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Article ; Online: Viroids as a Tool to Study RNA-Directed DNA Methylation in Plants.

Wassenegger, Michael / Dalakouras, Athanasios

2021 Volume 10, Issue 5

Abstract: Viroids are plant pathogenic, circular, non-coding, single-stranded RNAs (ssRNAs). Members of ... ...

Abstract	Viroids are plant pathogenic, circular, non-coding, single-stranded RNAs (ssRNAs). Members of the
MeSH term(s)	DNA Methylation ; DNA, Plant/genetics ; DNA, Plant/metabolism ; Gene Expression Regulation, Plant ; Plant Viruses/genetics ; Plants/genetics ; Plants/virology ; Plants, Genetically Modified/genetics ; Plants, Genetically Modified/virology ; RNA Interference ; RNA, Plant/genetics ; Viroids/genetics
Chemical Substances	DNA, Plant ; RNA, Plant
Language	English
Publishing date	2021-05-13
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	2661518-6
ISSN	2073-4409 ; 2073-4409
ISSN (online)	2073-4409
ISSN	2073-4409
DOI	10.3390/cells10051187
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Host-induced gene silencing - mechanisms and applications.

Koch, Aline / Wassenegger, Michael

The New phytologist

2021 Volume 231, Issue 1, Page(s) 54–59

Abstract: Host-induced gene silencing (HIGS) technology has emerged as a powerful alternative to chemical treatments for protecting plants from pathogens or pests. More than 170 HIGS studies have been published so far, and HIGS products have been launched. First, ... ...

Abstract	Host-induced gene silencing (HIGS) technology has emerged as a powerful alternative to chemical treatments for protecting plants from pathogens or pests. More than 170 HIGS studies have been published so far, and HIGS products have been launched. First, we discuss the strengths and limitations of this technology in a pathosystem-specific context. Next, we highlight the requirement for fundamental knowledge on the molecular mechanisms (i.e. uptake, processing and translocation of transgene-expressed double-stranded RNAs) that determine the efficacy and specificity of HIGS. Additionally, we speculate on the contribution of host and target RNA interference machineries, which may be incompatible depending on the lifestyle of the pathogen or pest. Finally, we predict that closing these gaps in knowledge will lead to the development of novel integrative concepts, precise risk assessment and tailor-made HIGS therapy for plant diseases.
MeSH term(s)	Gene Silencing ; Plant Diseases/genetics ; Plants/genetics ; RNA Interference ; RNA, Double-Stranded
Chemical Substances	RNA, Double-Stranded
Language	English
Publishing date	2021-05-02
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	208885-x
ISSN	1469-8137 ; 0028-646X
ISSN (online)	1469-8137
ISSN	0028-646X
DOI	10.1111/nph.17364
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Critical view on RNA silencing-mediated virus resistance using exogenously applied RNA.

Uslu, Veli V / Wassenegger, Michael

Current opinion in virology

2020 Volume 42, Page(s) 18–24

Abstract: In almost all eukaryotes, RNA interference (RNAi) is a natural defence mechanism against foreign nucleic acids, including transposons and viruses. It is generally triggered by long double stranded RNA molecules (dsRNA, >50bp) that are processed into ... ...

