The ubiquitous hammerhead ribozyme

Abstract

The hammerhead ribozyme is a small catalytic RNA motif capable of endonucleolytic (self-) cleavage. It is composed of a catalytic core of conserved nucleotides flanked by three helices, two of which form essential tertiary interactions for fast self-scission under physiological conditions. Originally discovered in subviral plant pathogens, its presence in several eukaryotic genomes has been reported since. More recently, this catalytic RNA motif has been shown to reside in a large number of genomes. We review the different approaches in discovering these new hammerhead ribozyme sequences and discuss possible biological functions of the genomic motifs.

FIGURE 1.

The hammerhead ribozyme. (A) The reaction proceeds via an S_N2 mechanism, in which the hydroxy moiety at C2′ attacks the neighboring 3′–5′ phosphodiester bond. The pentagonal bipyramidal transition state is adopted both in the forward cleavage and in the reverse ligation reaction. The cleavage products are a 2′,3′-cyclic phosphate at the 5′ product and 5′-hydroxyl at the 3′ cleavage product, which serve as substrates for the ligation reaction. (B) The catalytic core of HHR consists of conserved nucleotides (bold) flanked by helices I–III. The conventional numbering system is indicated (Hertel et al. 1992). Cleavage takes places between nucleotides 17 and 1.1, as indicated by the arrow. Dotted lines indicate backbone continuity. (C) Natural forms occur in three types, named after the open stem. Interactions between nucleotides of loops L1 and L2 are observed in all three topologies (double-headed arrows). Additionally, a base pair is formed between nucleotides C₃ and G₈, or variants thereof (see text). Only the three natural topologies are active under physiological magnesium ion conditions, while the minimal format is not. Pattern descriptors for RNABOB (D), PatScan (E), or RNAmotif (F), describing for each a type III motif with the following features: stem III of 3–6 bp, U, H, stem I of 4–7 bp, loop L1 of 4–100 nt, CUGANGA, stem II of 4–6 bp, loop L2 of 4–100 nt GAAA (for details, see D'Souza et al. 1997; Macke et al. 2001; Eddy 2005).

FIGURE 2.

Some examples of genomic HHRs detected in bacteria (A), plants (B), and metazoans (C). Main tertiary contacts based on previous experimental models (Martick and Scott 2006; Chi et al. 2008; Dufour et al. 2009), and the conserved nucleotides involved in the interactions are tentatively depicted in red.

FIGURE 3.

Structural versatility of stem–stem II interaction. Many different possible permutations that allow these two stems to interact are observed. (A) Tertiary interactions between stem I and II. Gray arrows illustrate the tertiary interaction. HHR examples pictured from left to right are found in the nematode Schistosoma mansoni (this example has been crystallized and shows the importance of the tertiary interaction), the viroid ASBVd, the bacterium Bukholderia oklahomensis, the mouse, and the viroid CChMVd (circled nucleotides are typically involved in this interaction for HHR sequences found in viroids). (B) Base-pairing interactions between stems I and II. The large array of different putative pseudoknots or kissing–loop interactions observed in HHR sequences is pictured. HHR examples pictured from left to right on the first row are found in the fungus Yarrowia lipolytica, the mouse and human microbiome, the bacterium Agrobacterium tumefaciens, and the fly Drosophila pseudoobscura. HHRs of the second row are found in the crickets Dolichopoda, marine metagenome, and in the bee Apis melifera.

FIGURE 4.

Possible function of the HHR in genomes. (A) HHR-mediated integration of nucleic acid sequences in genomes. (B) Self-cleavage of HHR sequences within transcripts of repeats or transposon clusters might interfere with the production of piRNAs or other small RNAs.

FIGURE 5.

Possible functions of HHR motifs in various transcriptional contexts. (A) Processing of polycistronic tRNA transcripts. (B) Generation of small RNAs. (C) Generation of alternative open reading frames by HHR self-cleavage and re-ligation. (D) Inhibition of RNAi by prevention of the generation of complementary RNA strands from fungal and retrotransposon promoters.