Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains
NS3 protein is a member of the non-structural protein of duck Tembusu virus (DTMUV), which contains three domains, each of which has serine protease, nucleotide triphosphatase, and RNA helicase activities, respectively. It performs a variety of biological functions that are involved in the regulatio...
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| Format: | Article |
| Language: | English |
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MDPI AG
2019-08-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/11/8/740 |
| _version_ | 1817982663619772416 |
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| author | Yawen Wang Shuai Zhang Yi Tang Youxiang Diao |
| author_facet | Yawen Wang Shuai Zhang Yi Tang Youxiang Diao |
| author_sort | Yawen Wang |
| collection | DOAJ |
| description | NS3 protein is a member of the non-structural protein of duck Tembusu virus (DTMUV), which contains three domains, each of which has serine protease, nucleotide triphosphatase, and RNA helicase activities, respectively. It performs a variety of biological functions that are involved in the regulation of the viral life cycle and host immune response. Based on the yeast two-hybrid system, we successfully transformed pGBKT7-NS3 bait plasmid into Y2H Gold, tested it to prove that it has no self-activation and toxicity, and then hybridized it with the prey yeast strain of the duck embryo fibroblast cDNA library for screening. After high-stringency selection, positive alignment with the National Center for Biotechnology Information database revealed nine potential interactive proteins: MGST1, ERCC4, WIF1, WDR75, ACBD3, PRDX1, RPS7, ND5, and LDHA. The most interesting one (PRDX1) was selected to be verified with full-length NS3 protein and its three domains S7/DEXDc/HELICc using yeast regressive verification and GST Pull-Down assay. It denoted that PRDX1 does indeed interact with HELICc domains of NS3. NS3 is involved in the RNA uncoiling process of viral replication, which may cause mitochondrial overload to create oxidative stress (OS) during DTMUV attack. We deduced that the HELICc domain binding partner PRDX1, which regulates the p38/mitogen-activated protein kinase pathway (p38/MAPK) to avert OS, causing apoptosis, making it possible for viruses to escape host immune responses. |
| first_indexed | 2024-04-13T23:23:27Z |
| format | Article |
| id | doaj.art-7a6a84aef7484e5fa26f009bfdf77d27 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-04-13T23:23:27Z |
| publishDate | 2019-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-7a6a84aef7484e5fa26f009bfdf77d272022-12-22T02:25:09ZengMDPI AGViruses1999-49152019-08-0111874010.3390/v11080740v11080740Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay DomainsYawen Wang0Shuai Zhang1Yi Tang2Youxiang Diao3College of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Road, Tai’an 271018, ChinaCollege of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Road, Tai’an 271018, ChinaCollege of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Road, Tai’an 271018, ChinaCollege of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Road, Tai’an 271018, ChinaNS3 protein is a member of the non-structural protein of duck Tembusu virus (DTMUV), which contains three domains, each of which has serine protease, nucleotide triphosphatase, and RNA helicase activities, respectively. It performs a variety of biological functions that are involved in the regulation of the viral life cycle and host immune response. Based on the yeast two-hybrid system, we successfully transformed pGBKT7-NS3 bait plasmid into Y2H Gold, tested it to prove that it has no self-activation and toxicity, and then hybridized it with the prey yeast strain of the duck embryo fibroblast cDNA library for screening. After high-stringency selection, positive alignment with the National Center for Biotechnology Information database revealed nine potential interactive proteins: MGST1, ERCC4, WIF1, WDR75, ACBD3, PRDX1, RPS7, ND5, and LDHA. The most interesting one (PRDX1) was selected to be verified with full-length NS3 protein and its three domains S7/DEXDc/HELICc using yeast regressive verification and GST Pull-Down assay. It denoted that PRDX1 does indeed interact with HELICc domains of NS3. NS3 is involved in the RNA uncoiling process of viral replication, which may cause mitochondrial overload to create oxidative stress (OS) during DTMUV attack. We deduced that the HELICc domain binding partner PRDX1, which regulates the p38/mitogen-activated protein kinase pathway (p38/MAPK) to avert OS, causing apoptosis, making it possible for viruses to escape host immune responses.https://www.mdpi.com/1999-4915/11/8/740DTMUVyeast two-hybrid systemHELICc domains of NS3yeast regressive verificationcellular co-localizationGST Pull-Down assay |
| spellingShingle | Yawen Wang Shuai Zhang Yi Tang Youxiang Diao Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains Viruses DTMUV yeast two-hybrid system HELICc domains of NS3 yeast regressive verification cellular co-localization GST Pull-Down assay |
| title | Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains |
| title_full | Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains |
| title_fullStr | Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains |
| title_full_unstemmed | Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains |
| title_short | Screening of Duck Tembusu Virus NS3 Interacting Host Proteins and Identification of Its Specific Interplay Domains |
| title_sort | screening of duck tembusu virus ns3 interacting host proteins and identification of its specific interplay domains |
| topic | DTMUV yeast two-hybrid system HELICc domains of NS3 yeast regressive verification cellular co-localization GST Pull-Down assay |
| url | https://www.mdpi.com/1999-4915/11/8/740 |
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