Effect of PVA infection on the mRNA expression of AGO1, AGO2, and SUO1 homologs from Nicotiana benthamiana

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http://urn.fi/URN:NBN:fi:hulib-202106183153
Title: Effect of PVA infection on the mRNA expression of AGO1, AGO2, and SUO1 homologs from Nicotiana benthamiana
Author: Xhelilaj, Kaltra
Other contributor: Helsingin yliopisto, Maatalous-metsätieteellinen tiedekunta
University of Helsinki, Faculty of Agriculture and Forestry
Helsingfors universitet, Agrikultur- och forstvetenskapliga fakulteten
Publisher: Helsingin yliopisto
Date: 2021
Language: eng
URI: http://urn.fi/URN:NBN:fi:hulib-202106183153
http://hdl.handle.net/10138/331555
Thesis level: master's thesis
Degree program: Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Specialisation: Kasvintuotantotieteet
Plant Production Sciences
Växtproduktionsvetenskaper
Abstract: Potyviruses are positive-sense single-stranded RNA viruses that can alter several functions of their host plants and consequently, cause significant economic losses in the infected crop plants. During the viral infection, the host transcriptome changes. Stress related genes are triggered, and genes allowing for susceptibility are target for viral-induced modifications. Therefore, in this study, we investigated whether the expression of potential proviral genes SUO1, AGO1, and the major antiviral player AGO2 change in Nicotiana benthamiana (N. benthamiana) in response to potato virus A (PVA; genus Potyvirus) infection. Moreover, we aimed to determine whether helper component protease (HCPro) and active replication have a role in the transcriptional regulation of these genes. Leaves infected with PVA tagged with Renilla luciferase were collected at 3, 6, and 9 days postinoculation, and the viral gene expression was quantified with a dual-luciferase assay. Total RNA was isolated, cDNA was synthesized, and samples were analyzed through qPCR. BLAST hit results revealed that N. benthamiana has three homologs of the SUO1 gene. qPCR data showed no significant change in neither the expression of SUO1 homologs nor the expression of AGO1 during wild-type PVA infection. Moreover, the lack of HCPro or viral replication did not affect the expression of these genes. On the other hand, the expression of AGO2 was approximately 6 and 5 fold up-regulated at 6 and 9 days post-inoculation, respectively. In contrast with the wild-type PVA infection, the mutated viruses had a pronounced effect on AGO2 transcripts at 3 days post-inoculation. Replication-deficient viral RNA increased AGO2 expression circa four-fold, followed by the HCPro-deficient viral RNA increasing expression circa two-fold. AGO2, the major player involved in antiviral defense, was up-regulated during the wild-type infection. Active viral replication and functional HCPro played a role in AGO2 regulation. However, Agrobacterium infiltration can be accounted for interfering with the interpretation of the AGO2 results. Although SUO1 and AGO1 may be potential genes allowing for susceptibility, this study revealed that the PVA infection does not affect the mRNA expression of these genes. Furthermore, it is concluded that active HCPro and viral replication do not have a role in the expression of these genes on mRNA level. To have a clearer view, integrating small RNA, mRNA, and protein quantification analysis of SUO1 homologs will be necessary. Keywords
Subject: potyviruses
PVA
RT-qPCR
SUO1
AGO1
AGO2
proviral
gene expression.


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