Abstract	In almost all eukaryotes, RNA interference (RNAi) is a natural defence mechanism against foreign nucleic acids, including transposons and viruses. It is generally triggered by long double stranded RNA molecules (dsRNA, >50bp) that are processed into small interfering RNAs (siRNAs). RNAi can be artificially activated by the expression of RNAi triggers through viruses (virus-induced gene silencing, VIGS) and transgenes. Moreover, for almost 10 years, exogenous RNA application methods are developed as tools to induce RNAi in plants. In this review, exogenous RNA application techniques having the potential to activate RNAi with a focus on RNAi-mediated virus resistance will be discussed. Limitations of exogenous RNA applications, targeting of virus vectors and open questions related to mechanistic details that still require further investigation will be pointed out.
MeSH term(s)	Disease Resistance ; Plant Diseases/genetics ; Plant Diseases/immunology ; Plant Diseases/virology ; RNA Interference ; RNA Viruses/genetics ; RNA Viruses/physiology ; RNA, Small Interfering/genetics ; RNA, Small Interfering/immunology
Chemical Substances	RNA, Small Interfering
Language	English
Publishing date	2020-05-01
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	2611378-8
ISSN	1879-6265 ; 1879-6257
ISSN (online)	1879-6265
ISSN	1879-6257
DOI	10.1016/j.coviro.2020.03.004
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Viroids as a Tool to Study RNA-Directed DNA Methylation in Plants

Michael Wassenegger / Athanasios Dalakouras

Cells, Vol 10, Iss 1187, p

2021 Volume 1187

Abstract: Viroids are plant pathogenic, circular, non-coding, single-stranded RNAs (ssRNAs). Members of the Pospiviroidae family replicate in the nucleus of plant cells through double-stranded RNA (dsRNA) intermediates, thus triggering the host’s RNA interference ( ...

Abstract	Viroids are plant pathogenic, circular, non-coding, single-stranded RNAs (ssRNAs). Members of the Pospiviroidae family replicate in the nucleus of plant cells through double-stranded RNA (dsRNA) intermediates, thus triggering the host’s RNA interference (RNAi) machinery. In plants, the two RNAi pillars are Post-Transcriptional Gene Silencing (PTGS) and RNA-directed DNA Methylation (RdDM), and the latter has the potential to trigger Transcriptional Gene Silencing (TGS). Over the last three decades, the employment of viroid-based systems has immensely contributed to our understanding of both of these RNAi facets. In this review, we highlight the role of Pospiviroidae in the discovery of RdDM, expound the gradual elucidation through the years of the diverse array of RdDM’s mechanistic details and propose a revised RdDM model based on the cumulative amount of evidence from viroid and non-viroid systems.
Keywords	viroids ; RNA-directed DNA methylation ; small RNAs ; RNA interference ; epigenetics ; bisulfite sequencing ; Biology (General) ; QH301-705.5
Subject code	580
Language	English
Publishing date	2021-05-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article: Host‐induced gene silencing – mechanisms and applications

Koch, Aline / Wassenegger, Michael

The new phytologist. 2021 July, v. 231, no. 1

2021

Abstract: Host‐induced gene silencing (HIGS) technology has emerged as a powerful alternative to chemical treatments for protecting plants from pathogens or pests. More than 170 HIGS studies have been published so far, and HIGS products have been launched. First, ... ...

Abstract	Host‐induced gene silencing (HIGS) technology has emerged as a powerful alternative to chemical treatments for protecting plants from pathogens or pests. More than 170 HIGS studies have been published so far, and HIGS products have been launched. First, we discuss the strengths and limitations of this technology in a pathosystem‐specific context. Next, we highlight the requirement for fundamental knowledge on the molecular mechanisms (i.e. uptake, processing and translocation of transgene‐expressed double‐stranded RNAs) that determine the efficacy and specificity of HIGS. Additionally, we speculate on the contribution of host and target RNA interference machineries, which may be incompatible depending on the lifestyle of the pathogen or pest. Finally, we predict that closing these gaps in knowledge will lead to the development of novel integrative concepts, precise risk assessment and tailor‐made HIGS therapy for plant diseases.
Keywords	RNA interference ; genes ; lifestyle ; pathogens ; pests ; risk assessment ; therapeutics
Language	English
Dates of publication	2021-07
Size	p. 54-59.
Publishing place	John Wiley & Sons, Ltd
Document type	Article
Note	NAL-AP-2-clean ; REVIEW
ZDB-ID	208885-x
ISSN	1469-8137 ; 0028-646X
ISSN (online)	1469-8137
ISSN	0028-646X
DOI	10.1111/nph.17364
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Beet Curly Top Iran Virus

Ebrahimi, Saeideh / Eini, Omid / Baßler, Alexandra / Hanke, Arvid / Yildirim, Zeynep / Wassenegger, Michael / Krczal, Gabi / Uslu, Veli Vural

Viruses

2023 Volume 15, Issue 10

Abstract: Beet curly top Iran ... ...

