Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well ...Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well studied in bacteria. N6-adenine-specific DNA methyltransferase（N6AMT） is the main enzyme catalyzing the methylation of the adenine base and knowledge of this enzyme was mainly derived from work in prokaryotic models. However, large-scale gene discovery at the genome level in many model organisms indicated that the N6AMT gene also exists in eukaryotes, such as humans, mice, fruit flies and plants. Here, we cloned a N6AMT gene from Nilaparvata lugens（Nlu-N6AMT） and amplified its fulllength transcript. Then, we carried out a systematic investigation of N6AMT in 33 publically available insect genomes, indicating that all studied insects had N6AMT. Genomic structure analysis showed that insect N6AMT has short introns compared with the mammalian homologs. Domain and phylogenetic analysis indicated that insect N6AMT had a conserved N6-adenine Mlase domain that is specific to catalyze the adenine methylation. Nlu-N6AMT was highly expressed in the adult female. We knocked down Nlu-N6AMT by feeding ds RNA from the second instar nymph to adult female, inducing retard development of adult female. In all, we provide the first genome-wide analysis of N6AMT in insects and presented the experimental evidence that N6AMT might have important functions in reproductive development and ovary maturation.展开更多
CRISPR-Cas systems provide the small RNA-based adaptive immunity to defend against invasive genetic elements in archaea and bacteria.Organisms of Sulfolobales,an order of thermophilic acidophiles belonging to the Cren...CRISPR-Cas systems provide the small RNA-based adaptive immunity to defend against invasive genetic elements in archaea and bacteria.Organisms of Sulfolobales,an order of thermophilic acidophiles belonging to the Crenarchaeotal Phylum,usually contain both type I and typeⅢCRISPR-Cas systems.Two species,Saccharolobus solfataricus and Sulfolobus islandicus,have been important models for CRISPR study in archaea,and knowledge obtained from these studies has greatly expanded our understanding of molecular mechanisms of antiviral defense in all three steps:adaptation,expression and crRNA processing,and interference.Four subtypes of CRISPR-Cas systems are common in these organisms,including I-A,I-D,Ⅲ-B,andⅢ-D.These cas genes form functional modules,e.g.,all genes required for adaptation and for interference in the I-A immune system are clustered together to form aCas and i Cas modules.Genetic assays have been developed to study mechanisms of adaptation and interference by different CRISPR-Cas systems in these model archaea,and these methodologies are useful in demonstration of the protospacer-adjacent motif(PAM)-dependent DNA interference by I-A interference modules and multiple interference activities byⅢ-B Cmr systems.Ribonucleoprotein effector complexes have been isolated for SulfolobalesⅢ-B andⅢ-D systems,and their biochemical characterization has greatly enriched the knowledge of molecular mechanisms of these novel antiviral immune responses.展开更多
Prokaryotic Argonautes(pAgos)provide bacteria and archaea with immunity against plasmids and viruses.Catalytically active pAgos utilize short oligonucleotides as guides to directly cleave foreign nucleic acids,while i...Prokaryotic Argonautes(pAgos)provide bacteria and archaea with immunity against plasmids and viruses.Catalytically active pAgos utilize short oligonucleotides as guides to directly cleave foreign nucleic acids,while inactive pAgos lacking catalytic residues employ auxiliary effectors,such as nonspecific nucleases,to trigger abortive infection upon detection of foreign nucleic acids.Here,we report a unique group of catalytically active pAgo proteins that frequently associate with a phos-pholipase D(PLD)family protein.We demonstrate that this particular system employs the catalytic center of the associated PLD protein rather than that of pAgo to restrict plasmid DNA,while interestingly,its immunity against a single-stranded DNA virus relies on the pAgo catalytic center and is enhanced by the PLD protein.We also find that this system selectively suppresses viral DNA propagation without inducing noticeable abortive infection outcomes.Moreover,the pAgo protein alone enhances gene editing,which is unexpectedly inhibited by the PLD protein.Our data highlight the ability of catalytically active pAgo proteins to employ auxiliary proteins to strengthen the targeted eradication of different genetic invaders and underline the trend of PLD nucleases to participate in host immunity.展开更多
基金supported by the National Basic Research Program of China (2012CB114102)
文摘Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well studied in bacteria. N6-adenine-specific DNA methyltransferase（N6AMT） is the main enzyme catalyzing the methylation of the adenine base and knowledge of this enzyme was mainly derived from work in prokaryotic models. However, large-scale gene discovery at the genome level in many model organisms indicated that the N6AMT gene also exists in eukaryotes, such as humans, mice, fruit flies and plants. Here, we cloned a N6AMT gene from Nilaparvata lugens（Nlu-N6AMT） and amplified its fulllength transcript. Then, we carried out a systematic investigation of N6AMT in 33 publically available insect genomes, indicating that all studied insects had N6AMT. Genomic structure analysis showed that insect N6AMT has short introns compared with the mammalian homologs. Domain and phylogenetic analysis indicated that insect N6AMT had a conserved N6-adenine Mlase domain that is specific to catalyze the adenine methylation. Nlu-N6AMT was highly expressed in the adult female. We knocked down Nlu-N6AMT by feeding ds RNA from the second instar nymph to adult female, inducing retard development of adult female. In all, we provide the first genome-wide analysis of N6AMT in insects and presented the experimental evidence that N6AMT might have important functions in reproductive development and ovary maturation.
