Yellow catfish (Pelteobagrus fulvidraco) is one of the most important freshwater aquaculture species in China. However, its small size and lower meat yield limit its edible value. Myostatin (MSTN) is a negative regulator of mammalian muscle growth. But, the function of Mstn in fish remains elusive. To explore roles of mstn gene in fish growth and create a strain of yellow catfish with high amount of muscle mass, we performed targeted disruption of mstn in yellow catfish using engineered zinc-finger nucleases (ZFNs). Employing zebrafish embryos as a screening system to identify ZFN activity, we obtained one pair of ZFNs that can edit mstn in yellow catfish genome. Using the ZFNs, we successfully obtained two founders (Founder July29-7 and Founder July29-8) carrying mutated mstn gene in their germ cells. The mutated mstn allele inherited from Founder July29-7 was a null allele (mstnnju6) containing a 4 bp insertion, predicted to encode function null Mstn. The mutated mstn inherited from Founder July29-8 was a complex type of mutation (mstnnju7), predicted to encode a protein lacking two amino acids in the N-terminal secretory signal of Mstn. Totally, we obtained 6 mstnnju6/+ and 14 mstnnju7/+ yellow catfish. To our best knowledge, this is the first endogenous gene knockout in aquaculture fish. Our result will help in understanding the roles of mstn gene in fish.
Engineered endonucleases are a powerful tool for editing DNA. However, sequence preferences may limit their application. We engineer a structure-guided endonuclease (SGN) composed of flap endonuclease-1 (FEN-1), which recognizes the 3′ flap structure, and the cleavage domain of Fok I (Fn1), which cleaves DNA strands. The SGN recognizes the target DNA on the basis of the 3′ flap structure formed between the target and the guide DNA (gDNA) and cut the target through its Fn1 dimerization. Our results show that the SGN, guided by a pair of gDNAs, cleaves transgenic reporter gene and endogenous genes in zebrafish embryonic genome.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-016-1038-5) contains supplementary material, which is available to authorized users.
Lung cancer is the main health threat in the world. Recently, oleuropein has been reported to have potent antioxidant and anticancer activities. However, the antitumor effects of oleuropein on H1299 cells are not well understood. Therefore, the purpose of this paper is tantamount to explore the effects of oleuropein on H1299 cells and its underlying mechanism that may be involved. Oleuropein treatment in H1299 cells resulted in cell cycle distribution at G2/M arrest and apoptosis in a dose‐dependent manner. Mitochondria‐mediated apoptosis was verified by the increase in Bax/Bcl‐2 ratio, release of cytochrome
c, and activation of caspase‐3 on oleuropein‐induced H1299 cells. In addition, our data also demonstrated that the p38 mitogen‐activated protein kinase (MAPK) signaling pathway has a critical role in oleuropein‐induced apoptosis. Moreover, we used transcriptome analysis to identify differentially expressed genes (DEGs) in H1299 cells by oleuropein and SB203580 treatment. Many DEGs were annotated to metabolic pathways, cell cycle, pathways in cancer, MAPK signaling pathway by Kyoto Encyclopedia of Genes and Genomes and Gene ontology enrichment methods. Network and expression analysis found that DEGs, including
RPS6A5,
GADD45A, and
MKP, play a key role in the p38 MAPK signaling pathway. In H1299 cells, oleuropein resulted in the expression of numerous genes related to cell signaling, metabolism pathway and directly associated with apoptosis. These results illustrated that oleuropein‐induced apoptosis via mitochondrial apoptotic cascade was activated by the p38 MAPK signaling pathway in H1299 cells. Thus, oleuropein as a natural compound and therapeutic drug has potential application value in the treatment of lung cancer.
Myostatin (Mstn), a member of the transforming growth factor b superfamily, plays an inhibiting role in mammalian muscle growth. Mammals like human, cattle, mouse, sheep, and dog carrying null alleles of Mstn display a double-muscle phenotype. Mstn is conserved in fish; however, little is known whether the fish with mutated mstn display a similar phenotype to mammals because of the lack of mutant fish with mstn null alleles. Previously, we knocked out one of the duplicated copies of myostatin gene (mstna) in yellow catfish using zincfinger nucleases. In this study, we report the identification of the second myostatin gene (mstnb) and knockout of mstnb in yellow catfish. The gene comprises three exons. It is predicted to encode 373 amino acid residues. The predicted protein exhibits 59.3% identity with yellow catfish Mstna and 57.3% identity with human MSTN. Employing TALEN (transcription activator-like effector nucleases) technology, we obtained two founders (from four randomly selected founders) of yellow catfish carrying the mutated mstnb gene in their germ cells. Totally, six mutated alleles of mstnb were obtained from the founders. Among the six alleles, four are nonframeshift and two are frameshift mutation. The frameshift mutated alleles include mstnb nju22 , an 8 bp deletion, and mstnb nju24 , a complex type of mutation comprising a 7 bp deletion and a 12 bp insertion. They are predicted to encode function null Mstnb. Our results will help to understand the roles of mstn genes in fish growth.
Background & Aims: Proton pump inhibitors (PPIs) have been reported to be associated with cholangitis and might possibly be carcinogenic. However, few studies have been conducted to investigate the association of PPIs with cholangiocarcinoma (CCA). Thus, a hospital-based case-control study was carried out in China to explore the association between PPIs and CCA. Methods: In this study, 1468 CCA cases (826 intrahepatic cholangiocarcinoma (ICC) and 642 extrahepatic cholangiocarcinoma (ECC)) were included, which were observed at Beijing Friendship Hospital, from February 2002 to October 2018. We retrospectively extracted PPI use and other possible risk factors from clinical records, followed by an investigation of the relationship with CCA via calculation of odds ratios (ORs), adjusted odds ratios (AORs), and 95% confidence intervals (CIs) using logistic regression analysis. Results: PPIs were used by 135 (9.2%) CCA cases and 173 (5.9%) controls. We found that PPI use was associated with a 1.61-fold elevated CCA odds (P < .001)
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