In my field of research – plant telomere biology – it happens quite often at conferences that non-plant telomere scientists ignore findings obtained in plants – at least one my parallel with Mendel’s unnoticed work, though not particularly satisfactory one. Once I even got a question from a recognised colleague working on human telomeres – do plants also have telomeres?
Well, they do. And not only that: telomeres have been discovered in plants (and in parallel in Drosophila flies). Plants can also "boast" other firsts – e.g., cells were discovered in plants (1667), genes were discovered in plants (at St. Thomas Abbey in Brno, 1865), as was also a semiconservative DNA replication (1957, one year before the E.coli experiment awarded by the Nobel Prize)... Apparently, a scientist working in plants has to be prepared to be ignored.
Why does it make sense to investigate plant telomeres and telomerases while most labs concentrate on human cells where the applicability of the telomere biology research in cancer and aging is certainly more direct?
My personal reason or answer is the following: in contrast to humans or other vertebrates, in plants there is no developmental shortening of telomeres (telomere-based aging clock) – the phenomenon associated with the reversible regulation of plant telomerase activity, in contrast to its permanent silencing in somatic cells during human embryonic development. Briefly, whenever a plant cell needs to divide, it can – somehow – re-activate its telomerase. Even differentiated somatic plant cells with a silenced telomerase, when contributing to plant regeneration or formation of new plant organs, re-activate their telomerase – something that we as humans can only envy to plants. When considering that these were among my first findings in telomere biology as a junior researcher (1995-98), then you can understand my very personal relationship to plant telomere biology (Fajkus et al. 1998, Fajkus et al. 1996, Fajkus et al. 1995, Riha et al. 1998).
The research story I would like to share with you spans over the last 6 years of our research.
Since 1995, it has been known that some plants possess a different kind of telomeric DNA than the most common (TTTAGGG)n repeat motif (Fuchs et al. 1995, Pich et al. 1996). The most interesting of these were plants of the genus Allium which lacked not only the typical plant telomere motif but also any other known variant of telomere repeat (Sykorova et al. 2003). It was even believed that Allium plants lacked telomerase and used some telomerase-independent mechanism of telomere maintenance, known e.g., in Drosophila. Nevertheless, in 2016 we were able to characterize unusual – 12 nt long – (TTATGGGCTCGG)n telomere repeats in 11 Allium species and demonstrated their synthesis by telomerase (Fajkus et al. 2016).
The extremely long telomere repeat size in Allium allowed us to identify putative telomerase RNA subunit (TR) in genomic and transcriptomic data of Allium species and verify its function experimentally. Subsequently it appeared that in contrast to the common opinion (based on findings in vertebrates, yeasts, and also previous reports in Arabidopsis (Cifuentes-Rojas et al. 2011), now retracted, we found that plant TRs are of monophyletic origin. Therefore, using the initial knowledge of TR in Allium, we could successfully identify TRs across the plant order of Asparagales, and then across the whole land plant phylogeny. The identified TRs and changes in their template regions provided molecular explanations of all previously reported evolutionary changes of plant telomere DNA repeats. Also, in contrast to the other known TRs (with the exception of Ciliates), plant TRs are transcribed with RNA polymerase III under the control of so-called type-3 promoter (and not by RNA pol II as in classical model organisms - vertebrates or yeasts). The type-3 promoter, consisting of the Upstream sequence element (USE) and TATA box is typical for a number of small nuclear RNAs, including spliceosomal U snRNAs. We believe that this study and the described relatively easy way to identify telomerase RNAs in any plant, including all important crops, introduced plants to the set of model species in telomerase research, at last (Fajkus et al. 2019).
