FunctionalPMS2hybrid alleles containing a pseudogene-specific missense variant trace back to a single ancient intrachromosomal recombination event

Abstract

Sequence exchange between PMS2 and its pseudogene PMS2CL, embedded in an inverted duplication on chromosome 7p22, has been reported to be an ongoing process that leads to functional PMS2 hybrid alleles containing PMS2-and PMS2CL-specific sequence variants at the 5′-and the 3′-end, respectively. The frequency of PMS2 hybrid alleles, their biological significance, and the mechanisms underlying their formation are largely unknown. Here we show that overall hybrid alleles account for one-third of 384 PMS2 alleles analyzed in individuals of different ethnic backgrounds. Depending on the population, 14-60% of hybrid alleles carry PMS2CL-specific sequences in exons 13-15, the remainder only in exon 15. We show that exons 13-15 hybrid alleles, named H1 hybrid alleles, constitute different haplotypes but trace back to a single ancient intrachromosomal recombination event with crossover. Taking advantage of an ancestral sequence variant specific for all H1 alleles we developed a simple gDNA-based polymerase chain reaction (PCR) assay that can be used to identify H1-allele carriers with high sensitivity and specificity (100 and 99%, respectively). Because H1 hybrid alleles harbor missense variant p.N775S of so far unknown functional significance, we assessed the H1-carrier frequency in 164 colorectal cancer patients. So far, we found no indication that the variant plays a major role with regard to cancer susceptibility.

Avoidance of pseudogene interference in the detection of 3′ deletions in PMS2

“…This gives the user no knowledge of the denominator in the ratio calculation, and thus it is difficult to draw a conclusion about the number of alleles harboring a particular PSV in the patient sample, that is, the numerator of the ratio calculation. Although the MLPA kit insert recommends pooling normal controls in an effort to diminish the variability introduced by control samples harboring unequal numbers of alleles with each PSV, this is unlikely to fully mitigate the problem as many samples harbor an unequal distribution of each PSV [Ganster et al, 2010;van der Klift et al, 2010]. Indeed, in our study, only 6/20 normal samples had two copies of each PSV for all sites within exons 12-15 (data not shown).…”

“…The newly designed MLPA kit also includes probes specific for both variants of PSV sites in PMS2 and PMS2CL for exons 11-15. Although the PSV site for exon 11 does not appear to undergo sequence transfer [Ganster et al, 2010;van der Klift et al, 2010], extensive sequence transfer between PMS2 exons 12-15 and PMS2CL means that the PSV-specific probes for these exons are also unable to assign the localization of detected deletions to the gene or pseudogene.…”

“…However, subsequent research showed that this portion of PMS2 undergoes gene conversion with PMS2CL, introducing variants that were thought to be exclusive to PMS2 into PMS2CL and vice versa [Auclair et al, 2007;Hayward et al, 2007]. The rate of exchange for the specific PSVs utilized in the MLPA probe design for these exons has recently been further characterized [Ganster et al, 2010;van der Klift et al, 2010]. Because sequences unique to PMS2 (as opposed to the pseudogene) are not present in exons 12-15, the original MLPA kit could not reliably detect deletions in this region of the gene.…”

Pseudogene in cancer: real functions and promising signature

“…Conversions from pseudogene to parental gene have been reported in several human diseases 32 33. Cytochrome P450 2A6 (CYP2A6), for example, enzyme metabolising precarcinogens including nicotine, has a pseudogene called CYP2A7.…”

PMS2 inactivation by a complex rearrangement involving an HERV retroelement and the inverted 100-kb duplicon on 7p22.1

“…5 Recombination-based sequence exchanges also affected the paralogs PMS2 and PMS2CL, situated at the inner ends of the LCR, which led to mainly functional hybrid PMS2 alleles that contain PMS2CL-derived sequences (as defined by NCBI RefSeq NC_000007.13), as well as hybrid PMS2CL alleles, with sequences derived from PMS2 (according to NCBI RefSeq NM_000535.5). [5][6][7] The high prevalence of these hybrid alleles complicates mutation analysis, because in the region of frequent sequence exchange, that is, downstream of PMS2 exon 12, gene and pseudogene cannot be reliably distinguished on the basis of sequence differences with respect to their NCBI RefSeqs. Recombination-based sequence exchanges between the paralogs are thought to be a still ongoing mechanism, which is also responsible for the generation of deleterious PMS2 alleles.…”