scholarly journals Dynamics of the VIGS-Mediated Chimeric Silencing of the Nicotiana benthamiana ChlH Gene and of the Tobacco Mosaic Virus Vector

2003 ◽  
Vol 16 (2) ◽  
pp. 99-106 ◽  
Author(s):  
Jean-Baptiste Hiriart ◽  
Eva-Mari Aro ◽  
Kirsi Lehto
Keyword(s):  
Gene Silencing ◽  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Target Gene ◽  
Virus Vector ◽  
Regulation System ◽  
Transcriptional Gene Silencing ◽  
Mrna Levels ◽  
Partial Recovery ◽  
Tmv Vector

The ChlH gene, encoding for the H subunit of the magnesium chelatase enzyme, was silenced in Nicotiana bentahamiana plants by virus-induced gene silencing (VIGS), using tobacco mosaic virus (TMV) expression vector. Strong silencing of the ChlH target gene was initiated only in the apical tissues, in which the endogenous transcription level of the target gene and the level of TMV vector RNA were both very high. The virus vector was also targeted by VIGS, and its suppression was correlated with the silencing of the ChlH mRNA. In the apical tissues, the suppression of both the virus vector and the ChlH mRNA led to a reduction of the silencing pressure and, consequently, to partial recovery of the new growth from the silencing. As the virus vector and the target mRNA levels increased, silencing was reestablished. The feedback regulation system, caused by the transient increase and reduction in levels of the virus vector and ChlH mRNA, led to a fluctuation of the silenced and recovered phenotypes in the plant apex. This TMV-vector mediated silencing system differed from previously analyzed VIGS systems; although the TMV vector was initially targeted by the silencing system, it was not permanently suppressed, indicating that, in this system, TMV was able to effectively escape post-transcriptional gene silencing.

scholarly journals Rapid induction of transcriptional and post-transcriptional gene silencing using a novel Cucumber mosaic virus vector

2006 ◽  
Vol 23 (3) ◽  
pp. 259-265 ◽  
Author(s):  
Shungo Otagaki ◽  
Makoto Arai ◽  
Akiko Takahashi ◽  
Kazunori Goto ◽  
Jin-Sung Hong ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Virus Vector ◽  
Transcriptional Gene Silencing ◽  
Rapid Induction ◽  
Post Transcriptional Gene Silencing

scholarly journals Stability of Barley stripe mosaic virus–Induced Gene Silencing in Barley

2007 ◽  
Vol 20 (11) ◽  
pp. 1323-1331 ◽  
Author(s):  
Marianne Bruun-Rasmussen ◽  
Christian Toft Madsen ◽  
Stine Jessing ◽  
Merete Albrechtsen
Keyword(s):  
Gene Silencing ◽  
Mosaic Virus ◽  
Barley Stripe Mosaic Virus ◽  
Mrna Levels ◽  
Virus Induced Gene Silencing ◽  
Qrt Pcr ◽  
Transient Nature ◽  
Polymerase Chain ◽  
Insert Length ◽  
Pds Gene

Virus-induced gene silencing (VIGS) can be used as a powerful tool for functional genomics studies in plants. With this approach, it is possible to target most genes and downregulate the messenger (m)RNA in a sequence-specific manner. Barley stripe mosaic virus (BSMV) is an established VIGS vector for barley and wheat; however, silencing using this vector is generally transient, with efficient silencing often being confined to the first two or three systemically infected leaves. To investigate this further, part of the barley Phytoene desaturase (PDS) gene was inserted into BSMV and the resulting photobleaching in infected barley plants was used as a reporter for silencing. In addition, downregulation of PDS mRNA was measured by quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR). Using fragments of PDS ranging from 128 to 584 nucleotides in BSMV, we observed that insert length influenced stability but not efficiency of VIGS. Silencing was transient in most cases; however, the decrease in PDS mRNA levels measured by qRT-PCR began earlier and lasted longer than the photobleaching. Occasionally, silencing persisted and could be transmitted through seed as well as via mechanical inoculation, although large parts of the insert had been lost from the virus vector. The instability of the insert, observed consistently throughout our experiments, offers an explanation for the transient nature of silencing when using BSMV as a VIGS vector.

scholarly journals Cucumber mosaic virus Infection Transiently Breaks dsRNA-Induced Transgenic Immunity to Potato virus Y in Tobacco

2003 ◽  
Vol 16 (10) ◽  
pp. 936-944 ◽  
Author(s):  
Neena Mitter ◽  
Emy Sulistyowati ◽  
Ralf G. Dietzgen
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Potato Virus ◽  
Potato Virus Y ◽  
Defense Mechanism ◽  
Transcriptional Gene Silencing ◽  
Mrna Levels ◽  
Post Transcriptional Gene Silencing ◽  
The Stability ◽  
High Level

