles, glycolysis, biosynthetic processes, catabolic processes, transcription, translation, oxidation and reduction. Several key enzymes involved in the synthesis of primary metabolic intermediates used as precursors for spinosad production, energy supply, polyketide chain assembly, deoxysugar methylation, and antioxidative stress were differentially expressed in the same pattern of facilitating spinosad production by the PR2 strain. Real-time reverse transcriptase polymerase chain reaction analysis revealed that four of five selected genes showed a positive correlation between changes at the translational and transcriptional expression level, which further confirmed the proteomic analysis.</p> <p>Conclusions</p> <p>The present study is the first comprehensive and comparative proteome analysis of <it>S. spinosa </it>strains. Our results highlight the differentially expressed proteins between the two <it>S. spinosa </it>strains and provide some clues to understand the molecular and metabolic mechanisms that could lead to the increased spinosad production yield.</p

Research Progress on the Polymeric Immunoglobulin Receptor (pIgR) in Fish

There are a large number of pathogens in the water where fish live, and the mucosal-associated lymphoid tissues (MALTs), such as skin, gill and intestine, are the first contact parts when pathogens infect fish. The secreted mucus of these tissues constitutes the first barrier for fish against the invasion of external pathogens. Mucosal immunity can identify and neutralize pathogens and induces immunocytes to devour pathogens and the like. As a key factor in the mucosal immune system, the polymeric immunoglobulin receptor (pIgR) is capable of mediating the transport and secretion of polymeric immunoglobulins towards mucus. The effective secretion of the pIgR is necessary for polymeric immunoglobulins (pIg) to exert mucosal defence and plays a significant role in fish immunity. With the deepening of research into fish immunoglobulins, the pIgR has become a research hotspot. The molecular structure, genetic structure and expression pattern of the pIgR and the important role it plays in mucosal immunity were summarized in this study, which contributed to a deeper understanding of fish mucosal immunity and laid a foundation for further exploration of the action mechanism and functions of the pIgR in fish

Induction of ROS Overload by Alantolactone Prompts Oxidative DNA Damage and Apoptosis in Colorectal Cancer Cells

Cancer cells typically display higher than normal levels of reactive oxygen species (ROS), which may promote cancer development and progression but may also render the cancer cells more vulnerable to further ROS insult. Indeed, many of the current anticancer therapeutics kill cancer cells via induction of oxidative stress, though they target both cancer and normal cells. Recently, alantolactone (ATL), a natural sesquiterpene lactone, has been shown to induce apoptosis by increasing ROS levels specifically in cancer cells; however, the molecular mechanisms linking ROS overproduction to apoptosis remain unclear. Here we show that the ATL-induced ROS overload in human SW480 and SW1116 colorectal cancer cells was followed by a prominent accumulation of cellular oxidized guanine (8-oxoG) and immediate increase in the number of DNA strand breaks, indicating that increased ROS resulted in extensive oxidative DNA damage. Consequently, the G1/S-CDK suppresser CDKN1B (p21) and pro-apoptotic proteins Bax and activated caspase-3 were upregulated, while anti-apoptotic Bcl-2 was downregulated, which were followed by cell cycle arrest at G1 and marked apoptosis in ATL-treated cancer but not non-cancer cells. These results suggest that the ATL-induced ROS overload triggers cell death through induction of massive oxidative DNA damage and subsequent activation of the intrinsic apoptosis pathway

Knockdown of SETDB1 inhibits breast cancer progression by miR-381-3p-related regulation

