Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endo...Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endonuclease adapt to the changes? The correlations between evolving pre-RNAs and adapting homodimeric endonuclease in lower prokaryotes is investigated in this paper. First, we construct and observe the appearance of a long branch in the phylogeny based on homodimeric endonuclease. To appreciate the finer aspects of accelerating evolution near this long branch, we delve deeper into the pre-RNA substrates of the endonuclease. Computational evidence of an as-yet-unreported noncoding RNA gene then emerges from this study. The capabilities of homodimeric endonuclease and the complexities of its pre-RNA substrates appear to evolve in steps together.展开更多
Circular intronic RNAs(ci RNAs) escaping from DBR1 debranching of intron lariats are co-transcriptionally produced from prem RNA splicing, but their turnover and mechanism of action have remained elusive. We report th...Circular intronic RNAs(ci RNAs) escaping from DBR1 debranching of intron lariats are co-transcriptionally produced from prem RNA splicing, but their turnover and mechanism of action have remained elusive. We report that RNase H1 degrades a subgroup of ci RNAs in human cells. Many ci RNAs contain high GC% and tend to form DNA:RNA hybrids(R-loops) for RNase H1 cleavage, a process that appears to promote Pol II transcriptional elongation at ci RNA-producing loci. One ci RNA, ciankrd52, shows a stronger ability of R-loop formation than that of its cognate pre-m RNA by maintaining a locally open RNA structure in vitro. This allows the release of pre-m RNA from R-loops by ci-ankrd52 replacement and subsequent ci RNA removal via RNase H1 for efficient transcriptional elongation. We propose that such an R-loop dependent ci RNA degradation likely represents a mechanism that on one hand limits ci RNA accumulation by recruiting RNase H1 and on the other hand resolves Rloops for transcriptional elongation at some GC-rich ci RNA-producing loci.展开更多
Ⅰ类内含子(group I intron)是研究RNAs结构与功能关系的理想元件,在解释RNA折叠理论、催化机制等方面起着重要作用;对其结构与功能关系的研究也因此成为一个非常重要的课题.本研究建立了一个基于卡那霉素抗性进行Ⅰ类内含子结构与功能...Ⅰ类内含子(group I intron)是研究RNAs结构与功能关系的理想元件,在解释RNA折叠理论、催化机制等方面起着重要作用;对其结构与功能关系的研究也因此成为一个非常重要的课题.本研究建立了一个基于卡那霉素抗性进行Ⅰ类内含子结构与功能关系研究系统,将源于海洋蓝细菌Nostoc punctiforme(Npu)核糖核酸还原酶基因(ribonucleotide reductase,Rir)中的1个Ⅰ类内含子插入到pDrive质粒的卡那霉素抗性基因(kanamycin resistance gene,KanR)内构建得pKR12质粒并转化大肠杆菌(E.coli).只有内含子剪接的阳性克隆才能生成KanR蛋白并在Kan抗性平板上生长.结果显示,pKR12转入E.coli后不能在Kan抗性平板上生长,RT-PCR检测仅可见前体带,表明插入到KanR中的Npu Rir内含子没有发生剪接.随后通过易错PCR建立内含子的随机突变库并用Kan抗性筛选进行定向演化,产生有剪接活性的内含子突变体,RT-PCR检测显示剪接发生.由于内含子剪接活性的改变可通过Kan抗性变化在LB平板上得以反映,因此该系统有望成为简单快速地研究Ⅰ类内含子结构与功能关系的有利工具.展开更多
Background:There is increasing evidence that circular RNAs(circRNAs)play a significant role in pathological processes including tumorigenesis.In contrast to exonic circRNAs,which are the most frequently reported circR...Background:There is increasing evidence that circular RNAs(circRNAs)play a significant role in pathological processes including tumorigenesis.In contrast to exonic circRNAs,which are the most frequently reported circRNAs in cancer so far,the studies of intronic circRNAs have been greatly lagged behind.Here,we aimed to investigate the regulatory role of intronic circRNAs in head and neck squamous cell carcinoma(HNSCC).Methods:We conducted whole-transcriptome sequencing with four pairs of primary tumor tissues and adjacent normal tissues from HNSCC patients.Then,we characterized circGNG7 expression in HNSCC tissues and cell lines and explored its association with