Nucleotide sequence and structural analysis of two satellite RNAs associated with chicory yellow mottle virus

Rubino, Luisa; Tousignant, M. E.; Steger, Gerhard; Kaper, J.M.

doi:10.1099/0022-1317-71-9-1897

Cited by 73 publications

(49 citation statements)

References 31 publications

(37 reference statements)

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“…The hammerheads were primarily chosen on the basis of having diverse loop 1 and loop 2 sequences; however, extremely large loops were avoided to minimize potential misfolding in the in vitro assays. While the cleavage properties of several (including CChMVd, rCSCVd, PLMVd, satTRsV) have previously been investigated biochemically (10,19,23,24), others (including satSCMoV) have only been shown to cleave in vitro (25), while still others (including satCYMoV, satArMV) are only proposed to self-cleave based upon their sequence (11,26).…”

Section: Resultsmentioning

confidence: 99%

Catalytic Diversity of Extended Hammerhead Ribozymes

Shepotinovskaya

Uhlenbeck

2008

Biochemistry

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Chimeras of the well characterized minimal hammerhead 16 and nine extended hammerheads derived from natural viroids and satellite RNAs were constructed with the goal of assessing whether their very different peripheral tertiary interactions modulate their catalytic properties. For each chimera, three different assays were used to determine the rate of cleavage and the fraction of full length hammerhead at equilibrium and thereby deduce the elemental cleavage (k 2 ) and ligation (k -2 ) rate constants. The nine chimeras were all more active than minimal hammerheads and showed a very broad range of catalytic properties, with values of k 2 varying by 750-fold and k -2 by 100-fold. At least two of the hammerheads showed an altered dependence of k obs on magnesium concentration. Since much less catalytic diversity is observed among minimal hammerheads that lack the tertiary interactions, a possible role for the different tertiary interaction is to modulate the hammerhead cleavage properties in viroids. For example, differing hammerhead cleavage and ligation rates could affect the steady state concentrations of linear, circular, and polymeric genomes in infected cells.

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Section: Resultsmentioning

confidence: 99%

Catalytic Diversity of Extended Hammerhead Ribozymes

Shepotinovskaya

Uhlenbeck

2008

Biochemistry

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“…Each of the three naturally occurring variants also shows a substrate purine and ribozyme pyrimidine at these positions ( Fig. 3B} (Rubino et al 1990). The selection experiments support a base pair at positions corresponding to g-4 " C13, but mismatches and wobble pairs are found more frequently than at the two preceding positions.…”

Section: In Vitro Selectionmentioning

confidence: 99%

Substrate selection rules for the hairpin ribozyme determined by in vitro selection, mutation, and analysis of mismatched substrates.

Joseph¹,

Berzal‐Herranz²,

Chowrira³

et al. 1993

Genes Dev.

104

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Substrate recognition by the hairpin ribozyme has been proposed to involve two short intermolecular helices, termed helix ! and helix 2. We have used a combination of three methods (cleavage of mismatched substrates, in vitro selection, and site-specific mutational analysis) to systematically determine the substrate recognition rules for this RNA enzyme. Assays measuring substrate cleavage in trans under multiple turnover conditions were conducted using the wild-type ribozyme and substrates containing mismatches in all sites potentially recognized by the ribozyme. Molecules containing single-and multiple-base mismatches in helix 2 at sites distant from the cleavage site (g_4c, u_sa, g-4c : u_sa) were cleaved with reduced efficiency, whereas those with mismatches proximal to the cleavage site (c_2a, a_~c, c_2a : a_3c) were not cut. Analogous results were obtained for helix 1, where mismatches distal from the cleavage site (u+Ta , u+sa, u+9a, u+Ta : U+sa : u+9a) were used much more eiiicienfly than those proximal to the cleavage site (c+4a, u_sa, g+6 c, c+4a : u+sa : g+6c). In vitro selection experiments were carried out to identify active variants from populations of molecules in which either helix 1 or helix 2 was randomized. Results constitute an artificial phylogenetic data base that proves base-pairing of nucleotides at five positions within helix 1 and three positions within helix 2 and reveals a significant sequence bias at 3 bp (c+4" G6, c-2" Gll, and a_ 3 9 U12 ). This sequence bias was confirmed at two sites by measuring relative cleavage rates of all 16 possible dinucleotide combinations at base pairs c+4" G6 and c-2" Gll. The strong sequence preference suggests that tertiary structure (e.g., base triples) or alternative secondary structures at these sites may be important for ribozyme function. Together with results from previous work, we conclude that hairpin ribozymes can be engineered to cleave substrates containing the sequence 5'-

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“…However such a polypeptide has never been observed in infected plants. Other nepoviruses, including tomato black ring virus (TBRV) (Fritsch et al, 1978), strawberry latent ringspot virus (Mayo et al, 1982), myrobalan latent ringspot virus (Fritsch et al, 1984), ArMV (Liu et al, 1990) virus (Rubino et al, 1990), also contain satellite RNAs which encode a satellite protein in the range of 38K to 48K (reviewed in Fritsch & Mayo, 1989). For TBRV, a polypeptide which coelectrophoresed with the in vitro translation product of the satellite RNA of TBRV was detected in infected protoplasts but not in plants (Fritsch et al, 1978).…”

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confidence: 99%