RESUMEN
The CRISPR system is an adaptive immune system found in prokaryotes that defends host cells against the invasion of foreign DNA1. As part of the ongoing struggle between phages and the bacterial immune system, the CRISPR system has evolved into various types, each with distinct functionalities2. Type II Cas9 is the most extensively studied of these systems and has diverse subtypes. It remains uncertain whether members of this family can evolve additional mechanisms to counter viral invasions3,4. Here we identify 2,062 complete Cas9 loci, predict the structures of their associated proteins and reveal three structural growth trajectories for type II-C Cas9. We found that novel associated genes (NAGs) tended to be present within the loci of larger II-C Cas9s. Further investigation revealed that CbCas9 from Chryseobacterium species contains a novel ß-REC2 domain, and forms a heterotetrameric complex with an NAG-encoded CRISPR-Cas-system-promoting (pro-CRISPR) protein of II-C Cas9 (PcrIIC1). The CbCas9-PcrIIC1 complex exhibits enhanced DNA binding and cleavage activity, broader compatibility for protospacer adjacent motif sequences, increased tolerance for mismatches and improved anti-phage immunity, compared with stand-alone CbCas9. Overall, our work sheds light on the diversity and 'growth evolutionary' trajectories of II-C Cas9 proteins at the structural level, and identifies many NAGs-such as PcrIIC1, which serves as a pro-CRISPR factor to enhance CRISPR-mediated immunity.
Asunto(s)
Bacterias , Bacteriófagos , Proteína 9 Asociada a CRISPR , Sistemas CRISPR-Cas , Bacterias/virología , Bacterias/genética , Bacterias/inmunología , Bacteriófagos/genética , Bacteriófagos/inmunología , Chryseobacterium/genética , Chryseobacterium/inmunología , Chryseobacterium/virología , Proteína 9 Asociada a CRISPR/química , Proteína 9 Asociada a CRISPR/genética , Proteína 9 Asociada a CRISPR/metabolismo , Sistemas CRISPR-Cas/genética , Sistemas CRISPR-Cas/inmunología , División del ADN , Sitios Genéticos/genética , Modelos Moleculares , Dominios ProteicosRESUMEN
Microplastics (MPs) are abundant in aquaculture water, including in bioflocs aquaculture systems. Compared with other aquaculture systems, biofloc technology systems have the richest microbes and are beneficial to cultivated organisms. Therefore, this study provides a comprehensive assessment of the potential effects of MPs on aquaculture organisms in bioflocs systems. Here, Nile Tilapia (Oreochromis niloticus) were exposed to MPs (polystyrene; 32-40 µm diameter) with 0, 80 items/L (30 µg/L), and 800 items/L (300 µg/L) for 28 days in a bioflocs aquaculture system. The results showed that the MPs generally had no apparent effect on water quality, tilapia growth, or digestive enzyme activity. However, MPs accumulated the most in the liver (5.65 ± 0.74 µg/mg) and significantly increased the hepato-somatic index of tilapia and reduced the crude protein and lipid of tilapia muscle (p < 0.05). The levels of the antioxidant enzymes catalase and glutathione S-transferase increased significantly in response to MPs (p < 0.05). In contrast, MPs did not affect the content of glutathione, glutathione peroxidase, oxidized glutathione, and malondialdehyde, or the enzyme activity of Na+/K+-ATPase. Moreover, using an improved integrated biomarker response index, growth performance was found to be less responsive to MPs than to oxidative stress and digestive activity. Exposure to MPs did not significantly influence the microbial communities of the bioflocs and tilapia guts (p < 0.05). These results suggest that MPs barely affected tilapia in the bioflocs system. This study contributes to the evaluation of the ecological risk of MPs in aquaculture systems and a better understanding of the integrated response of cultivated vertebrates to MPs in biofloc technology systems.
