Assembly of the Archaeal Box C/D sRNP Can Occur via Alternative Pathways and Requires Temperature-facilitated sRNA Remodeling

Gagnon, Keith T.; Zhang, Xinxin; Agris, Paul F.; Maxwell, Elizabeth S.

doi:10.1016/j.jmb.2006.07.091

Cited by 22 publications

(36 citation statements)

References 58 publications

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“…Biological function of sRNA motion in specific 2 0 -O-methylation Gagnon et al 90 also confirmed the induced fitting mechanism of K-turn sRNA upon binding with L7Ae and Nop56/58. The assembly and sRNA's structure change are temperature-dependent, indicating the key role of the sRNA's flexibility in sRNP assembly.…”

Section: Comparison With Experiments and Previous MD Simulationsmentioning

confidence: 64%

Kink turn sRNA folding upon L7Ae binding using molecular dynamics simulations

Wei¹,

Yang²,

Yu³

et al. 2013

Phys. Chem. Chem. Phys.

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Section: Comparison With Experiments and Previous MD Simulationsmentioning

confidence: 64%

Kink turn sRNA folding upon L7Ae binding using molecular dynamics simulations

Wei¹,

Yang²,

Yu³

et al. 2013

Phys. Chem. Chem. Phys.

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“…Analysis of protein-protein and protein-RNA interactions by affinity copurification was performed as previously described Gagnon et al 2006;Zhang et al 2006). Briefly, His(6X)-tagged wild-type or mutant L7Ae was incubated at 70°C with untagged L7Ae wild-type or mutant protein with or without the indicated RNAs.…”

Section: Protein and Rnp Affinity Copurificationmentioning

confidence: 99%

“…In vitro 29-O-methylation assays for assembled box C/D sRNPs were carried out as previously described Gagnon et al 2006). Aliquots were removed at selected time points, TCAprecipitated, and washed, and incorporation of In vivo assessment of mutant M. jannaschii L7Ae, mouse 15.5kD, and yeast Snu13p function Yeast strain YGALSNU13 with SNU13 under the control of the GAL1 promoter was a gift from Raymond O'Keefe (Dobbyn and O'Keefe 2004).…”

Section: Protein and Rnp Affinity Copurificationmentioning

confidence: 99%

Signature amino acids enable the archaeal L7Ae box C/D RNP core protein to recognize and bind the K-loop RNA motif

Gagnon¹,

Zhang²,

Qu³

et al. 2009

RNA

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The archaeal L7Ae and eukaryotic 15.5kD protein homologs are members of the L7Ae/15.5kD protein family that characteristically recognize K-turn motifs found in both archaeal and eukaryotic RNAs. In Archaea, the L7Ae protein uniquely binds the K-loop motif found in box C/D and H/ACA sRNAs, whereas the eukaryotic 15.5kD homolog is unable to recognize this variant K-turn RNA. Comparative sequence and structural analyses, coupled with amino acid replacement experiments, have demonstrated that five amino acids enable the archaeal L7Ae core protein to recognize and bind the K-loop motif. These signature residues are highly conserved in the archaeal L7Ae and eukaryotic 15.5kD homologs, but differ between the two domains of life. Interestingly, loss of K-loop binding by archaeal L7Ae does not disrupt C9/D9 RNP formation or RNA-guided nucleotide modification. L7Ae is still incorporated into the C9/D9 RNP despite its inability to bind the K-loop, thus indicating the importance of protein-protein interactions for RNP assembly and function. Finally, these five signature amino acids are distinct for each of the L7Ae/L30 family members, suggesting an evolutionary continuum of these RNA-binding proteins for recognition of the various K-turn motifs contained in their cognate RNAs.

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“…The similarity between pre-tRNA Trp intron and its circularly permuted version was not unexpected, since the intron is normally spliced out as circular RNA and can be detected inside the cells (Salgia et al 2003), and both linear and circular versions of this intron can guide 29-O-methylation (Singh et al 2004). Gel-shift analyses have indicated that only one L7Ae binds to archaeal box C/D-guide RNAs at a lower protein:RNA ratio (Rashid et al 2003;Tran et al 2003;Gagnon et al 2006). However, it has been difficult to specifically assign this asymmetric binding of L7Ae to the box C/D or box C9/D9 motif.…”

Section: L7ae Induced Structural Rearrangements In Srnasmentioning

confidence: 99%

“…Binding of L7Ae to the K-turn changes the conformation of the box C/D motif, and this altered RNA conformation is then recognized by aNop5p (Rashid et al 2003). Structural probing of the sRNP complexes during assembly has shown that sRNA remodeling is critical for the assembly and functioning of these complexes (Gagnon et al 2006). Although three archaeal core proteins assemble symmetrically at the two box motifs of sRNA, the complex formed at one of the two motifs may have a preference for modification (Rashid et al 2003).…”

Section: Introductionmentioning

confidence: 99%

Dynamic guide–target interactions contribute to sequential 2′-O-methylation by a unique archaeal dual guide box C/D sRNP

Singh

Gurha²,

Gupta³

2008

RNA

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Assembly and guide-target interaction of an archaeal box C/D-guide sRNP was investigated under various conditions by analyzing the lead (II)-induced cleavage of the guide RNA. Guide and target RNAs derived from Haloferax volcanii pre-tRNA Trp were used with recombinant Methanocaldococcus jannaschii core proteins in the reactions. Core protein L7Ae binds differentially to C/D and C9/D9 motifs of the guide RNA, and interchanging the two motifs relative to the termini of the guide RNA did not affect L7Ae binding or sRNA function. L7Ae binding to the guide RNA exposes its D9-guide sequence first followed by the D guide. These exposures are reduced when aNop5p and aFib proteins are added. The exposed guide sequences did not pair with the target sequences in the presence of L7Ae alone. The D-guide sequence could pair with the target in the presence of L7Ae and aNop5p, suggesting a role of aNop5p in target recruitment and rearrangement of sRNA structure. aFib binding further stabilizes this pairing. After box C/D-guided modification, target-guide pairing at the D-guide sequence is disrupted, suggesting that each round of methylation may require some conformational change or reassembly of the RNP. Asymmetric RNPs containing only one L7Ae at either of the two box motifs can be assembled, but a functional RNP requires L7Ae at the box C/D motif. This arrangement resembles the asymmetric eukaryal snoRNP. Observations of initial D-guidetarget pairing and the functional requirement for L7Ae at the box C/D motif are consistent with our previous report of the sequential 29-O-methylations of the target RNA.

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Assembly of the Archaeal Box C/D sRNP Can Occur via Alternative Pathways and Requires Temperature-facilitated sRNA Remodeling

Cited by 22 publications

References 58 publications

Kink turn sRNA folding upon L7Ae binding using molecular dynamics simulations

Kink turn sRNA folding upon L7Ae binding using molecular dynamics simulations

Signature amino acids enable the archaeal L7Ae box C/D RNP core protein to recognize and bind the K-loop RNA motif

Dynamic guide–target interactions contribute to sequential 2′-O-methylation by a unique archaeal dual guide box C/D sRNP

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