Abstract	Beet curly top Iran virus
MeSH term(s)	RNA Interference ; Beta vulgaris ; Viral Proteins/genetics ; Viral Proteins/metabolism ; Iran ; Geminiviridae ; Nicotiana ; Plant Diseases
Chemical Substances	Viral Proteins
Language	English
Publishing date	2023-09-26
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2516098-9
ISSN	1999-4915 ; 1999-4915
ISSN (online)	1999-4915
ISSN	1999-4915
DOI	10.3390/v15101996
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: High-pressure sprayed siRNAs influence the efficiency but not the profile of transitive silencing.

Uslu, Veli Vural / Dalakouras, Athanasios / Steffens, Victor A / Krczal, Gabi / Wassenegger, Michael

The Plant journal : for cell and molecular biology

2022 Volume 109, Issue 5, Page(s) 1199–1212

Abstract: In plants, small interfering RNAs (siRNAs) are a quintessential class of RNA interference (RNAi)-inducing molecules produced by the endonucleolytic cleavage of double-stranded RNAs (dsRNAs). In order to ensure robust RNAi, siRNAs are amplified through a ... ...

Abstract	In plants, small interfering RNAs (siRNAs) are a quintessential class of RNA interference (RNAi)-inducing molecules produced by the endonucleolytic cleavage of double-stranded RNAs (dsRNAs). In order to ensure robust RNAi, siRNAs are amplified through a positive feedback mechanism called transitivity. Transitivity relies on RNA-DIRECTED RNA POLYMERASE 6 (RDR6)-mediated dsRNA synthesis using siRNA-targeted RNA. The newly synthesized dsRNA is subsequently cleaved into secondary siRNAs by DICER-LIKE (DCL) endonucleases. Just like primary siRNAs, secondary siRNAs are also loaded into ARGONAUTE proteins (AGOs) to form an RNA-induced silencing complex reinforcing the cleavage of the target RNA. Although the molecular players underlying transitivity are well established, the mode of action of transitivity remains elusive. In this study, we investigated the influence of primary target sites on transgene silencing and transitivity using the green fluorescent protein (GFP)-expressing Nicotiana benthamiana 16C line, high-pressure spraying protocol, and synthetic 22-nucleotide (nt) long siRNAs. We found that the 22-nt siRNA targeting the 3' of the GFP transgene was less efficient in inducing silencing when compared with the siRNAs targeting the 5' and middle region of the GFP. Moreover, sRNA sequencing of locally silenced leaves showed that the amount but not the profile of secondary RNAs is shaped by the occupancy of the primary siRNA triggers on the target RNA. Our findings suggest that RDR6-mediated dsRNA synthesis is not primed by primary siRNAs and that dsRNA synthesis appears to be generally initiated at the 3'-end of the target RNA.
MeSH term(s)	Green Fluorescent Proteins/genetics ; RNA Interference ; RNA, Double-Stranded/genetics ; RNA, Small Interfering/genetics ; RNA, Small Interfering/metabolism ; RNA-Dependent RNA Polymerase/genetics ; RNA-Induced Silencing Complex/genetics
Chemical Substances	RNA, Double-Stranded ; RNA, Small Interfering ; RNA-Induced Silencing Complex ; Green Fluorescent Proteins (147336-22-9) ; RNA-Dependent RNA Polymerase (EC 2.7.7.48)
Language	English
Publishing date	2022-01-19
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1088037-9
ISSN	1365-313X ; 0960-7412
ISSN (online)	1365-313X
ISSN	0960-7412
DOI	10.1111/tpj.15625
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: High-Pressure-Sprayed Double Stranded RNA Does Not Induce RNA Interference of a Reporter Gene.

