CGM - Department Genes Expression
Structure, functions and evolution of catalytic RNA
Group leader: François MICHEL
Last update: 02-Sept-2011
Group members
Maria Costa, Chargé de Recherche, CNRS
Jean-Luc Ferat, Maître de Conférence, UVSQ
François Michel, Directeur de Recherche, CNRS
Dario Monachello, Ingénieur de Recherche
Boubekeur Saïfi, Doctorant
Our address
CNRS - Centre de Génétique Moléculaire
Avenue de la Terrasse - Bât. 26
91198 GIF-SUR-YVETTE Cedex
FRANCE
Phone: 01 69 82 31 88
Telecopy: 01 69 82 43 86
Structure and in vitro activity of group II introns
M. Costa, C.-F. Li, F. Michel
Group II introns, which are widely believed to be the progenitors of the nuclear splicing machinery (the spliceosome) and its substrates, comprise a large ribozyme (catalytic RNA) (Fig. 1) and the coding sequence of a reverse transcriptase. Group II introns are found in mitochondrial, chloroplast and bacterial genomes and a majority of them behave as retrotransposons. Our main research themes are the three-dimensional structure of group II ribozymes and their interactions with the intron-encoded protein.
Figure 1: Secondary structure and tertiary interactions of a group II ribozyme (Pl.LSU/2 from the large ribosomal RNA precursor of Pylaiella littoralis). We are using this molecule as a model system for in vitro studies. The secondary structure is organized into six 'domains' (subtrees I to VI). EBS1-IBS1, EBS2-IBS2, EBS3-IBS3 are interactions between the intron and exons; tertiary interactions within the intron are designated by greek letters. The asterisk shows the location of the adenosine in dVI whose 2'-OH group attacks the phosphodiester bond between the intron and 5' exon during the first transesterification reaction of the splicing process. The terminal loop of dIV (1846 nucleotides) includes the coding sequence of a protein (with reverse transcriptase and endonuclease domains) which is involved in intron mobility. Colored sections are the ones that have been modeled (see Fig. 2).
We have combined computer analyses of sequence alignments, nucleotide substitution experiments, footprinting, in vitro selection (SELEX) and kinetic analyses of reactions in order to identify tertiary interactions and eventually be able to generate three-dimensional models of the ribozyme architecture. An example is provided by the model in Fig. 2, which was built in collaboration with Eric Westhof (IBMC, Strasbourg) and includes part of the complex formed by the two exons and two of the six structural domains of the ribozyme (Costa et al., 2000).
Figure 2: Three-dimensional model of the complex formed by the ligated exons (dark grey), domain V and part of domain I (Costa et coll., 2000).
The crystal structure of part of a group II intron was recently solved (reviewed in Michel et al. (2009) Trends Biochem. Sci., 34, 189-199), so that the model in Fig. 2 is now known to have been essentially correct. We are currently attempting to add the missing sections, precise the role of ions and characterize the conformational rearrangements undergone by the ribozyme and its exon substrates during the splicing process. These data are essential in order eventually to understand at the atomic level the mechanics of transposition by reverse splicing and reverse transcription – a process that is peculiar to group II introns.
Bacterial group II introns
J.-L. Ferat, B. Saïfi
Large-scale sequencing of bacterial genomes has changed our perception of the biology of group II introns. The sequences of some 40 group II introns from bacteria and archaebacteria have already been published (some strains carry up to several dozen copies of the same intron) and it is becoming clear that the diversity of group II molecules is significantly greater in the bacterial world than within organelles. We are particularly interested in the few introns that happen to interrupt important genes, as well as in a subclass which is widespread in eubacteria and whose members are notable for targeting transcription terminators as well as showing important differences in sequence and structure from the rest of group II molecules (Granlund et al., 2001).
We are looking for host-encoded factors potentially involved in splicing and/or transposition and are attempting to answer questions such as "how are transcription terminators, rather than a specific sequence, recognized by the complex between the ribozyme and intron-encoded protein ?" or "what might be the impact of those introns on the evolution of host genomes ?".
Publications depuis 1993
Saifi, B., Ferat, J.-L. (2012) Replication Fork Reactivation in a dnaC2 Mutant at Non-Permissive Temperature in Escherichia coli. PLoS One, 7 (3) e33613.
