Luc Jaeger - Academia.edu (original) (raw)

Papers by Luc Jaeger

Current Biology, 1992

RNA pseudoknots result from Watson-Crick base pairing involving a stretch of bases located betwee... more RNA pseudoknots result from Watson-Crick base pairing involving a stretch of bases located between paired strands and a distal single-stranded region. Recently, significant advances in our understanding of their structural and functional aspects have been accomplished. At the structural level, modelling and NMR studies have shown that a defined subset of pseudoknots may be considered as tertiary motifs in RNA foldings. At the functional level, there is evidence that the realm of functions encompassed by RNA pseudoknots extends from the control of translation in prokaryotes, retroviruses and coronaviruses to the control of catalytic activity in ribozymes and the control of replication in some plant viruses.

Nature Communications, 2020

5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5... more 5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5S rRNA and the reasons for its evolutionary preservation as an independent molecule remain unclear. Here we used ribosome engineering to investigate whether 5S rRNA autonomy is critical for ribosome function and cell survival. By linking circularly permutated 5S rRNA with 23S rRNA we generated a bacterial strain devoid of free 5S rRNA. Viability of the engineered cells demonstrates that autonomous 5S rRNA is dispensable for cell growth under standard conditions and is unlikely to have essential functions outside the ribosome. The fully assembled ribosomes carrying 23S-5S rRNA are highly active in translation. However, the engineered cells accumulate aberrant 50S subunits unable to form stable 70S ribosomes. Cryo-EM analysis revealed a malformed peptidyl transferase center in the misassembled 50S subunits. Our results argue that the autonomy of 5S rRNA is preserved due to its role in ribo...

F1000 - Post-publication peer review of the biomedical literature, 2013

The pentatricopeptide repeat (PPR) is a helical repeat motif found in an exceptionally large fami... more The pentatricopeptide repeat (PPR) is a helical repeat motif found in an exceptionally large family of RNA-binding proteins that functions in mitochondrial and chloroplast gene expression. PPR proteins harbor between 2 and 30 repeats and typically bind single-stranded RNA in a sequence-specific fashion. However, the basis for sequence-specific RNA recognition by PPR tracts has been unknown. We used computational methods to infer a code for nucleotide recognition involving two amino acids in each repeat, and we validated this model by recoding a PPR protein to bind novel RNA sequences in vitro. Our results show that PPR tracts bind RNA via a modular recognition mechanism that differs from previously described RNA-protein recognition modes and that underpins a natural library of specific protein/RNA partners of unprecedented size and diversity. These findings provide a significant step toward the prediction of native binding sites of the enormous number of PPR proteins found in nature. Furthermore, the extraordinary evolutionary plasticity of the PPR family suggests that the PPR scaffold will be particularly amenable to redesign for new sequence specificities and functions.

F1000 - Post-publication peer review of the biomedical literature, 2013

F1000 - Post-publication peer review of the biomedical literature, 2013

Angewandte Chemie International Edition, 2000

Journal of Molecular Biology, 1993

Journal of Molecular Biology, 1994

Terminal loops with a GNRA consensus sequence are widespread in RNA. It has been suggested that t... more Terminal loops with a GNRA consensus sequence are widespread in RNA. It has been suggested that these loops act as "anchors" during tertiary folding, by interacting in a sequence-specific way with helices at distant locations along the molecule. We now show that a GUGA loop changes state upon disruption of the tertiary architecture of a self-splicing group I intron. Successful replacement of the postulated loop-helix contact by classical base-pairing points to binding of the loop into the shallow (minor) groove of the helix, as also indicated by partial restoration of ribozyme stability upon a specific double nucleotide substitution.

Nature Chemistry, 2010

Supra-molecular assembly is a powerful strategy used by nature for building nano-scale architectu... more Supra-molecular assembly is a powerful strategy used by nature for building nano-scale architectures with predefined sizes and shapes. Numerous challenges remain however to be solved in order to demonstrate precise control over the synthesis, folding and assembly of rationally designed three-dimensional (3D) nano-objects made of RNA. Using the transfer RNA molecule as a structural building block, we report the design, efficient synthesis and structural characterization of stable, modular 3D particles adopting the polyhedral geometry of a non-uniform square antiprism. The spatial control within the final architecture allows precise positioning and encapsulation of proteins. This work demonstrates that a remarkable degree of structural control can be achieved with RNA structural motifs to build thermostable 3D nano-architectures that do not rely on helix bundles or tensegrity. RNA 3D particles can potentially be used as carriers or scaffolds in nano-medicine and synthetic biology.