Uslu, Veli Vural / Bassler, Alexandra / Krczal, Gabi / Wassenegger, Michael

Frontiers in plant science

2020 Volume 11, Page(s) 534391

Abstract: In plants, RNA interference (RNAi) is an effective defense mechanism against pathogens and pests. RNAi mainly involves the micro RNA and the small interfering RNA (siRNA) pathways. The latter pathway is generally based on the processing of long double ... ...

Abstract	In plants, RNA interference (RNAi) is an effective defense mechanism against pathogens and pests. RNAi mainly involves the micro RNA and the small interfering RNA (siRNA) pathways. The latter pathway is generally based on the processing of long double stranded RNAs (dsRNA) into siRNAs by DICER-LIKE endonucleases (DCLs). SiRNAs are loaded onto ARGONAUTE proteins to constitute the RNA-induced silencing complex (RISC). Natural dsRNAs derive from transcription of inverted repeats or of specific RNA molecules that are transcribed by RNA-directed RNA polymerase 6 (RDR6). Moreover, replication of infecting viruses/viroids results in the production of dsRNA intermediates that can serve as substrates for DCLs. The high effectiveness of RNAi both locally and systemically implicated that plants could become resistant to pathogens, including viruses, through artificial activation of RNAi by topical exogenous application of dsRNA. The most preferable procedure to exploit RNAi would be to simply spray naked dsRNAs onto mature plants that are specific for the attacking pathogens serving as a substitute for pesticides applications. However, the plant cell wall is a difficult barrier to overcome and only few reports claim that topical application of naked dsRNA triggers RNAi in plants. Using a transgenic
Language	English
Publishing date	2020-12-16
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2613694-6
ISSN	1664-462X
ISSN	1664-462X
DOI	10.3389/fpls.2020.534391
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Article ; Online: RNA-directed DNA methylation efficiency depends on trigger and target sequence identity.

Dalakouras, Athanasios / Dadami, Elena / Wassenegger, Michèle / Krczal, Gabi / Wassenegger, Michael

The Plant journal : for cell and molecular biology

2016 Volume 87, Issue 2, Page(s) 202–214

Abstract: RNA-directed DNA methylation (RdDM) in plants has been extensively studied, but the RNA molecules guiding the RdDM machinery to their targets are still to be characterized. It is unclear whether these molecules require full complementarity with their ... ...

Abstract	RNA-directed DNA methylation (RdDM) in plants has been extensively studied, but the RNA molecules guiding the RdDM machinery to their targets are still to be characterized. It is unclear whether these molecules require full complementarity with their target. In this study, we have generated Nicotiana tabacum (Nt) plants carrying an infectious tomato apical stunt viroid (TASVd) transgene (Nt-TASVd) and a non-infectious potato spindle tuber viroid (PSTVd) transgene (Nt-SB2). The two viroid sequences exhibit 81% sequence identity. Nt-TASVd and Nt-SB2 plants were genetically crossed. In the progeny plants (Nt-SB2/TASVd), deep sequencing of small RNAs (sRNAs) showed that TASVd infection was associated with the accumulation of abundant small interfering RNAs (siRNAs) that mapped along the entire TASVd but only partially matched the SB2 transgene. TASVd siRNAs efficiently targeted SB2 RNA for degradation, but no transitivity was detectable. Bisulfite sequencing in the Nt-SB2/TASVd plants revealed that the TASVd transgene was targeted for dense cis-RdDM along its entire sequence. In the same plants, the SB2 transgene was targeted for trans-RdDM. The SB2 methylation pattern, however, was weak and heterogeneous, pointing to a positive correlation between trigger-target sequence identity and RdDM efficiency. Importantly, trans-RdDM on SB2 was also detected at sites where no homologous siRNAs were detected. Our data indicate that RdDM efficiency depends on the trigger-target sequence identity, and is not restricted to siRNA occupancy. These findings support recent data suggesting that RNAs with sizes longer than 24 nt (>24-nt RNAs) trigger RdDM.
MeSH term(s)	Blotting, Northern ; DNA Methylation/genetics ; DNA Methylation/physiology ; Plants, Genetically Modified/genetics ; Plants, Genetically Modified/metabolism ; Plants, Genetically Modified/physiology ; RNA, Plant/genetics ; RNA, Plant/physiology ; Nicotiana/genetics ; Nicotiana/metabolism ; Nicotiana/physiology
Chemical Substances	RNA, Plant
Language	English
Publishing date	2016-07-26
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1088037-9
ISSN	1365-313X ; 0960-7412
ISSN (online)	1365-313X
ISSN	0960-7412
DOI	10.1111/tpj.13193
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: High‐pressure sprayed siRNAs influence the efficiency but not the profile of transitive silencing