the prognosis of HNSCC patients.We also identified interactions between circGNG7 and functional proteins,which alter downstream signaling that regulate HNSCC progression.Results:In this study,we identified a new intronic circRNA,circGNG7,and validated its functional roles in HNSCC progression.CircGNG7 was predominately localized to the cytoplasm,and its expression was downregulated in both HNSCC tissues andCAL27,CAL33,SCC4,SCC9,HN6,and HN30 cells.Low expression of circGNG7 was significantly correlated with poor prognosis in HNSCC patients.Consistent with this finding,overexpression of circGNG7 strongly inhibited tumor cell proliferation,colony formation,in vitro migration,and in vivo tumor growth.Mechanistically,the expression of circGNG7 in HNSCC cells was regulated by the transcription factor SMAD family member 4(SMAD4).Importantly,we discovered that circGNG7 could bind to serine residues 78 and 82 of the functional heat shock protein 27(HSP27),occupying its phosphorylation sites and hindering its phosphorylation,which reduced HSP27-JNK/P38 mitogen-activated protein kinase(MAPK)oncogenic signaling.Downregulation of circGNG7 expression in HNSCC increased HSP27-JNK/P38 MAPK signaling and promoted tumor progression.Conclusions:Our results revealed that a new intronic circRNA,circGNG7,functions as a strong tumor suppressor and that circGNG7/HSP27-JNK/P38 MAPK signaling is a novel mechanism by which HNSCC progression can be controlled.展开更多
Objective: To classify four new Sparassis strains(CLM1, CKM1, CKM2, and KJM1) using the internal transcribed spacer sequence and to elucidate their β-glucan content and mycelial growth.Methods: Two different microbio...Objective: To classify four new Sparassis strains(CLM1, CKM1, CKM2, and KJM1) using the internal transcribed spacer sequence and to elucidate their β-glucan content and mycelial growth.Methods: Two different microbiological media were used to determine growth rate. The β-glucan contents were analyzed using the Megazyme Mushroom and Yeast Beta-Glucan kit. To determine the genetic relationships, phylogenetic trees were constructed using ClustalX. Multiple sequence alignments were printed and shaded with the BOXSHADE 3.21 program. Results: In this study, four new Sparassis strains were isolated from the southern region of the Korea Peninsula. They were all classified into the Sparassis latifolia clade as a monophyletic group based on the internal transcribed spacer sequence. Mycelial growth rate of the CLM1 strain was highest in potato dextrose agar and potato dextrose agar larch. The β-glucan content of the CLM1 strain was highest at 29.5%(w/w). A high degree of sequence divergence was detected in the RNA polymerase second largest subunit II gene(RPB2) within Sparassis spp. tested. The putative amino acid sequences of the RPB2 had a distinct sequence. The nucleotide sequences of the RPB2's intron were also divergent among Sparassis spp., even though their nucleotide length was well conserved within Sparassis latifolia. Conclusions: These results indicate that the nucleotide sequences and the amino acid sequences of RPB2 can be used to identify individual Sparassis sp. The Sparassis strain CLM1 may be best for developing a remedy to prevent or treat cancer and other chronic diseases.展开更多