Nucleic Acids and Molecular Biology, 1996

Group I introns are large catalytic RNAs which, like the group II introns and the RNA component o... more Group I introns are large catalytic RNAs which, like the group II introns and the RNA component of bacterial RNase P, can function alone in salt solutions (Cech 1993). In vitro, these ribozymes carry out self-splicing via two consecutive transesterification reactions, leading to their own excision from a pre-messenger RNA and the ligation of the flanking exon sequences, generally, in the absence of protein (Cech 1990). This relatively complex process requires the folding of the intron into a defined tertiary structure that allows the recognition of the substrates and catalysis. Thus, a good understanding of the series of events occurring between the synthesis of the ribozyme and the self-splicing reaction requires answers to the following questions. (1) What does a group I intron look like in three-dimensional space? (2) How does it reach its folded structure and recognize its different substrates for catalysis?

F1000 - Post-publication peer review of the biomedical literature, 2013

Analysis of the 2.4-Å resolution crystal structure of the large ribosomal subunit from Haloarcula... more Analysis of the 2.4-Å resolution crystal structure of the large ribosomal subunit from Haloarcula marismortui reveals the existence of an abundant and ubiquitous structural motif that stabilizes RNA tertiary and quaternary structures. This motif is termed the A-minor motif, because it involves the insertion of the smooth, minor groove edges of adenines into the minor groove of neighboring helices, preferentially at C-G base pairs, where they form hydrogen bonds with one or both of the 2 OHs of those pairs. A-minor motifs stabilize contacts between RNA helices, interactions between loops and helices, and the conformations of junctions and tight turns. The interactions between the 3 terminal adenine of tRNAs bound in either the A site or the P site with 23S rRNA are examples of functionally significant A-minor interactions. The A-minor motif is by far the most abundant tertiary structure interaction in the large ribosomal subunit; 186 adenines in 23S and 5S rRNA participate, 68 of which are conserved. It may prove to be the universally most important long-range interaction in large RNA structures.

F1000 - Post-publication peer review of the biomedical literature, 2013

Motivation: Systematic Evolution of Ligands by EXponential Enrichment (SELEX) represents a state-... more Motivation: Systematic Evolution of Ligands by EXponential Enrichment (SELEX) represents a state-of-the-art technology to isolate single-stranded (ribo)nucleic acid fragments, named aptamers, which bind to a molecule (or molecules) of interest via specific structural regions induced by their sequence-dependent fold. This powerful method has applications in designing protein inhibitors, molecular detection systems, therapeutic drugs and antibody replacement among others. However, full understanding and consequently optimal utilization of the process has lagged behind its wide application due to the lack of dedicated computational approaches. At the same time, the combination of SELEX with novel sequencing technologies is beginning to provide the data that will allow the examination of a variety of properties of the selection process. Results: To close this gap we developed, Aptamotif, a computational method for the identification of sequence-structure motifs in SELEXderived aptamers. To increase the chances of identifying functional motifs, Aptamotif uses an ensemble-based approach. We validated the method using two published aptamer datasets containing experimentally determined motifs of increasing complexity. We were able to recreate the author's findings to a high degree, thus proving the capability of our approach to identify binding motifs in SELEX data. Additionally, using our new experimental dataset, we illustrate the application of Aptamotif to elucidate several properties of the selection process.

F1000 - Post-publication peer review of the biomedical literature, 2013

Biophysical Journal, 2014

Advances in synthetic biology are transforming our ability to design and build synthetic biologic... more Advances in synthetic biology are transforming our ability to design and build synthetic biological systems. While progress has been made in the design of complex genetic circuits, capabilities for constructing large genetic systems currently surpass our ability to design such systems. This growing 'design gap' has highlighted the need to develop methods that support the generation of new functional biological components and scalable design strategies for complex genetic circuits that will lay the foundation for integrated biological devices and systems. As examples of functional RNA molecules playing key roles in the behavior of natural biological systems have grown over the past decade, there has been growing interest in the design and implementation of synthetic counterparts. Researchers are taking advantage of the relative ease with which RNA molecules can be modeled and designed to engineer functional RNA molecules that act as diverse components including sensors, regulators, controllers (ligand-responsive RNA regulators), and scaffolds. These synthetic regulatory RNAs are providing new tools for temporal and spatial control in biological systems. I will describe recent advances in the design of RNA controllers and in addressing challenges in their implementation as user-programmed cellular control systems. In particular, I will discuss how the application of synthetic RNA controllers in biological pathways is leading to the elucidation of integrated systems design strategies and new capabilities for programming genetic systems.