Uslu, Veli Vural / Dalakouras, Athanasios / Steffens, Victor A. / Krczal, Gabi / Wassenegger, Michael

plant journal. 2022 Mar., v. 109, no. 5

2022

Abstract: In plants, small interfering RNAs (siRNAs) are a quintessential class of RNA interference (RNAi)‐inducing molecules produced by the endonucleolytic cleavage of double‐stranded RNAs (dsRNAs). In order to ensure robust RNAi, siRNAs are amplified through a ... ...

Abstract	In plants, small interfering RNAs (siRNAs) are a quintessential class of RNA interference (RNAi)‐inducing molecules produced by the endonucleolytic cleavage of double‐stranded RNAs (dsRNAs). In order to ensure robust RNAi, siRNAs are amplified through a positive feedback mechanism called transitivity. Transitivity relies on RNA‐DIRECTED RNA POLYMERASE 6 (RDR6)‐mediated dsRNA synthesis using siRNA‐targeted RNA. The newly synthesized dsRNA is subsequently cleaved into secondary siRNAs by DICER‐LIKE (DCL) endonucleases. Just like primary siRNAs, secondary siRNAs are also loaded into ARGONAUTE proteins (AGOs) to form an RNA‐induced silencing complex reinforcing the cleavage of the target RNA. Although the molecular players underlying transitivity are well established, the mode of action of transitivity remains elusive. In this study, we investigated the influence of primary target sites on transgene silencing and transitivity using the green fluorescent protein (GFP)‐expressing Nicotiana benthamiana 16C line, high‐pressure spraying protocol, and synthetic 22‐nucleotide (nt) long siRNAs. We found that the 22‐nt siRNA targeting the 3ʹ of the GFP transgene was less efficient in inducing silencing when compared with the siRNAs targeting the 5ʹ and middle region of the GFP. Moreover, sRNA sequencing of locally silenced leaves showed that the amount but not the profile of secondary RNAs is shaped by the occupancy of the primary siRNA triggers on the target RNA. Our findings suggest that RDR6‐mediated dsRNA synthesis is not primed by primary siRNAs and that dsRNA synthesis appears to be generally initiated at the 3ʹ‐end of the target RNA.
Keywords	Nicotiana benthamiana ; RNA interference ; RNA-directed RNA polymerase ; green fluorescent protein ; mechanism of action ; transgenes
Language	English
Dates of publication	2022-03
Size	p. 1199-1212.
Publishing place	John Wiley & Sons, Ltd
Document type	Article
Note	JOURNAL ARTICLE
ZDB-ID	1088037-9
ISSN	1365-313X ; 0960-7412
ISSN (online)	1365-313X
ISSN	0960-7412
DOI	10.1111/tpj.15625
Database	NAL-Catalogue (AGRICOLA)

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