Unfortunately, sequence divergence of TRs across the whole Viridiplantae is too high to use simple homology searches beyond land plants group. Therefore, in the subsequent study, we had to design a novel strategy, presuming a wider conservation of the type-3 promoter also in early diverging plants – e.g., Bryophytes, Streptophyte algae and green algae. We started with identification of the USE element in snRNAs, and identification of telomere DNA repeats (usually the most frequent short tandem repeats in genomic data), as identified by Tandem Repeats Finder tool. The telomere DNA sequence provided the information about a putative template region of TR - serving to synthesize telomere repeats (complementary to telomere DNA repeats). These two motifs (TR template and specific USE sequence) then served to identify TRs in selected species of Viridiplantae and other selected species of the phylogenetic megagroup Diaphoretickes (including e.g., Rhodophytes, Ciliates or Stramenopiles). Homologous TRs were then used to build covariance models to identify TRs in more distant species. Example TRs were confirmed experimentally (Fajkus et al. 2021). Together, our results elucidated evolution of the earliest eukaryotic TRs, demonstrated the common origin of TRs across Diaphoretickes, and explained evolutionary transitions in telomere repeats. Our current studies continue to investigate the evolution of telomerases even beyond Diaphoretickes megagroup. In parallel, we characterise TR secondary structure and the whole telomerase ribonucleoprotein composition, structure and assembly. We believe the studies can address the molecular principle of reversible telomerase regulation.
Acknowledgements: I would like to thank Petr Fajkus, Vratislav Peška and the other members of my research group for their significant contribution to these results. The research was supported by ERDF project SYMBIT, reg. no. CZ.02.1.01/0.0/0.0/15.003/0000477, Czech Science Foundation (20-01331X) and ERDF project SINGING Plant, reg. no. CZ.02.01/0.0/0.0/16_026/0008446.
References:
Cifuentes-Rojas, C., Kannan, K., Tseng, L. and Shippen, D.E. (2011) Two RNA subunits and POT1a are components of Arabidopsis telomerase. P Natl Acad Sci USA, 108, 73-78.
Fajkus, J., Fulneckova, J., Hulanova, M., Berkova, K., Riha, K. and Matyasek, R. (1998) Plant cells express telomerase activity upon transfer to callus culture, without extensively changing telomere lengths. Mol Gen Genet, 260, 470-474.
Fajkus, J., Kovarik, A. and Kralovics, R. (1996) Telomerase activity in plant cells. Febs Lett, 391, 307-309.
Fajkus, J., Kovarik, A., Kralovics, R. and Bezdek, M. (1995) Organization of Telomeric and Subtelomeric Chromatin in the Higher-Plant Nicotiana-Tabacum. Mol Gen Genet, 247, 633-638.
Fajkus, P., Kilar, A., Nelson, A.D.L., Hola, M., Peska, V., Goffova, I., Fojtova, M., Zachova, D., Fulneckova, J. and Fajkus, J. (2021) Evolution of plant telomerase RNAs: farther to the past, deeper to the roots. Nucleic Acids Res, 49, 7680-7694.
Fajkus, P., Peska, V., Sitova, Z., Fulneckova, J., Dvorackova, M., Gogela, R., Sykorova, E., Hapala, J. and Fajkus, J. (2016) Allium telomeres unmasked: the unusual telomeric sequence (CTCGGTTATGGG)(n) is synthesized by telomerase. Plant J, 85, 337-347.
Fajkus, P., Peska, V., Zavodnik, M., Fojtova, M., Fulneckova, J., Dobias, S., Kilar, A., Dvorackova, M., Zachova, D., Necasova, I., Sims, J., Sykorova, E. and Fajkus, J. (2019) Telomerase RNAs in land plants. Nucleic Acids Res, 47, 9842-9856.
Fuchs, J., Brandes, A. and Schubert, I. (1995) Telomere Sequence Localization and Karyotype Evolution in Higher-Plants. Plant Syst Evol, 196, 227-241.
Pich, U., Fuchs, J. and Schubert, I. (1996) How do Alliaceae stabilize their chromosome ends in the absence of TTTAGGG sequences? Chromosome Res, 4, 207-213.
Riha, K., Fajkus, J., Siroky, J. and Vyskot, B. (1998) Developmental control of telomere lengths and telomerase activity in plants. Plant Cell, 10, 1691-1698.
Sykorova, E., Lim, K.Y., Kunicka, Z., Chase, M.W., Bennett, M.D., Fajkus, J. and Leitch, A.R. (2003) Telomere variability in the monocotyledonous plant order Asparagales. P Roy Soc B-Biol Sci, 270, 1893-1904.