Post-transcriptional gene silencing (PTGS), an intrinsic plant defense mechanism, can be efficiently triggered by double stranded (ds)RNA-producing transgenes and can provide high level virus resistance by specific targeting of cognate viral RNA. The discovery of virus-encoded suppressors of PTGS led to concerns about the stability of such resistance. Here, we show that Cucumber mosaic virus (CMV) is able to suppress dsRNA-induced PTGS and the associated Potato virus Y (PVY) immunity in tobacco. CMV suppression supported only a transient PVY accumulation and did not prevent recovery of the transgenic plants from PVY infection. CMV inoculation resulted in strongly increased transgene mRNA levels due to suppression of PTGS, but accumulation of PVY-specific small interfering (si)RNA was unaffected. However, PVY accumulation in previously immune plants resulted in increased PVY siRNA levels and transgene mRNA was no longer detected, despite the presence of CMV. Transgene mRNA returned to high levels once PVY was no longer detected in CMV-infected plants. Recovered and chronically CMV-infected tissues were immune to further PVY infection.

scholarly journals A Foxtail mosaic virus Vector for Virus-Induced Gene Silencing in Maize

2016 ◽  
pp. pp.00172.2016 ◽  
Author(s):  
Yu Mei ◽  
Chunquan Zhang ◽  
Bliss M. Kernodle ◽  
John H. Hill ◽  
Steven A. Whitham
Keyword(s):  
Gene Silencing ◽  
Mosaic Virus ◽  
Virus Vector ◽  
Virus Induced Gene Silencing

scholarly journals A High Throughput Barley Stripe Mosaic Virus Vector for Virus Induced Gene Silencing in Monocots and Dicots

PLoS ONE ◽  
2011 ◽  
Vol 6 (10) ◽  
pp. e26468 ◽  
Author(s):  
Cheng Yuan ◽  
Cui Li ◽  
Lijie Yan ◽  
Andrew O. Jackson ◽  
Zhiyong Liu ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Mosaic Virus ◽  
High Throughput ◽  
Virus Vector ◽  
Barley Stripe Mosaic Virus ◽  
Virus Induced Gene Silencing

scholarly journals Discriminating Mutations of HC-Pro of Zucchini yellow mosaic virus with Differential Effects on Small RNA Pathways Involved in Viral Pathogenicity and Symptom Development

2010 ◽  
Vol 23 (1) ◽  
pp. 17-28 ◽  
Author(s):  
Hui-Wen Wu ◽  
Shih-Shun Lin ◽  
Kuan-Chun Chen ◽  
Shyi-Dong Yeh ◽  
Nam-Hai Chua
Keyword(s):  
Amino Acids ◽  
Gene Silencing ◽  
Mosaic Virus ◽  
Symptom Severity ◽  
Zucchini Yellow Mosaic Virus ◽  
Yellow Mosaic Virus ◽  
Transcriptional Gene Silencing ◽  
Molecular Evidence ◽  
Virus Induced Gene Silencing ◽  
Conserved Amino Acids

Helper component-proteinase (HC-Pro), the gene-silencing suppressor of Potyvirus spp., interferes with microRNA (miRNA) and short-interfering RNA (siRNA) pathways. Our previous studies showed that three mutations of highly conserved amino acids of HC-Pro, R180I (mutation A), F205L (B), and E396N (C), of Zucchini yellow mosaic virus (ZYMV) affect symptom severity and viral pathogenicity. The mutant ZYMV GAC (ZGAC) with double mutations, R180I/E396N, induces transient leaf mottling in host plants followed by recovery. This mutant confers complete cross protection against subsequent infection by the parental ZYMV (ZG) strain. Here, we sought to obtain molecular evidence on the roles of the three highly conserved amino acids of HC-Pro in miRNA and siRNA pathways using transgenic Arabidopsis plants expressing comparable levels of wild-type and mutant HC-Pro proteins. We demonstrated that amino acid residues 180, 205, and 396 of HC-Pro are critical for suppression of miRNA, trans-acting siRNA (ta-siRNA), and virus-induced gene silencing (VIGS) pathways but not for sense-post transcriptional gene silencing (s-PTGS). Because the HC-Pro double mutant (R180I/E396N) does not interfere with miRNA and ta-siRNA pathways, the ZGAC mutant virus elicits only attenuated symptoms. Furthermore, the recovery seen on ZGAC-infected plants likely results from the weak VIGS suppression by the HC-Pro double AC mutant. Thus, through manipulating these three conserved amino acids on HC-Pro, symptom severity of diseases caused by Potyvirus spp. can be modulated to generate useful cross protectants for field application. Although some of our mutated HC-Pro proteins do not interfere with miRNA and ta-siRNA pathways, they still retain the ability to suppress s-PTGS.

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