Alternative splicing （AS） is a vital genetic mechanism that enhances the diversity of eukaryotic transcriptomes. Here, we generated 8.3 Gb high-quality RNA-sequencing data from cotton （Gossypium raimondii） and per...Alternative splicing （AS） is a vital genetic mechanism that enhances the diversity of eukaryotic transcriptomes. Here, we generated 8.3 Gb high-quality RNA-sequencing data from cotton （Gossypium raimondii） and performed a systematic, comparative analysis of AS events. We mapped 85% of the RNA-sequencing data onto the reference genome and identified 154 368 splice junctions with 16 437 as events in 10197 genes. I ntron retention constituted the majority （40%） of all AS events in G. raimondii. Comparison across 11 eukaryote species showed that intron retention is the most common AS type in higher plants. Although transposable elements （TEs） were found in only 2.9% of all G. raimondii introns, they are present in 43% of the retained introns, suggesting that TE-insertion may be an important mechanism for intron retention during AS. The majority of the TE insertions are concentrated 0-40 nt upstream of the 3＇-splice site, substantially altering the distribution of branch points from preferred positions and reducing the efficiency of intron splicing by decreasing RNA secondary structure flexibility. Our data suggest that TE-insertion-induced changes in branch point-site distribution are important for intron retention-type AS. Our findings may help explain the vast differences in intron-retention frequencies between vertebrates and higher plants.展开更多
文摘Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endonuclease adapt to the changes? The correlations between evolving pre-RNAs and adapting homodimeric endonuclease in lower prokaryotes is investigated in this paper. First, we construct and observe the appearance of a long branch in the phylogeny based on homodimeric endonuclease. To appreciate the finer aspects of accelerating evolution near this long branch, we delve deeper into the pre-RNA substrates of the endonuclease. Computational evidence of an as-yet-unreported noncoding RNA gene then emerges from this study. The capabilities of homodimeric endonuclease and the complexities of its pre-RNA substrates appear to evolve in steps together.
基金This work was supported by the National Natural Science Foundation of China(NSFC)(91940303,31725009)the HHMI International Program(55008728)to L.-L.C.+2 种基金NSFC(31730111,31925011)to L.Y.Young Elite Scientists Sponsorship Program(2020QNRC001)to X.L.L.-L.C.the support from the XPLORER PRIZE.
文摘Circular intronic RNAs(ci RNAs) escaping from DBR1 debranching of intron lariats are co-transcriptionally produced from prem RNA splicing, but their turnover and mechanism of action have remained elusive. We report that RNase H1 degrades a subgroup of ci RNAs in human cells. Many ci RNAs contain high GC% and tend to form DNA:RNA hybrids(R-loops) for RNase H1 cleavage, a process that appears to promote Pol II transcriptional elongation at ci RNA-producing loci. One ci RNA, ciankrd52, shows a stronger ability of R-loop formation than that of its cognate pre-m RNA by maintaining a locally open RNA structure in vitro. This allows the release of pre-m RNA from R-loops by ci-ankrd52 replacement and subsequent ci RNA removal via RNase H1 for efficient transcriptional elongation. We propose that such an R-loop dependent ci RNA degradation likely represents a mechanism that on one hand limits ci RNA accumulation by recruiting RNase H1 and on the other hand resolves Rloops for transcriptional elongation at some GC-rich ci RNA-producing loci.
基金This study was supported by grants from the National Natural Science Foundation of China(No.81902748 and 81872185)Shanghai Sailing Program(No.19YF1427100)sponsored by Program of Innovative Research Team of High-level Local Universities in Shanghai.