Science Advances, 2018

We report a new algorithm and a battery of blind challenges for the prediction of complex RNA str... more We report a new algorithm and a battery of blind challenges for the prediction of complex RNA structures at atomic accuracy.

RNA, 2004

It is theoretically possible to obtain a catalytic site of an artificial ribozyme from a random s... more It is theoretically possible to obtain a catalytic site of an artificial ribozyme from a random sequence consisting of a limited numbers of nucleotides. However, this strategy has been inadequately explored. Here, we report an in vitro selection technique that exploits modular construction of a structurally constrained RNA to acquire a catalytic site for RNA ligation from a short random sequence. To practice the selection, a sequence of 30 nucleotides was located close to the putative reaction site in a derivative of a naturally occurring self-folding RNA whose crystal structure is known. RNAs whose activity depended on the starting three-dimensional structure were selected with 3′–5′ ligation specificity, indicating that the strategy can be used to acquire a variety of catalytic sites and other functional RNA modules.

Journal of the American Chemical Society, 2010

The structure of a DNA-dimethyldidodecylammonium bromide (DDAB) film was recently described to un... more The structure of a DNA-dimethyldidodecylammonium bromide (DDAB) film was recently described to undergo a distinctive transition in response to the water content in the surrounding environment. 1 The existence, preparation, and basic properties of DNA-surfactant films have been known in the literature for some time.2 , 3 Here, we describe the structural response of DNA-DDAB films to environmental changes, particularly temperature and humidity, in greater detail revealing new structural states. We can direct the lamellar structure of the film into three distinct states-doublestranded DNA (dsDNA) paired with an interdigitated bilayer of DDAB (bDDAB), single-stranded DNA (ssDNA) with monolayer of DDAB (mDDAB), and ssDNA with bDDAB. Both temperature and humidity cause the molecules composing the lamellar structure to change reversibly from ssDNA to dsDNA and/or from mDDAB to bDDAB. We found that the structural transition from dsDNA to ssDNA and bDDAB to mDDAB is concerted and follows apparent first order kinetics. We also found that the double-stranded conformation of DNA in the film can be stabilized with the inclusion of cholesterol even while the DDAB in the film is able to form either a monolayer or bilayer depending on the environmental conditions. Films treated with ethidium bromide prompt switching of dsDNA to ssDNA before bDDAB transitions to mDDAB. Swelling experiments have determined that there is a direct proportionality between the macroscopic increase in volume and the nanoscopic increase in lamellar spacing when a film is allowed to swell in water. Finally, experiments with phosphatebuffered saline (PBS) indicate that the films can disassemble in a simulated biological environment due to screening of their charges by buffer salt. We conclude that the structure of DNA in the film depends on the water content (as measured by the relative humidity) and temperature of the environment, while the state of DDAB depends essentially only on the water content. The structure of the film is quite flexible, and can be altered by changing environmental conditions as well as the chemical ingredients. These films will have useful, new applications as responsive materials, e.g. in drug and gene delivery.

Genes & Development, 1992

Over 1000 nucleotides may separate the ribozyme core of some group I introns from their 3' sp... more Over 1000 nucleotides may separate the ribozyme core of some group I introns from their 3' splice junctions. Using the sunY intron of bacteriophage T4 as a model system, we have investigated the mechanisms by which proximal splicing events are suppressed in vitro, as well as in vivo. Exon ligation as well as cleavage at the 5' splice site are shown to require long-range pairing between one of the peripheral components of the ribozyme core and some of the nucleotides preceding the authentic 3' splice junction. Consistent with our three-dimensional modeling of the entire sunY ribozyme, we propose that this novel interaction is necessary to drive 5' exon-core transcripts into an active conformation. A requirement for additional stabilizing interactions, either RNA-based or mediated by proteins, appears to be a general feature of group I self-splicing. A role for these interactions in mediating putative alternative splicing events is discussed.

