Background
RNA silencing
Higher eukaryotes have developed a mechanism of sequence-specific RNA degradation known as ‘RNA silencing', an idiom combining the terms ‘posttranscriptional gene silencing (PTGS)' and ‘RNA interference (RNAi)'. Despite common features of RNA silencing, there are differences between the animal and plant kingdoms and also amongst species (for review see e.g . [1-6]. The central path of the RNA degradation pathway is the generation of short interfering RNA (siRNA) from a double-stranded RNA by a double-strand-specific RNase, called Dicer. The siRNAs are incorporated into the RNA-induced silencing complex (RISC) and after strand separation, the remaining single-stranded RNA strand guides the sequence- specific cleavage of a target RNA.
The mechanism of RNA silencing in plants is more complex than in most animals. Different size classes of siRNAs ranging from 21 to 24 nt can be found [7], as well as different forms of Dicers [8]. Further, plant RNA silencing is non-cell-autonomous. It can spread from an initially silenced cell to a neighboring cell and silencing can spread over a long distance to different parts of the plant [9-11].
Further information on RNA silencing phenomena can be found in the literature cited above but also in open source sites such as the ones given below.
http://en.wikipedia.org/wiki/RNA_interference
http://en.wikipedia.org/wiki/Small_interfering_RNA
Viroids
Viroids are infectious, naked circular RNAs sized from 246 to 401nt, capable of infecting a wide range of hosts, causing important economic losses [1]. They are divided into two families, Pospiviroidae and Avsunviroidae [12–14]. Potato spindle tuber viroid (PSTVd), a type species of the Pospiviroidae family, has a rod-like secondary structure with five distinct domains and replicates in the nucleus through an asymmetric rolling circle mechanism using RNA polymerase II (RNAPII) [12, 15–17]. It is known to infect crop plants of the Solanaceae family such as tomato and potato, as well as some ornamental plants of the Scrophulariaceae and Asteracae family, but does not infect the plant model Arabidopsis thaliana systemically.
Since viroids do not encode any protein they rely on plant available resources and / or mechanisms for their infectivity. One of the mechanisms they have been proposed to exploit is RNA interference (RNAi), especially because of their particular double stranded RNA structures (dsRNA) [15,16]. Further information on viroids can be found in the literature cited above but also in open source sites such as the ones given below.
https://en.wikipedia.org/wiki/Viroid
Research Interests
• RNA Silencing Pathways: RNA-mediated Regulation and Defence mechanisms
• Viroids and Viroid-Plant interactions
• Stability of Transgene expression
• Plant/RNA-Virus interactions
• Virus resistance, Viral suppressors of RNA silencing
Major contributions
- Our group was one of the first to report a number of important features of RNA silencing such as the susceptibility of viroids to the silencing mechanism [18], the conservation of the RNA silencing mechanism between plants and nematodes [19], the effects of temperature on the generation of siRNAs in plants [20] and the effects of light on S-PTGS [21].
- The group has also made important contributions in differentiating between short-range [22]) and long-range silencing and in characterising some of the rules defining the systemic movement of the silencing signal [23]. Our group was one of the first to characterise miRNA promoters in plants [24].
- The group was also the first to show the regulation of c-myc by the miR let-7 in mammalian cells [25]. Moreover our laboratory in a collaboration with the Computational Biology lab at IMBB which lead to the publication of a combined computational as well as experimental approach for predicting and verifying novel miRNA gene candidates within CAGRs [26].
- Later, we showed that the plant homologue of Enhancer of RNAi, ERL1, functions as a 3' exonuclease in the processing of rRNAs in the chloroplasts, and not in RNA silencing [27].
- Finally, we have shown that DCL4, the plant Dicer a key player in antiviral silencing pathways has a contrasting effect on viroid pathogenicity [28]. In addition, we have shown that in in contrast, it is the combined activity of DCL2 and DCL3 that defend the plant from Pospiviroids [29].
Current Research
The groups' main focus is on viroid/virus-plant interactions and RNA silencing pathways in plants. However, we are interested also in RNA silencing phenomena in diatoms and marine unicellular photosynthetic organisms with a wide genetic reservoir. In the plant RNA silencing field, we are currently trying to understand the phenomenon of spontaneous silencing of transgenes, i.e. the ability of plants to silence specifically transgene expression, a process that often leads to unstable transgenic phenotype.
In relation to viroids, we are studying how these tiny RNA pathogens not encoding for any protein can escape the plants defense mechanisms and especially the hosts RNA silencing machinery. In addition, we are aiming in elucidating how specific host factors, such as Virp1, are used by the viroid to achieve infection.
The group is also interested in RNA viruses that infect plants and especially how these pathogens manage to overcome the plant resistance mechanisms. In this direction, we have isolated virally encoded suppressors of RNA silencing and are working in elucidating their function.
Support
During the last ten years the laboratory has been successful in a number of Horizon 2020 and National grant applications securing more than 1500K research funding for the lab. In three of these projects we functioned as coordinators.
Bibliography
1. Vance, V. and H. Vaucheret, RNA silencing in plants--defense and counterdefense. Science, 2001. 292 (5525): p. 2277-80.
2. Voinnet, O., RNA silencing: small RNAs as ubiquitous regulators of gene expression. Curr Opin Plant Biol, 2002. 5 (5): p. 444-51.