Abstract Background SET domain bifurcated 1 (SETDB1) has been widely considered as an oncogene playing a critical role in many human cancers, including breast cancer. Nevertheless, the molecular mechanism by which SETDB1 regulates breast cancer tumorigenesis is still unknown. Methods qRT-PCR assay or western blot analysis was performed to assess the expression level of SETDB1 mRNA or protein, respectively. siSETDB1, pCMV6-XL5-SETDB1, miR-381-3p mimic, or miR-381-3p inhibitor was transfected into cells to regulate the expression of SETDB1 or miR-381-3p. MiRNA directly interacted with SETDB1 was verified by luciferase reporter assay and RNA immunoprecipitation. CCK-8 assay, colony formation assay, flow cytometric analysis, and transwell assay were used to detect the abilities of cell proliferation, cell cycle progression and migration, respectively. Animal model of xenograft tumor was used to observe the regulatory effect of SETDB1 on tumor growth in vivo. Results We verified that SETDB1 mRNA level was upregulated in breast cancer tissues and cell lines, and SETDB1 depletion led to a suppression of cell proliferation, cell cycle progression and migration in vitro, as well as tumor growth in vivo. SETDB1 was verified to be a target of miR-381-3p. Moreover, miR-381-3p overexpression suppressed cell proliferation, cell cycle progression and migration, whereas SETDB1 abated miR-381-3p-mediated regulatory function on breast cancer cells. Conclusions This study revealed that SETDB1 knockdown might suppress breast cancer progression at least partly by miR-381-3p-related regulation, providing a novel prospect in breast cancer therapy

A Dual-Crosslinked Hydrogel Based on Gelatin Methacryloyl and Sulfhydrylated Chitosan for Promoting Wound Healing

The skin is the largest organ of the human body. Skin injuries, especially full-thickness injuries, are a major treatment challenge in clinical practice. Therefore, wound dressing materials with therapeutic effects have great practical significance in healthcare. This study used photocrosslinkable gelatin methacryloyl (GelMA) and sulfhydrylated chitosan (CS-SH) to design a double-crosslinked hydrogel for wound dressing. When crosslinked together, the resulting hydrogels showed a highly porous inner structure, and enhanced mechanical properties and moisture retention capacity. The compression modulus of the GelMA/CS-SH hydrogel (GCH) reached up to about 40 kPa and was much higher than that of pure GelMA hydrogel, and the compression modulus was increased with the amount of CS-SH. In vitro study showed no cytotoxicity of obtained hydrogels. Interestingly, a higher concentration of CS-SH slightly promoted the proliferation of cells. Moreover, the double-crosslinked hydrogel exhibited antibacterial properties because of the presence of chitosan. In vivo study based on rats showed that full-thickness skin defects healed on the 15th day. Histological results indicate that the hydrogel accelerated the repair of hair follicles and encouraged the orderly growth of collagen fibers in the wound. Furthermore, better blood vessel formation and a higher expression of VEGFR were observed in the hydrogel group when compared with the untreated control group. Based on our findings, GCH could be a promising candidate for full-thickness wound dressing

A Dual-Crosslinked Hydrogel Based on Gelatin Methacryloyl and Sulfhydrylated Chitosan for Promoting Wound Healing

The skin is the largest organ of the human body. Skin injuries, especially full-thickness injuries, are a major treatment challenge in clinical practice. Therefore, wound dressing materials with therapeutic effects have great practical significance in healthcare. This study used photocrosslinkable gelatin methacryloyl (GelMA) and sulfhydrylated chitosan (CS-SH) to design a double-crosslinked hydrogel for wound dressing. When crosslinked together, the resulting hydrogels showed a highly porous inner structure, and enhanced mechanical properties and moisture retention capacity. The compression modulus of the GelMA/CS-SH hydrogel (GCH) reached up to about 40 kPa and was much higher than that of pure GelMA hydrogel, and the compression modulus was increased with the amount of CS-SH. In vitro study showed no cytotoxicity of obtained hydrogels. Interestingly, a higher concentration of CS-SH slightly promoted the proliferation of cells. Moreover, the double-crosslinked hydrogel exhibited antibacterial properties because of the presence of chitosan. In vivo study based on rats showed that full-thickness skin defects healed on the 15th day. Histological results indicate that the hydrogel accelerated the repair of hair follicles and encouraged the orderly growth of collagen fibers in the wound. Furthermore, better blood vessel formation and a higher expression of VEGFR were observed in the hydrogel group when compared with the untreated control group. Based on our findings, GCH could be a promising candidate for full-thickness wound dressing