基金grants from the Chinese National Transgenic Science and Technology Program(2019ZX08010003 to QS)the National Natural Science Foundation of China(31771380 to QS)+1 种基金the Qingdao Applied Research Fund for postdoctoral researchers(62450079311107 to ZY)the State Key Laboratory of Microbial Technology and Shandong University。
文摘CRISPR-Cas systems provide the small RNA-based adaptive immunity to defend against invasive genetic elements in archaea and bacteria.Organisms of Sulfolobales,an order of thermophilic acidophiles belonging to the Crenarchaeotal Phylum,usually contain both type I and typeⅢCRISPR-Cas systems.Two species,Saccharolobus solfataricus and Sulfolobus islandicus,have been important models for CRISPR study in archaea,and knowledge obtained from these studies has greatly expanded our understanding of molecular mechanisms of antiviral defense in all three steps:adaptation,expression and crRNA processing,and interference.Four subtypes of CRISPR-Cas systems are common in these organisms,including I-A,I-D,Ⅲ-B,andⅢ-D.These cas genes form functional modules,e.g.,all genes required for adaptation and for interference in the I-A immune system are clustered together to form aCas and i Cas modules.Genetic assays have been developed to study mechanisms of adaptation and interference by different CRISPR-Cas systems in these model archaea,and these methodologies are useful in demonstration of the protospacer-adjacent motif(PAM)-dependent DNA interference by I-A interference modules and multiple interference activities byⅢ-B Cmr systems.Ribonucleoprotein effector complexes have been isolated for SulfolobalesⅢ-B andⅢ-D systems,and their biochemical characterization has greatly enriched the knowledge of molecular mechanisms of these novel antiviral immune responses.
基金This work was supported by the Science&Technology Fundamental Resources Investigation Program(2022FY101100)the Strategic Priority Research Program of the Chinese Academy of Sciences(Precision seed design and breeding)(XDA24020101)+4 种基金the National Natural Science Foundation of China(Nos.31970544,31970545,32022003,32150020,32200057,and 32270092)the Youth Innovation Promotion Association of CAS(No.2020090)the China National Postdoctoral Program for Innovative Talents(BX20220331)the China Postdoctoral Science Foundation(2022M720160)the Special Research Assistant Program of Chinese Academy of Sciences(No.2023000056).
文摘Prokaryotic Argonautes(pAgos)provide bacteria and archaea with immunity against plasmids and viruses.Catalytically active pAgos utilize short oligonucleotides as guides to directly cleave foreign nucleic acids,while inactive pAgos lacking catalytic residues employ auxiliary effectors,such as nonspecific nucleases,to trigger abortive infection upon detection of foreign nucleic acids.Here,we report a unique group of catalytically active pAgo proteins that frequently associate with a phos-pholipase D(PLD)family protein.We demonstrate that this particular system employs the catalytic center of the associated PLD protein rather than that of pAgo to restrict plasmid DNA,while interestingly,its immunity against a single-stranded DNA virus relies on the pAgo catalytic center and is enhanced by the PLD protein.We also find that this system selectively suppresses viral DNA propagation without inducing noticeable abortive infection outcomes.Moreover,the pAgo protein alone enhances gene editing,which is unexpectedly inhibited by the PLD protein.Our data highlight the ability of catalytically active pAgo proteins to employ auxiliary proteins to strengthen the targeted eradication of different genetic invaders and underline the trend of PLD nucleases to participate in host immunity.