Li, C. F., Costa, M., Bassi, G., Lai, YK., Michel, F. (2011) Recurrent insertion of 5'-terminal nucleotides and loss of the branchpoint motif in lineages of group II introns inserted in mitochondrial preribosomal RNAs. RNA, 17 (7)1321-35.
Li, C. F., Costa, M., Michel, F. (2011) Linking the branchpoint helix to a newly found receptor allows lariat formation by a group II intron. EMBO J, 30 (15) 3040-51.
Mullineux, ST., Costa, M., Bassi, GS., Michel, F., Hausner, G. (2010) A group II intron encodes a functional LAGLIDADG homing endonuclease and self-splices under moderate temperature and ionic conditions. RNA, 16 (9) :1818-31.
Michel, F., Costa, M., Westhof, E. (2009) The ribozyme core of group II introns: a structure in want of partners. Trends Biochem Sci, 34 (4) 189-99.
James, L. and Sargueil, B. (2008) RNA secondary structure of the feline immunodeficiency virus 5'UTR and Gag coding region. Nucleic Acids Res, 36 (14) 4653-6.
Michel, F., Costa, M., Doucet, A. J. and Ferat, J.-L. (2007) Specialized lineages of bacterial group II introns. Biochimie, 89 (4) 542-53.
Balvay, L., Lopez Lastra, M., Sargueil, B., Darlix, J. and Ohlmann, T. (2007) Translational control of retroviruses. Nature Reviews Microbiology 5128-140.
Brezellec, P., Hoebeke, M., Hiet, M. S., Pasek, S. and Ferat, J.-L. (2006) DomainSieve: a protein domain-based screen that led to the identification of dam-associated genes with potential link to DNA maintenance. Bioinformatics, 22 (16) 1935-1941.
Weill, L. (2006) Etude de la fonction d'ARN structurés : Recherche d'un ribozyme de fonction inédite et initiation de la traduction de l'ARN génomique d'HIV-2. Thèse, Paris-Sud 11 Orsay.
Costa, M., Michel, F., Molina-Sanchez, M., Martinez-Abarca, F. and Toro, N. (2006) An alternative intron-exon pairing scheme implied by unexpected in vitro activities of group II intron RmInt1 from Sinorhizobium meliloti. Biochimie, 88 (6) 711-7.
Costa, M., Michel, F. and Toro, N. (2006) Potential for alternative intron-exon pairings in group II intron RmInt1 from Sinorhizobium meliloti and its relatives. RNA, 12 (3) 338-41.
Herbreteau, C., Weill, L., Decimo, D., Prévôt, D., Darlix, J., Sargueil, B., and Ohlman, T. (2005). HIV-2 genomic RNA contains a novel type of IRES located downstream to its initiation codon. Mol Struct Biol 12 (11) 1001-7
Longo, A., Leonard, C., Bassi, G., Berndt, D., Krahn, J., Hall, T., and Weeks, K. (2005). Evolution from DNA to RNA recognition by the bI3 LAGLIDADG maturase. Nat Struct Mol Biol 12, 779-787.
Sargueil, B. (2004) Etude de la structure fonctionnelle d'ARN "actifs", HDR, Université Paris-Sud 11.
Weill, L., Louis, D., and Sargueil, B. (2004). Selection and evolution of NTP specific aptamers. Nucleic Acids Research 32, 5045-5050
Ferat, J., Le Gouar, M., and Michel, F. (2003). A group II intron has invaded the genus Azotobacter and is inserted within the termination codon of the essential groEL gene. Mol Microbiol 49, 1407-1423.
Michel, F. (2003). Some questions about RNA folding. In Folding and Self-Assembly of Biological Macromolecules, Compte-Rendus d'un colloque de l'IHES, (Singapour, World Scientific).
Sargueil, B., Hampel, K., Lambert, D., and Burke, J. (2003). In vitro selection of second site revertants analysis of the hairpin ribozyme active site. J Biol Chem 278, 52783-52791.
