Recent advances with the type II clustered regularly interspaced short palindromic repeats (CRISPR) system promise an improved approach to genome editing. However, the applicability and efficiency of this system in model organisms, such as zebrafish, are little studied. Here, we report that RNA-guided Cas9 nuclease efficiently facilitates genome editing in both mammalian cells and zebrafish embryos in a simple and robust manner. Over 35% of site-specific somatic mutations were found when specific Cas/gRNA was used to target either etsrp, gata4 or gata5 in zebrafish embryos in vivo. The Cas9/gRNA efficiently induced biallelic conversion of etsrp or gata5 in the resulting somatic cells, recapitulating their respective vessel phenotypes in etsrpy11 mutant embryos or cardia bifida phenotypes in fautm236a mutant embryos. Finally, we successfully achieved site-specific insertion of mloxP sequence induced by Cas9/gRNA system in zebrafish embryos. These results demonstrate that the Cas9/gRNA system has the potential of becoming a simple, robust and efficient reverse genetic tool for zebrafish and other model organisms. Together with other genome-engineering technologies, the Cas9 system is promising for applications in biology, agriculture, environmental studies and medicine.

Microbes appear in every corner of human life, and microbes affect every aspect of human life. The human oral cavity contains a number of different habitats. Synergy and interaction of variable oral microorganisms help human body against invasion of undesirable stimulation outside. However, imbalance of microbial flora contributes to oral diseases and systemic diseases. Oral microbiomes play an important role in the human microbial community and human health. The use of recently developed molecular methods has greatly expanded our knowledge of the composition and function of the oral microbiome in health and disease. Studies in oral microbiomes and their interactions with microbiomes in variable body sites and variable health condition are critical in our cognition of our body and how to make effect on human health improvement.

Aberrant activation of c-kit proto-oncogene contributes to abnormal cell proliferation by altering the tyrosine kinase signaling and constitutes a crucial impetus for leukemogenesis. Epigenetic silencing of tumor-suppressive microRNAs (miRNAs) is a key oncogenic mechanism for the activation of oncogenes in tumors. In this study, several miRNAs potentially binding to the 3 0 -untranslated region of human c-kit mRNA were screened by luciferase reporter assays. Among these miRNAs, miR-193a was embedded in a CpG island and epigenetically repressed by promoter hypermethylation in acute myeloid leukemia (AML) cell lines and primary AML blasts, but not in normal bone marrow cells. Importantly, miR-193a levels were inversely correlated with c-kit levels measured in 9 leukemia cell lines and 27 primary AML samples. Restoring miR-193a expression in AML cells harboring c-kit mutation and/or overexpression, either by synthetic miR-193a transfection or by DNA hypomethylating agent 5-azacytidine (5-aza) treatment, resulted in a significant reduction in c-kit expression at both RNA and protein levels and inhibition of cell growth. The growth-inhibitory activity of miR-193a was associated with apoptosis and granulocytic differentiation. Moreover, 5-aza-induced c-kit reduction could be partially blocked by miR-193a inhibitor, leading to a reversal of antiproliferative and proapoptotic effects of 5-aza. These data reveal a critical role for methylation-repressed miR-193a in myeloid leukemogenesis and the therapeutic promise of upregulating miR-193a expression for c-kit-positive AML.

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A comprehensive review that offers mechanistic insight into the CRISPR–Cas toolbox for analytical and diagnostic assay development.

A limit to the clinical benefit of radiotherapy is not an incapacity to eliminate tumor cells but rather a limit on its capacity to do so without destroying normal tissue and inducing inflammation. Recent evidence reveals that the inflammasome is essential for mediating radiation-induced cell and tissue damage. In this study, using primary cultured bone marrow-derived macrophages (BMDM) and a mouse radiation model, we explored the role of NLRP3 inflammasome activation and the secondary pyroptosis underlying radiation-induced immune cell death. We observed an increasing proportion of pyroptosis and elevating Caspase-1 activation in 10 and 20 Gy radiation groups. Nlrp3 knock out significantly diminished the quantity of cleaved-Caspase-1 (p10) and IL-1β as well as the proportion of pyroptosis. Additionally, in vivo research shows that 9.5 Gy of radiation promotes Caspase-1 activation in marginal zone cells and induces death in mice, both of which can be significantly inhibited by knocking out Nlrp3. Thus, based on these findings, we conclude that the NLRP3 inflammasome activation mediates radiation-induced pyroptosis in BMDMs. Targeting NLRP3 inflammasome and pyroptosis may serve as effective strategies to diminish injury caused by radiation.

Geosmin provides the characteristic but sometimes undesirable "earthy" flavor to red table beets. To date, it is not known whether geosmin is a byproduct of beet metabolism or synthesized by soil-borne microorganisms and taken up by the beets during maturation. Analysis of mature beet roots revealed that peels contained 6 times the amount of geosmin compared to the bodies and cores. Sterilized beet seeds were aseptically grown in a basal medium prior to analysis for the presence of geosmin. Using a headspace solid-phase microextraction (HSPME) method, the relative recovery of geosmin from beet seedling extracts was 72.0 +/- 4.2% with (-)-menthone as the internal standard. The presence of geosmin in aseptically grown beet seedlings was confirmed by gas chromatography-mass spectrometry using authentic geosmin as the standard. During aseptic growth, the concentration of geosmin in seedlings remained constant for up to 5 months but increased at 6 months. Geosmin added to the growth medium was not absorbed by the seedlings. These studies support the conclusion that red beets are capable of endogenous synthesis of geosmin.

The zebrafish possesses a remarkable capacity of adult heart regeneration, but the underlying mechanisms are not well understood. Here we report that chromatin remodelling factor Brg1 is essential for adult heart regeneration. Brg1 mRNA and protein are induced during heart regeneration. Transgenic over-expression of dominant-negative Xenopus Brg1 inhibits the formation of BrdU+/Mef2C+ and Tg(gata4:EGFP) cardiomyocytes, leading to severe cardiac fibrosis and compromised myocardial regeneration. RNA-seq and RNAscope analyses reveal that inhibition of Brg1 increases the expression of cyclin-dependent kinase inhibitors such as cdkn1a and cdkn1c in the myocardium after ventricular resection; and accordingly, myocardial-specific expression of dn-xBrg1 blunts myocardial proliferation and regeneration. Mechanistically, injury-induced Brg1, via its interaction with Dnmt3ab, suppresses the expression of cdkn1c by increasing the methylation level of CpG sites at the cdkn1c promoter. Taken together, our results suggest that Brg1 promotes heart regeneration by repressing cyclin-dependent kinase inhibitors partly through Dnmt3ab-dependent DNA methylation.