文摘Background:There is increasing evidence that circular RNAs(circRNAs)play a significant role in pathological processes including tumorigenesis.In contrast to exonic circRNAs,which are the most frequently reported circRNAs in cancer so far,the studies of intronic circRNAs have been greatly lagged behind.Here,we aimed to investigate the regulatory role of intronic circRNAs in head and neck squamous cell carcinoma(HNSCC).Methods:We conducted whole-transcriptome sequencing with four pairs of primary tumor tissues and adjacent normal tissues from HNSCC patients.Then,we characterized circGNG7 expression in HNSCC tissues and cell lines and explored its association with the prognosis of HNSCC patients.We also identified interactions between circGNG7 and functional proteins,which alter downstream signaling that regulate HNSCC progression.Results:In this study,we identified a new intronic circRNA,circGNG7,and validated its functional roles in HNSCC progression.CircGNG7 was predominately localized to the cytoplasm,and its expression was downregulated in both HNSCC tissues andCAL27,CAL33,SCC4,SCC9,HN6,and HN30 cells.Low expression of circGNG7 was significantly correlated with poor prognosis in HNSCC patients.Consistent with this finding,overexpression of circGNG7 strongly inhibited tumor cell proliferation,colony formation,in vitro migration,and in vivo tumor growth.Mechanistically,the expression of circGNG7 in HNSCC cells was regulated by the transcription factor SMAD family member 4(SMAD4).Importantly,we discovered that circGNG7 could bind to serine residues 78 and 82 of the functional heat shock protein 27(HSP27),occupying its phosphorylation sites and hindering its phosphorylation,which reduced HSP27-JNK/P38 mitogen-activated protein kinase(MAPK)oncogenic signaling.Downregulation of circGNG7 expression in HNSCC increased HSP27-JNK/P38 MAPK signaling and promoted tumor progression.Conclusions:Our results revealed that a new intronic circRNA,circGNG7,functions as a strong tumor suppressor and that circGNG7/HSP27-JNK/P38 MAPK signaling is a novel mechanism by which HNSCC progression can be controlled.
基金Supported by the Regional Technology Innovation Program(A001100501)funded by the Ministry of Knowledge Economy,Korea+1 种基金the Basic Science Research Program through the National Research Foundation of Koreafunded by the Ministry of Education,Science and Technology(NRF-2012R1A1A2006934)
文摘Objective: To classify four new Sparassis strains(CLM1, CKM1, CKM2, and KJM1) using the internal transcribed spacer sequence and to elucidate their β-glucan content and mycelial growth.Methods: Two different microbiological media were used to determine growth rate. The β-glucan contents were analyzed using the Megazyme Mushroom and Yeast Beta-Glucan kit. To determine the genetic relationships, phylogenetic trees were constructed using ClustalX. Multiple sequence alignments were printed and shaded with the BOXSHADE 3.21 program. Results: In this study, four new Sparassis strains were isolated from the southern region of the Korea Peninsula. They were all classified into the Sparassis latifolia clade as a monophyletic group based on the internal transcribed spacer sequence. Mycelial growth rate of the CLM1 strain was highest in potato dextrose agar and potato dextrose agar larch. The β-glucan content of the CLM1 strain was highest at 29.5%(w/w). A high degree of sequence divergence was detected in the RNA polymerase second largest subunit II gene(RPB2) within Sparassis spp. tested. The putative amino acid sequences of the RPB2 had a distinct sequence. The nucleotide sequences of the RPB2's intron were also divergent among Sparassis spp., even though their nucleotide length was well conserved within Sparassis latifolia. Conclusions: These results indicate that the nucleotide sequences and the amino acid sequences of RPB2 can be used to identify individual Sparassis sp. The Sparassis strain CLM1 may be best for developing a remedy to prevent or treat cancer and other chronic diseases.
文摘Alternative splicing （AS） is a vital genetic mechanism that enhances the diversity of eukaryotic transcriptomes. Here, we generated 8.3 Gb high-quality RNA-sequencing data from cotton （Gossypium raimondii） and performed a systematic, comparative analysis of AS events. We mapped 85% of the RNA-sequencing data onto the reference genome and identified 154 368 splice junctions with 16 437 as events in 10197 genes. I ntron retention constituted the majority （40%） of all AS events in G. raimondii. Comparison across 11 eukaryote species showed that intron retention is the most common AS type in higher plants. Although transposable elements （TEs） were found in only 2.9% of all G. raimondii introns, they are present in 43% of the retained introns, suggesting that TE-insertion may be an important mechanism for intron retention during AS. The majority of the TE insertions are concentrated 0-40 nt upstream of the 3＇-splice site, substantially altering the distribution of branch points from preferred positions and reducing the efficiency of intron splicing by decreasing RNA secondary structure flexibility. Our data suggest that TE-insertion-induced changes in branch point-site distribution are important for intron retention-type AS. Our findings may help explain the vast differences in intron-retention frequencies between vertebrates and higher plants.
