MBE Advance Access originally published online on April 18, 2006
Molecular Biology and Evolution 2006 23(7):1339-1340; doi:10.1093/molbev/msk024
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Letter |
Evidence that Protein Length Expansion and Contraction Is Partly Due to Mutational Events in Premeiotic Cells
Department of Biology and Biochemistry, University of Bath, Bath, United Kingdom
E-mail: bssaew{at}bath.ac.uk.
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Abstract |
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Studies on the rate of evolution of proteins typically concentrate on rates of change of orthologous amino acids rather than on changes in size (i.e., generation of nonorthologous domains). Recent work has focused attention on Ser/Thr-rich regions in yeast as these tend to undergo size changes rapidly, with size polymorphisms commonly being found, especially in proteins with cell-surface localization. The underlying mechanism generating the indels is presently unclear though, due to a lack of correlation with the location of meiotic double-strand breaks, it has, by exclusion, been conjectured to be replication slippage. Here we provide new evidence to support this possibility. Notably, we show that Ser/Thr-rich repeat regions are more generally associated with the location of Mre11p in premeiotic cells. This is to be expected if the repeats were produced by mutational events in mitotic cells possibly through replication slippage.
Key Words: replication slippage • Ser/Thr rich • polymorphisms • yeast
Although the rate of evolution of orthologous amino acids within orthologous proteins in yeast is well established (Dujon et al. 2004
; Byrne and Wolfe 2005), the rate of evolution of size within individual proteins is still not fully understood. Indel regions in alignments are often ignored in preference to conserved regions when considering protein evolution. Of particular interest in this regard is the finding that, in yeast, many cell-surface proteins are polymorphic for size and that this polymorphism is often associated with indel events in DNA that can give rise to repeated regions rich in serines and threonines (Bowen et al. 2005; Verstrepen et al. 2005). Recent evidence suggests that the polymorphism is likely to be under selection as differently sized orthologs show differing levels of mutual attraction (Verstrepen et al. 2005). The polymorphism may hence be associated with selection for yeasts to aggregate, for example, on first colonizing a new location, and with dispersal (i.e. nonaggregation) when that location becomes saturated.
Though selection may well be acting on the polymorphism, the question remains as to how the indel events are generated. Two possibilities are conventionally discussed: gene conversion through nonhomologous recombination (Jeffreys et al. 1998; Bishop et al. 2000; Pâques et al. 2001) and replication-associated slippage (Rolfsmeier et al. 2001; Verstrepen et al. 2005). Recently, Richard and Dujon (2006) argued against the former possibility as they observed no correlation between the location of meiotic double-strand breaks and tandem repeats. They hence argued that, by exclusion, replication slippage is the more likely mechanism. They sensibly caution, however, that this should not be considered in any manner to be definitive as the location of hot and cold spots of recombination may change over time. Here we provide an alternative line of evidence, which we suggest provides support for the hypothesis of replication-associated slippage.
Our test requires 2 assumptions. First, that oligopeptide repeats rich in serine and threonine residues are, in yeast, subject to selection. Second, that data on the premeiotic location of Mre11p in the data set of Borde et al. (2004) indicate the location of mutational events, such as replication slippage or strand breaks. Mre11 is a nuclease that forms part of a DNA repair complex involved in the processing of specific DNA structures such as double-stranded breaks in meiosis and certain secondary structures in mitosis (Richard et al. 2000; Debrauwère et al. 2001; Lewis et al. 2004). Experimental evidence in mammalian cells has found that the Mre11p complex can act as a DNA damage sensor following S phase in the cell cycle (Carson et al. 2003). Mutational events through replication slippage occur in premeiotic cells most probably because meiotic entry requires cell cycle arrest and alternative replication and DNA damage checkpoints (Viguera et al. 2001; Murakami and Nurse 2000).
Given this, we can then ask whether Ser/Thr-rich oligopeptide repeats are associated with premeiotic Mre11 locations or meiotic ones. Coding DNA sequences of Saccharomyces cerevisiae S288C in FASTA format were downloaded from the Saccharomyces Genome Database (http://www.yeastgenome.org, Dolinski et al. 2005). Genomic occurrence of repeats was evaluated using Tandem Repeat Finder (Benson 1999). Polymorphisms are also known to occur through trinucleotide repeats that encode single amino acid tracts (Young et al. 2000); these were therefore removed from the data set leaving open reading frames (ORFs) that mainly encode Ser/Thr-rich oligopeptides (n = 188; Supplementary Table 1, Supplementary Material online). A Perl script was used to determine Ser and Thr genomic content in translated ORFs. Raw microarray data of Mre11p chromatin association were downloaded from http://nicolas.curie.fr/mre11-hotspots.html (Borde et al. 2004). Correlations between the median value of raw microarray data were determined first against repeat copy number and then Ser/Thr content, of translated ORFs containing oligopeptide repeats.
As Richard and Dujon (2006) have shown, there is no evidence for a correlation between ORFs containing repeated regions and meiotic recombination, a result that we can replicate (fig. 1). Significantly, however, we observe a striking correlation between repeat content and premeiotic locations, consistent with the replication slippage model (fig. 2). Simulations (100,000) of the data set were used to give an estimate of significance for each calculated r value in premeiotic (P = 0.00049) and meiotic (P = 0.5415) cells. In the simulations, a randomization strategy was used by which the observed Mre11p association values are reordered randomly and used to correlate with the observed repeat copy number in ORFs. Moreover, we find an even higher correlation between premeiotic data and Ser/Thr content (see supplementary figures, Supplementary Material online). These results in turn suggest that DNA regions encoding a high Ser/Thr content are associated with repetitive structures that may be due to DNA damage or replication slippage prior to meiosis. Further Mre11p analysis based on indel size through various stages of the cell cycle would allow a greater understanding of these mechanisms.
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The above results suggest therefore that, at least in yeast, intragenic indels are largely associated with premeiotic DNA damage that may include replication slippage, reinforcing the suggestion of Richard and Dujon (2006)
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Supplementary Material |
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TOPAbstractSupplementary MaterialAcknowledgementsReferences |
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Supplementary Table 1 and figures are available at Molecular Biology and Evolution online (http://www.mbe.oxfordjournals.org/).
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Acknowledgements |
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TOPAbstractSupplementary MaterialAcknowledgementsReferences |
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The authors would like to thank Laurence Hurst for assistance with statistics and in compiling the manuscript.
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Footnotes |
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Edward Holmes, Associate Editor
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References |
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TOPAbstractSupplementary MaterialAcknowledgementsReferences |
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