Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method
Selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) is a widely used technique for RNA structure analysis using N-methylisatoic anhydride (NMIA) treatment that has been proven to be applicable to different types of RNA templates. In this study, we performed the structural analysis o...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2023-07-01
|
| Series: | Agronomy |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2073-4395/13/8/1990 |
| _version_ | 1797585835465375744 |
|---|---|
| author | Zhifei Liu Xinran Cao Chengming Yu Xuefeng Yuan |
| author_facet | Zhifei Liu Xinran Cao Chengming Yu Xuefeng Yuan |
| author_sort | Zhifei Liu |
| collection | DOAJ |
| description | Selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) is a widely used technique for RNA structure analysis using N-methylisatoic anhydride (NMIA) treatment that has been proven to be applicable to different types of RNA templates. In this study, we performed the structural analysis of the viral satellite RNA of cucumber mosaic virus TA-Tb (satCMV TA-Tb) using the SHAPE method. In the preliminary experiment, we optimized the protocol of SHAPE method for analyzing satCMV TA-Tb by determining a suitable quantity of template RNA. This optimization effectively reduced the appearance of a large number of intense bands in the NMIA-untreated sample lane, along with a strong overall background signal that prevented the clear elucidation of the RNA structure. SHAPE analysis indicated the presence of non-structured, single-stranded flexible regions throughout satCMV TA-Tb with prominent flexible stretches located around nucleotide positions 145 to 200. The positions of these flexible regions were particularly consistent with a secondary structure of satCMV TA-Tb predicted by mfold software v.2.3, which consisted of five 5′- and 3′-proximal stem-loops and one internal large multi-branched stem-loop. Sequence alignment and secondary RNA structure prediction of other satCMV sequences that are phylogenetically the same group with satCMV TA-Tb also suggested the presence of 5′- and 3′-proximal stem-loop structures. Our data provide the structural basis for elucidating the mechanism by which satCMV TA-Tb regulates the pathogenicity and replication of its helper virus. |
| first_indexed | 2024-03-11T00:12:43Z |
| format | Article |
| id | doaj.art-9de66487255e48ac97a487167a7b7b01 |
| institution | Directory Open Access Journal |
| issn | 2073-4395 |
| language | English |
| last_indexed | 2024-03-11T00:12:43Z |
| publishDate | 2023-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Agronomy |
| spelling | doaj.art-9de66487255e48ac97a487167a7b7b012023-11-18T23:53:31ZengMDPI AGAgronomy2073-43952023-07-01138199010.3390/agronomy13081990Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE MethodZhifei Liu0Xinran Cao1Chengming Yu2Xuefeng Yuan3College of Plant Protection, Shandong Agricultural University, Tai’an 271018, ChinaCollege of Plant Health and Medicine, Qingdao Agricultural University, Qingdao 266109, ChinaCollege of Plant Protection, Shandong Agricultural University, Tai’an 271018, ChinaCollege of Plant Protection, Shandong Agricultural University, Tai’an 271018, ChinaSelective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) is a widely used technique for RNA structure analysis using N-methylisatoic anhydride (NMIA) treatment that has been proven to be applicable to different types of RNA templates. In this study, we performed the structural analysis of the viral satellite RNA of cucumber mosaic virus TA-Tb (satCMV TA-Tb) using the SHAPE method. In the preliminary experiment, we optimized the protocol of SHAPE method for analyzing satCMV TA-Tb by determining a suitable quantity of template RNA. This optimization effectively reduced the appearance of a large number of intense bands in the NMIA-untreated sample lane, along with a strong overall background signal that prevented the clear elucidation of the RNA structure. SHAPE analysis indicated the presence of non-structured, single-stranded flexible regions throughout satCMV TA-Tb with prominent flexible stretches located around nucleotide positions 145 to 200. The positions of these flexible regions were particularly consistent with a secondary structure of satCMV TA-Tb predicted by mfold software v.2.3, which consisted of five 5′- and 3′-proximal stem-loops and one internal large multi-branched stem-loop. Sequence alignment and secondary RNA structure prediction of other satCMV sequences that are phylogenetically the same group with satCMV TA-Tb also suggested the presence of 5′- and 3′-proximal stem-loop structures. Our data provide the structural basis for elucidating the mechanism by which satCMV TA-Tb regulates the pathogenicity and replication of its helper virus.https://www.mdpi.com/2073-4395/13/8/1990SHAPERNA structure probingsatellite RNAcucumber mosaic virus |
| spellingShingle | Zhifei Liu Xinran Cao Chengming Yu Xuefeng Yuan Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method Agronomy SHAPE RNA structure probing satellite RNA cucumber mosaic virus |
| title | Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method |
| title_full | Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method |
| title_fullStr | Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method |
| title_full_unstemmed | Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method |
| title_short | Probing the RNA Structure of a Satellite RNA of Cucumber Mosaic Virus Using SHAPE Method |
| title_sort | probing the rna structure of a satellite rna of cucumber mosaic virus using shape method |
| topic | SHAPE RNA structure probing satellite RNA cucumber mosaic virus |
| url | https://www.mdpi.com/2073-4395/13/8/1990 |
| work_keys_str_mv | AT zhifeiliu probingthernastructureofasatelliternaofcucumbermosaicvirususingshapemethod AT xinrancao probingthernastructureofasatelliternaofcucumbermosaicvirususingshapemethod AT chengmingyu probingthernastructureofasatelliternaofcucumbermosaicvirususingshapemethod AT xuefengyuan probingthernastructureofasatelliternaofcucumbermosaicvirususingshapemethod |