<![if !vml]><![endif]>QLTR 2001-02098
Pararetroviruses, diseases, integration and genomes
October 2002 until April 2006
PARADIGM is a programme in the EU framework V coordinated by Marie-Line Caruana and Pierre-Yves Teycheney of the Centre International de Recherche Agronomique pour le Développement (CIRAD), France, and involves seven partners from France, UK, Austria, Switzerland and Spain.
Project duration: October 2002 – April 2006
Numerous endogenous pararetroviruses (EPRVs) are found to be integrated in variable copy numbers into the genome of various crops such as banana, petunia and tobacco. Although EPRVs might in some cases be simply neutral components of plant genomes, in other instances they can be infectious, express viral particles and cause severe diseases in plants. Recent experimental evidences show that EPRVs are widespread in the genome of a large number of crops and might be highly detrimental to existing and future crop improvement-oriented breeding programmes, in vitro mass propagation of vegetative crops and germplasm conservation. Our project aims at studying the biodiversity and roles of EPRVs in plant germplasms, their interactions with plant and viral genomes, the associated risks in pathogenesis, and their potential to confer viral resistance in order to develop appropriate risk assessment strategies for the control of potentially pathogenic EPRV sequences in crops relevant to European agriculture.
Overall objectives of the PARADIGM consortium
The overall research that is carried out by the consortium will be primarily based on existing experimental models (banana, tobacco, petunia) for the development of risk assessment strategies, then applied to crops relevant to European agriculture for the implementation stage. The consortium will share and apply their acknowledged complementary expertise and experience to develop experimental approaches with the specific objectives of:
<![if !supportLists]>· <![endif]>Investigating the biodiversity and evolutionary status of EPRVs in such crops;
<![if !supportLists]>· <![endif]>Screening economically important crops to European agriculture for possibly deleterious EPRVs;
<![if !supportLists]>· <![endif]>Evaluating the risks of EPRVs for germplasm conservation and for ongoing and future breeding programmes, including those aimed at creating genetically modified crops;
<![if !supportLists]>· <![endif]>Determining the mechanisms of (i) integration of EPRVs in plant genomes and (ii) those of activation, excision and expression of EPRVs leading to infectious episomal viral genomes;
<![if !supportLists]>· <![endif]>Defining the effect of EPRVs on plant genomes stability and dynamics;
<![if !supportLists]>· <![endif]>Characterizing the mechanisms developed by plants to control EPRVs, such as gene silencing, and the strategies developed by pararetroviruses to suppress such controls;
<![if !supportLists]>· <![endif]>Devising risk assessment and control strategies for EPRVs, and implementing such strategies through industrial partnerships with European plant biotech and plant breeding
To address these concerns, the PARADIGM project has been divided into 4 scientific workpackages aimed at answering clearly defined questions:
WPI: What is the exact extent and biodiversity of EPRVs within plant germplasms?
WPII: What are the mechanisms of movement of viral sequences into and out of the plant genomes?
WPIII: What are the interactions between EPRVs and plant genomes?
WPIV: What is the role of, and risk related to, EPRVs in pathogenesis?
The team in Leicester, lead by Dr Trude Schwarzacher,
was coordinating WPI and plays a leading role in examining the organisation and diversity of EPRV and related sequences in the host plant nuclear genome. Together with other partners, we are developing universal tools to isolate EPRVs from a limited number of crops where EPRV activation has already been observed, and investigate the biodiversity represented in EPRV sequences. By examination of short and long clones, and by PCR, we plan to determine the nature, organisation and sequence relationships of EPRVs between accessions of two target species groups, examining the copy numbers and chromosomal arrangement, long-range organisation and fine structure of EPRVs. Partners will develop evolutionary and structural models of EPRVs that will allow us to predict infection and expression routes. In the final task, partners will design molecular tools for identifying candidate EPRVs in any species, concentrating on five major European crops and, attempt implement them for routine screening; results, probes and primers will be delivered. We will develop S-SAP-based and inverse PCR (I-PCR) approaches for the characterisation of EPRVs integration sites and develop methods for the analysis of longer range flanking sequences.
Banana chromosomes stained with DAPI (blue) and after in situ hybridization with Banana Streak Virus probe (red). Two to eight integration sites have been detected (arrows). See Harper et al. (1999) for details.
Chromosomes of Petunia hybrida ‘Himmelsroeschen’ after DAPI staining (blue) and in situ hybridization with Petunia Vein Clearing Virus probe (red) and 5Sr DNA (green). See Richert-Poeggeler et al. (2003) for more detail.
Recent publications relevant to the subject:
Harper, G., Osuji, J.O., Heslop-Harrison, P. Hull, R. (1999). Integration of banana streak badnavirus into the Musa genome : molecular evidence. Virology 255: 207-213.
Teo CH, Tan SH, Othman YR, Schwarzacher T (2002) The cloning of Ty 1-copia-like retrotransposons from 10 varieties of banana (Musa sp). Journal of Biochemistry, Molecular Biology and Biophysics 6,193-201.
Richert-Pöggeler, K.R., Noreen, F., Schwarzacher, T., Harper, G. and Hohn, T. (2003) Induction of infectious Petunia vein clearing (pararetro) virus from endogenous provirus in petunia. EMBO Journal 22: 4836-4845. doi: 10.1093/emboj/cdg443
Hansen CN, and Heslop-Harrison JS (2004) Sequences and Phylogenies of Plant Pararetroviruses, Viruses and Transposable Elements. Advances in Botanical Research ISBN-13: 978-0-12-005941-6 41: 165-193
Hansen CN, Harper G, and Heslop-Harrison JS (2004) Characterization of pararetrovirus-like sequences in the genome of potato (Solanum tuberosum). Cytogenet Genome Res 559-565. DOI: 10.1159/000084989
C., Gregor, F., Mette, M.F.,
Hohn T, Richert-Pöggeler KR, Staginnus C, Harper G, Schwarzacher T, Chee How Teo CH, Teycheney P-Y, Iskra-Caruana ML, Hull R (2008). Evolution of integrated plant viruses. In: Plant virus evolution. (Rossinck M, ed.), Springer, Berlin, ISBN 978-3-540-75762-7. pp 53-82.
Schwarzacher T (2008) Fluorescent In Situ Hybridization to Detect Transgene Integration into Plant Genomes. in Transgenic Wheat, Barley and Oats. Huw D. Jones and Peter R. Shewry (eds.), Methods in Molecular Biology, 478, 227-246 doi: 10.1007/978-1-59745-379-0