3. Sontheimer, E.J., Assembly and function of RNA silencing complexes. Nat Rev Mol Cell Biol, 2005. 14 : p. 14.
4. Baulcombe, D., RNA silencing in plants. Nature, 2004. 431 (7006): p. 356-63.
5. Meister, G. and T. Tuschl, Mechanisms of gene silencing by double-stranded RNA. Nature, 2004. 431 (7006): p. 343-9.
6. Hutvagner, G., et al., Sequence-specific inhibition of small RNA function. PLoS Biol, 2004. 2 (4): p. E98.
7. Hamilton, A., et al., Two classes of short interfering RNA in RNA silencing. Embo J, 2002. 21 (17): p. 4671-9.
8. Schauer, S.E., et al., DICER-LIKE1: blind men and elephants in Arabidopsis development. Trends Plant Sci, 2002. 7 (11): p. 487-91.
9. Winston, W.M., C. Molodowitch, and C.P. Hunter, Systemic RNAi in C. elegans requires the putative transmembrane protein SID-1. Science, 2002. 295 (5564): p. 2456-9.
10. Palauqui, J.C., et al., Systemic acquired silencing: transgene-specific post-transcriptional silencing is transmitted by grafting from silenced stocks to non-silenced scions. Embo J, 1997. 16 (15): p. 4738-45.
11. Voinnet, O. and D.C. Baulcombe, Systemic signalling in gene silencing. Nature, 1997. 389 (6651): p. 553.
12. Flores R, Di Serio F, Navarro B, Duran-Vila N, Owens R. Viroids and viroid diseases of plants. Wiley-Blackwell. 2011; 307–341.
13. Gago-Zachert S. Viroids, infectious long non-coding RNAs with autonomous replication. Virus Res.; 2015;15.
14. Katsarou K, Rao ALN, Tsagris M, Kalantidis K. Infectious long non-coding RNAs. Biochimie. 2015;117: 37–47.
15. Tabler M, Tsagris M. Viroids: petite RNA pathogens with distinguished talents. Trends Plant Sci. 2004;9: 339–348.
16. Tsagris EM, et al. Viroids. Cell Microbiol. 2008;10: 2168–2179.
17. Rao ALN, Kalantidis K. Virus-associated small satellite RNAs and viroids display similarities in their replication strategies. Virology. 2015;479–480: 627–636.
18. Papaefthimiou, I., et al., Replicating potato spindle tuber viroid RNA is accompanied by short RNA fragments that are characteristic of post-transcriptional gene silencing. Nucleic Acids Res, 2001. 29 (11): p. 2395-2400.
19. Boutla, A., et al., Induction of RNA interference in Caenorhabditis elegans by RNAs derived from plants exhibiting post-transcriptional gene silencing. Nucleic Acids Res, 2002. 30 (7): p. 1688-94.
20. Kalantidis, K., et al., The occurrence of CMV-specific short Rnas in transgenic tobacco expressing virus-derived double-stranded RNA is indicative of resistance to the virus. Mol Plant Microbe Interact, 2002. 15 (8): p. 826-33.
21. Kotakis, C., et al., Light intensity affects RNA silencing of a transgene in Nicotiana benthamiana plants. BMC Plant Biol. 10 : p. 220.
22. Tournier, B, et al. Phloem flow strongly influences the systemic spread of silencing in GFP Nicotiana benthamiana plants. Plant J, 2006. 47 (3): p. 383-94.
23. Megraw M, et al. MicroRNA Promoter Element Discovery in Arabidopsis. RNA, 2006, 12(9): 1612.
24. Koscianska, E., et al., Prediction and preliminary validation of oncogene regulation by miRNAs. BMC Mol Biol, 2007. 8 : p. 79.
25. Oulas, A., et al., Prediction of novel microRNA genes in cancer-associated genomic regions--a combined computational and experimental approach. Nucleic Acids Res, 2009. 37 (10): p. 3276-87.
26. Mermigka G, Verret F., and Kalantidis K.*. 2016. RNA silencing movement in plants
Invited expert review, JIPB, 2016 Apr;58(4):328-42.
27. Dadami E. et al. . 2013. RNA silencing pathways may have a positive effect on Potato spindle tuber viroid infectivity in Nicotiana benthamiana. Mol. Plant 6:232-234.
28. Katsarou K, et al. Combined Activity of DCL2 and DCL3 is crucial in The Defense Against Potato Spindle Tuber Viroid.. PLoS Path., 2016, In Press
29. Dimopoulou A, Theologidis I, Liebmann B, Kalantidis K, Vassilakos N, Skandalis N. 2019. Bacillus amyloliquefaciens MBI600 differentially induces tomato defense signaling pathways depending on plant part and dose of application. Sci Rep. 2019 Dec 13;9(1):19120. doi: 10.1038/s41598-019-55645-2.
30. Katsarou K, Bardani E, Kallemi P, Kalantidis K. 2019. Viral Detection: Past, Present, and Future. Bioessays. 2019 Aug 22:e1900049. doi: 10.1002/bies.201900049.
31. Katsarou K, Chiumenti M, Kalantidis K, Mathioudakis MM. 2020. First Report of Citrus Viroids Infecting Persian (Tahiti) Lime in Greece. Plant Disease, PDIS-07-19-1385-PDN
32. Pecinka A, Chevalier C, Colas I, Kalantidis K, Varotto S, Krugman T, Michaelidis C, Vallés M-P, r Muñoz A, Pradillo M. 2020. Chromatin dynamics during interphase and cell division: similarities and differences between model and crop plants. J. of Exp. Botany, In Press.