Current Biology, 1992

RNA pseudoknots result from Watson-Crick base pairing involving a stretch of bases located betwee... more RNA pseudoknots result from Watson-Crick base pairing involving a stretch of bases located between paired strands and a distal single-stranded region. Recently, significant advances in our understanding of their structural and functional aspects have been accomplished. At the structural level, modelling and NMR studies have shown that a defined subset of pseudoknots may be considered as tertiary motifs in RNA foldings. At the functional level, there is evidence that the realm of functions encompassed by RNA pseudoknots extends from the control of translation in prokaryotes, retroviruses and coronaviruses to the control of catalytic activity in ribozymes and the control of replication in some plant viruses.

Nature Communications, 2020

5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5... more 5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5S rRNA and the reasons for its evolutionary preservation as an independent molecule remain unclear. Here we used ribosome engineering to investigate whether 5S rRNA autonomy is critical for ribosome function and cell survival. By linking circularly permutated 5S rRNA with 23S rRNA we generated a bacterial strain devoid of free 5S rRNA. Viability of the engineered cells demonstrates that autonomous 5S rRNA is dispensable for cell growth under standard conditions and is unlikely to have essential functions outside the ribosome. The fully assembled ribosomes carrying 23S-5S rRNA are highly active in translation. However, the engineered cells accumulate aberrant 50S subunits unable to form stable 70S ribosomes. Cryo-EM analysis revealed a malformed peptidyl transferase center in the misassembled 50S subunits. Our results argue that the autonomy of 5S rRNA is preserved due to its role in ribo...

F1000 - Post-publication peer review of the biomedical literature, 2013

The pentatricopeptide repeat (PPR) is a helical repeat motif found in an exceptionally large fami... more The pentatricopeptide repeat (PPR) is a helical repeat motif found in an exceptionally large family of RNA-binding proteins that functions in mitochondrial and chloroplast gene expression. PPR proteins harbor between 2 and 30 repeats and typically bind single-stranded RNA in a sequence-specific fashion. However, the basis for sequence-specific RNA recognition by PPR tracts has been unknown. We used computational methods to infer a code for nucleotide recognition involving two amino acids in each repeat, and we validated this model by recoding a PPR protein to bind novel RNA sequences in vitro. Our results show that PPR tracts bind RNA via a modular recognition mechanism that differs from previously described RNA-protein recognition modes and that underpins a natural library of specific protein/RNA partners of unprecedented size and diversity. These findings provide a significant step toward the prediction of native binding sites of the enormous number of PPR proteins found in nature. Furthermore, the extraordinary evolutionary plasticity of the PPR family suggests that the PPR scaffold will be particularly amenable to redesign for new sequence specificities and functions.

F1000 - Post-publication peer review of the biomedical literature, 2013

F1000 - Post-publication peer review of the biomedical literature, 2013

Angewandte Chemie International Edition, 2000

Journal of Molecular Biology, 1993

Journal of Molecular Biology, 1994

Terminal loops with a GNRA consensus sequence are widespread in RNA. It has been suggested that t... more Terminal loops with a GNRA consensus sequence are widespread in RNA. It has been suggested that these loops act as "anchors" during tertiary folding, by interacting in a sequence-specific way with helices at distant locations along the molecule. We now show that a GUGA loop changes state upon disruption of the tertiary architecture of a self-splicing group I intron. Successful replacement of the postulated loop-helix contact by classical base-pairing points to binding of the loop into the shallow (minor) groove of the helix, as also indicated by partial restoration of ribozyme stability upon a specific double nucleotide substitution.

Nature Chemistry, 2010

Supra-molecular assembly is a powerful strategy used by nature for building nano-scale architectu... more Supra-molecular assembly is a powerful strategy used by nature for building nano-scale architectures with predefined sizes and shapes. Numerous challenges remain however to be solved in order to demonstrate precise control over the synthesis, folding and assembly of rationally designed three-dimensional (3D) nano-objects made of RNA. Using the transfer RNA molecule as a structural building block, we report the design, efficient synthesis and structural characterization of stable, modular 3D particles adopting the polyhedral geometry of a non-uniform square antiprism. The spatial control within the final architecture allows precise positioning and encapsulation of proteins. This work demonstrates that a remarkable degree of structural control can be achieved with RNA structural motifs to build thermostable 3D nano-architectures that do not rely on helix bundles or tensegrity. RNA 3D particles can potentially be used as carriers or scaffolds in nano-medicine and synthetic biology.

