October 16, 2012

Frameshifting occurs at slippery sequences

  • Programmed frameshifting is required for expression of the protein sequences coded beyond a specific site at which a +1 or -1 frameshift occurs at some typical frequency.
  • The reading frame may be influenced by the sequence of mRNA and the ribosomal environment.
  • Slippery sequences allow a tRNA to shift by 1 base after it has paired with its anticodon, thereby changing the reading frame.
  • Translation of some genes depends upon the regular occurrence of programmed frameshifting. 

Frameshifting is associated with specific tRNAs in two circumstances (for review see Farabaugh and Bjorkk, 1999):
  • Some mutant tRNA suppressors recognize a "codon" for 4 bases instead of the usual 3 bases.
  • Certain "slippery" sequences allow a tRNA to move a base up or down mRNA in the A site.
Frameshift mutants result from the insertion or deletion of a base. They can be suppressed by restoring the original reading frame. This can be achieved by compensating base deletions and insertions within a gene (see 1.21 The genetic code is triplet). However, extragenic frameshift suppressors also can be found in the form of tRNAs with aberrant properties.
The simplest type of external frameshift suppressor corrects the reading frame when a mutation has been caused by inserting an additional base within a stretch of identical residues. For example, a G may be inserted in a run of several contiguous G bases. The frameshift suppressor is a tRNAGly that has an extra base inserted in its anticodon loop, converting the anticodon from the usual triplet sequence CCC to the quadruplet sequence CCCC . The suppressor tRNA recognizes a 4-base "codon".
Some frameshift suppressors can recognize more than one 4-base "codon". For example, a bacterial tRNALys suppressor can respond to either AAAA or AAAU, instead of the usual codon AAA. Another suppressor can read any 4-base "codon" with ACC in the first three positions; the next base is irrelevant. In these cases, the alternative bases that are acceptable in the fourth position of the longer "codon" are not related by the usual wobble rules. The suppressor tRNA probably recognizes a 3 base codon, but for some other reasonmost likely steric hindrancethe adjacent base is blocked. This forces one base to be skipped before the next tRNA can find a codon.
Situations in which frameshifting is a normal event are presented by phages and viruses. Such events may affect the continuation or termination of protein synthesis, and result from the intrinsic properties of the mRNA.

In retroviruses, translation of the first gene is terminated by a nonsense codon in phase with the reading frame. The second gene lies in a different reading frame, and (in some viruses) is translated by a frameshift that changes into the second reading frame and therefore bypasses the termination codon (see Figure 7.29) (Jacks et al., 1988) (see 17.3 Retroviral genes codes for polyproteins). The efficiency of the frameshift is low, typically ~5%. In fact, this is important in the biology of the virus; an increase in efficiency can be damaging. Figure 7.31 illustrates the similar situation of the yeast Ty element, in which the termination codon of tya must be bypassed by a frameshift in order to read the subsequent tyb gene.
Such situations makes the important point that the rare (but predictable) occurrence of "misreading" events can be relied on as a necessary step in natural translation. This is called programmed frameshifting (for review see Farabaugh, 1995; Gesteland and Atkins, 1996). It occurs at particular sites at frequencies that are 100-1000× greater than the rate at which errors are made at nonprogrammed sites (~3 × 105 per codon).
There are two common features in this type of frameshifting:
  • A "slippery" sequence allows an aminoacyl-tRNA to pair with its codon and then to move +1 (rare) or 1 base (more common) to pair with an overlapping triplet sequence that can also pair with its anticodon.
  • The ribosome is delayed at the frameshifting site to allow time for the aminoacyl-tRNA to rearrange its pairing. The cause of the delay can be an adjacent codon that requires a scarce aminoacyl-tRNA, a termination codon that is recognized slowly by its release factor, or a structural impediment in mRNA (for example, a "pseudoknot," a particular conformation of RNA) that impedes the ribosome.
Slippery events can involve movement in either direction; a 1 frameshift is caused when the tRNA moves backwards, and a +1 frameshift is caused when it moves forwards. In either case, the result is to expose an out-of-phase triplet in the A site for the next aminoacyl-tRNA. The frameshifting event occurs before peptide bond synthesis. In the most common type of case, when it is triggered by a slippery sequence in conjunction with a downstream hairpin in mRNA, the surrounding sequences influence its efficiency.
The frameshifting in Figure 7.31 shows the behavior of a typical slippery sequence. The 7 nucleotide sequence CUUAGGC is usually recognized by Leu-tRNA at CUU followed by Arg-tRNA at AGC. However, the Arg-tRNA is scarce, and when its scarcity results in a delay, the Leu-tRNA slips from the CUU codon to the overlapping UUA triplet. This causes a frameshift, because the next triplet in phase with the new pairing (GGC) is read by Gly-tRNA. Slippage usually occurs in the P site (when the Leu-tRNA actually has become peptidyl-tRNA, carrying the nascent chain).
Frameshifting at a stop codon causes readthrough of the protein. The base on the 3 side of the stop codon influences the relative frequencies of termination and frameshifting, and thus affects the efficiency of the termination signal. This helps to explain the significance of context on termination.