Granlund, M, Michel, F, Norgren, M. (2001) Mutually Exclusive Distribution of IS1548 and GBSi1, an Active Group II Intron Identified in Human Isolates of Group B Streptococci. J. Bacteriol. 183, 2560-2569 Abstract Text
Liu, Z. R., Sargueil, B., and Smith, C. W. (2000) RNA-protein methylen blue-mediated photo-cross-linking. Methods in Enzymology 318, 22-33
Sargueil B., McKenna JK., and Burke J.M. (2000) Analysis of the functional role of a GoA sheared base pair by in vitro genetics. J. Biol. Chem. 275, 32157-32166 Abstract Text
Costa, M., Michel, F., Westhof, E. (2000) A three-dimensional perspective on exon binding by a group II self-splicing intron. EMBO J. 19, 5007-5018 Abstract Text
Michel, F., Costa, M., Massire, C., Westhof, E. (2000) Modelling RNA tertiary structure from patterns of sequence variation. Methods in Enzymology 317 491-510
Michel, F. (2000) Vers une utilisation thérapeutique des introns de groupe II ? Médecine sciences 16 1430-1431
Costa, M., Michel, F. (1999) Tight binding of the 5' exon to domain I of a group II self-splicing intron requires completion of the intron active site. EMBO J. 18, 1025-1037 Abstract Text
Aubert, J., Legal, L., Descimon, H., Michel, F. (1999) Molecular phylogeny of Swallowtail butterflies of the tribe Papilionini (Papilionidae, Lepidoptera). Molec. Phyl. Evol. 12, 156-167Abstract Text
Brion, P., Michel, F., Schroeder, R., Westhof, E. (1999) Analysis of the cooperative thermal unfolding of the td intron of bacteriophage T4 Nucleic Acids Res. 27, 2494-2502 Abstract Text
Brion, P., Schroeder, R., Michel, F., Westhof, E. (1999) Influence of specific mutations on the thermal stability of the td group I intron in vitro and on its splicing efficiency in vivo: a comparative study RNA 5, 947-958 Abstract Text
Einvik, C., Nielsen, H., Westhof, E., Michel, F., Johansen, S. Group I-like ribozymes with a novel core organization perform obligate sequential hydrolytic cleavages at two processing sites. RNA 4, 530-541 (1998) Abstract Text
Costa, M., Christian, E.L., Michel, F. (1998) Differential chemical probing of a group II self-splicing intron identifies bases involved in tertiary interactions and supports an alternative secondary structure model of domain V. RNA 4, 1055-1068 Abstract Text
Westhof, E., Michel, F. (1998) Ribozyme architectural diversity made visible. Science 282, 251-252
Michel, F., Costa, M. (1998) Inferring RNA structure by phylogenetic and genetic analyses. in 'RNA Structure and Function', Cold Spring Harbor Laboratory Press, R. Simons and M. Grunberg-Manago eds., pp. 175-202
Costa, M., Fontaine, J.-M., Loiseaux-de Goër, S., Michel, F. (1997) A group II self-splicing intron from the brown alga Pylaiella littoralis is active at unusually low magnesium concentrations and forms populations of molecules with a uniform conformation. J. Mol. Biol. 274, 353-364 Abstract Text
Costa, M., Dème, E., Jacquier, A., Michel, F. (1997) Multiple tertiary interactions involving domain II of group II self-splicing introns. J. Mol. Biol. 267, 520-536 Abstract Text
Robineau, S., Bergantino, E., Carignani, G., Michel, F., Netter, P. (1997) Suppressors of cis-acting splicing-deficient mutations that affect the ribozyme core of a group II intron. J. Mol. Biol. 267, 537-547Abstract Text
Costa, M., Michel, F. (1997) Rules for RNA recognition of GNRA tetraloops deduced by in vitro selection: comparison with in vivo evolution. EMBO J. 11, 3289-3302 Abstract Text
Aubert, J., Barascud, B., Descimon, H., Michel, F. (1997) Ecology and genetics of interspecific hybridization in the swallowtails, Papilio hospiton Géné and Papilio machaon L., in Corsica (Lepidoptera: Papilionidae). Biological Journal of the Linnean Society 60, 467-492 Abstract Text
Aubert, J., Barascud, B., Descimon, H., Michel, F. Systématique moléculaire des Argynnes. C.R. Acad. Sci. Paris, Sciences de la vie 319, 647-651 (1996)
Aubert, J., Descimon, H., Michel, F. Population biology and conservation of the Corsican Swallowtail butterfly Papilio hospiton. Géné. Biological Conservation 78, 247-255 (1996)
Michel, F., Westhof, E. Visualizing the logic behind RNA self-assembly. Science 273, 1676-1677 (1996)
Jaeger, L., Westhof, E., Michel, F. (1996) Function of a pseudoknot in the suppression of an alternative splicing event in a group I intron. Biochimie 78, 466-47.