Nucleic Acids and Molecular Biology, 1996

Group I introns are large catalytic RNAs which, like the group II introns and the RNA component o... more Group I introns are large catalytic RNAs which, like the group II introns and the RNA component of bacterial RNase P, can function alone in salt solutions (Cech 1993). In vitro, these ribozymes carry out self-splicing via two consecutive transesterification reactions, leading to their own excision from a pre-messenger RNA and the ligation of the flanking exon sequences, generally, in the absence of protein (Cech 1990). This relatively complex process requires the folding of the intron into a defined tertiary structure that allows the recognition of the substrates and catalysis. Thus, a good understanding of the series of events occurring between the synthesis of the ribozyme and the self-splicing reaction requires answers to the following questions. (1) What does a group I intron look like in three-dimensional space? (2) How does it reach its folded structure and recognize its different substrates for catalysis?

F1000 - Post-publication peer review of the biomedical literature, 2013

Analysis of the 2.4-Å resolution crystal structure of the large ribosomal subunit from Haloarcula... more Analysis of the 2.4-Å resolution crystal structure of the large ribosomal subunit from Haloarcula marismortui reveals the existence of an abundant and ubiquitous structural motif that stabilizes RNA tertiary and quaternary structures. This motif is termed the A-minor motif, because it involves the insertion of the smooth, minor groove edges of adenines into the minor groove of neighboring helices, preferentially at C-G base pairs, where they form hydrogen bonds with one or both of the 2 OHs of those pairs. A-minor motifs stabilize contacts between RNA helices, interactions between loops and helices, and the conformations of junctions and tight turns. The interactions between the 3 terminal adenine of tRNAs bound in either the A site or the P site with 23S rRNA are examples of functionally significant A-minor interactions. The A-minor motif is by far the most abundant tertiary structure interaction in the large ribosomal subunit; 186 adenines in 23S and 5S rRNA participate, 68 of which are conserved. It may prove to be the universally most important long-range interaction in large RNA structures.

F1000 - Post-publication peer review of the biomedical literature, 2013

Motivation: Systematic Evolution of Ligands by EXponential Enrichment (SELEX) represents a state-... more Motivation: Systematic Evolution of Ligands by EXponential Enrichment (SELEX) represents a state-of-the-art technology to isolate single-stranded (ribo)nucleic acid fragments, named aptamers, which bind to a molecule (or molecules) of interest via specific structural regions induced by their sequence-dependent fold. This powerful method has applications in designing protein inhibitors, molecular detection systems, therapeutic drugs and antibody replacement among others. However, full understanding and consequently optimal utilization of the process has lagged behind its wide application due to the lack of dedicated computational approaches. At the same time, the combination of SELEX with novel sequencing technologies is beginning to provide the data that will allow the examination of a variety of properties of the selection process. Results: To close this gap we developed, Aptamotif, a computational method for the identification of sequence-structure motifs in SELEXderived aptamers. To increase the chances of identifying functional motifs, Aptamotif uses an ensemble-based approach. We validated the method using two published aptamer datasets containing experimentally determined motifs of increasing complexity. We were able to recreate the author's findings to a high degree, thus proving the capability of our approach to identify binding motifs in SELEX data. Additionally, using our new experimental dataset, we illustrate the application of Aptamotif to elucidate several properties of the selection process.

F1000 - Post-publication peer review of the biomedical literature, 2013

Biophysical Journal, 2014

Advances in synthetic biology are transforming our ability to design and build synthetic biologic... more Advances in synthetic biology are transforming our ability to design and build synthetic biological systems. While progress has been made in the design of complex genetic circuits, capabilities for constructing large genetic systems currently surpass our ability to design such systems. This growing 'design gap' has highlighted the need to develop methods that support the generation of new functional biological components and scalable design strategies for complex genetic circuits that will lay the foundation for integrated biological devices and systems. As examples of functional RNA molecules playing key roles in the behavior of natural biological systems have grown over the past decade, there has been growing interest in the design and implementation of synthetic counterparts. Researchers are taking advantage of the relative ease with which RNA molecules can be modeled and designed to engineer functional RNA molecules that act as diverse components including sensors, regulators, controllers (ligand-responsive RNA regulators), and scaffolds. These synthetic regulatory RNAs are providing new tools for temporal and spatial control in biological systems. I will describe recent advances in the design of RNA controllers and in addressing challenges in their implementation as user-programmed cellular control systems. In particular, I will discuss how the application of synthetic RNA controllers in biological pathways is leading to the elucidation of integrated systems design strategies and new capabilities for programming genetic systems.

Science Advances, 2018

We report a new algorithm and a battery of blind challenges for the prediction of complex RNA str... more We report a new algorithm and a battery of blind challenges for the prediction of complex RNA structures at atomic accuracy.

RNA, 2004

It is theoretically possible to obtain a catalytic site of an artificial ribozyme from a random s... more It is theoretically possible to obtain a catalytic site of an artificial ribozyme from a random sequence consisting of a limited numbers of nucleotides. However, this strategy has been inadequately explored. Here, we report an in vitro selection technique that exploits modular construction of a structurally constrained RNA to acquire a catalytic site for RNA ligation from a short random sequence. To practice the selection, a sequence of 30 nucleotides was located close to the putative reaction site in a derivative of a naturally occurring self-folding RNA whose crystal structure is known. RNAs whose activity depended on the starting three-dimensional structure were selected with 3′–5′ ligation specificity, indicating that the strategy can be used to acquire a variety of catalytic sites and other functional RNA modules.

Journal of the American Chemical Society, 2010

The structure of a DNA-dimethyldidodecylammonium bromide (DDAB) film was recently described to un... more The structure of a DNA-dimethyldidodecylammonium bromide (DDAB) film was recently described to undergo a distinctive transition in response to the water content in the surrounding environment. 1 The existence, preparation, and basic properties of DNA-surfactant films have been known in the literature for some time.2 , 3 Here, we describe the structural response of DNA-DDAB films to environmental changes, particularly temperature and humidity, in greater detail revealing new structural states. We can direct the lamellar structure of the film into three distinct states-doublestranded DNA (dsDNA) paired with an interdigitated bilayer of DDAB (bDDAB), single-stranded DNA (ssDNA) with monolayer of DDAB (mDDAB), and ssDNA with bDDAB. Both temperature and humidity cause the molecules composing the lamellar structure to change reversibly from ssDNA to dsDNA and/or from mDDAB to bDDAB. We found that the structural transition from dsDNA to ssDNA and bDDAB to mDDAB is concerted and follows apparent first order kinetics. We also found that the double-stranded conformation of DNA in the film can be stabilized with the inclusion of cholesterol even while the DDAB in the film is able to form either a monolayer or bilayer depending on the environmental conditions. Films treated with ethidium bromide prompt switching of dsDNA to ssDNA before bDDAB transitions to mDDAB. Swelling experiments have determined that there is a direct proportionality between the macroscopic increase in volume and the nanoscopic increase in lamellar spacing when a film is allowed to swell in water. Finally, experiments with phosphatebuffered saline (PBS) indicate that the films can disassemble in a simulated biological environment due to screening of their charges by buffer salt. We conclude that the structure of DNA in the film depends on the water content (as measured by the relative humidity) and temperature of the environment, while the state of DDAB depends essentially only on the water content. The structure of the film is quite flexible, and can be altered by changing environmental conditions as well as the chemical ingredients. These films will have useful, new applications as responsive materials, e.g. in drug and gene delivery.

Genes & Development, 1992

Over 1000 nucleotides may separate the ribozyme core of some group I introns from their 3' sp... more Over 1000 nucleotides may separate the ribozyme core of some group I introns from their 3' splice junctions. Using the sunY intron of bacteriophage T4 as a model system, we have investigated the mechanisms by which proximal splicing events are suppressed in vitro, as well as in vivo. Exon ligation as well as cleavage at the 5' splice site are shown to require long-range pairing between one of the peripheral components of the ribozyme core and some of the nucleotides preceding the authentic 3' splice junction. Consistent with our three-dimensional modeling of the entire sunY ribozyme, we propose that this novel interaction is necessary to drive 5' exon-core transcripts into an active conformation. A requirement for additional stabilizing interactions, either RNA-based or mediated by proteins, appears to be a general feature of group I self-splicing. A role for these interactions in mediating putative alternative splicing events is discussed.