Lehnert, V., Jaeger, L., Michel, F., Westhof, E. (1996) New loop-loop tertiary interactions in self-splicing introns of subgroup IC and ID: a complete 3D model of the Tetrahymena thermophila ribozyme. Chemistry & Biology 3, 993-1009.
Jaeger, L., Michel, F., Westhof, E. (1996) The structure of group I ribozymes. In 'Nucleic Acids and Molecular Biology', vol. 10, F. Eckstein & D.M.J. Lilley Eds, Springer Verlag Berlin, pp. 33-51.
Costa, M., Michel, F. (1995) Frequent use of the same tertiary motif by self-folding RNAs. EMBO J. 14, 1276-1285 Abstract
Michel, F., Ferat, J.-L. (1995) Structure and activities of group II introns. Annual Review of Biochemistry 64, 435-461.
Michel, F., Westhof, E. (1994) Slippery substrates. Nature Structural Biology 1, 5-7 .
Lisacek, F., Diaz, Y., Michel, F. (1994) Automatic identification of group I intron cores in genomic DNA sequences. J. Mol. Biol. 235, 1206-1217 . Abstract Text
Ferat, J.-L., Le Gouar, M., Michel, F. (1994) Multiple group II self-splicing introns in mobile DNA from Escherichia coli. Comptes Rendus Acad. Sci. Paris 317, 141-148.
Jaeger, L., Michel, F., Westhof, E. (1994) Involvement of a GNRA tetraloop in long-range RNA tertiary interactions. J. Mol. Biol. 236, 1271-1276 .
Westhof, E., Michel, F. (1994) Prediction and experimental investigation of RNA secondary and tertiary foldings. In 'RNA-Protein Interactions: Frontiers in Molecular Biology', K. Nagai & I.W. Mattaj Eds, IRL Press at Oxford University Press.
Tian, G.-L., Michel, F., Macadre, C., Lazowska, J. (1993) Sequence of the mitochondrial gene encoding subunit I of cytochrome oxidase in Saccharomyces douglasii. Gene 124, 153-163.
Westhof, E., Michel, F. (1993) Model of the catalytic core of group I introns. Structure: Introductory issue, viii-ix.
Ohta, E., Oda, K., Yamato, K., Nakamura, Y., Takemura, M., Nozato, N., Akashi, K., Ohyama, K., Michel, F. (1993) Group I introns in the liverwort mitochondrial genome: the gene coding for subunit 1 of cytochrome oxidase shares five intron positions with its fungal counterparts. Nucleic Acids Res. 21, 1297-1305. Abstract
Ferat, J.-L., Michel, F. (1993) Group II self-splicing introns in bacteria. Nature 364, 358-361.
Haouazine, N., Takvorian, A., Jubier, M.-F., Michel, F., Lejeune, B. (1993) nad6 gene and nad1 exon d are co-transcribed in wheat mitochondria. Current Genetics 24, 533-538.
Jaeger, L., Westhof, E., Michel, F. (1993) Monitoring of the cooperative unfolding of the sf the sunY group I intron of bacteriophage T4. The active form of the sunY ribozyme core is stabilized by multiple interactions with 3' terminal intron components. J. Mol. Biol. 234, 331-346. Abstract Text
Goldschmidt-Clermont, M., Choquet, Y., Girard-Bascou, J., Michel, F., Rochaix, J.-D. (1993) Post-transcriptional Control of Chloroplast Gene Expression in Chlamydomonas: the Case of psaA Trans-splicing. In "Plant Mitochondria", A. Brennicke, U. Kück Eds, VCH Weinheim, pp. 233-240 .
