binding nuclear proteins which regulate alternative splicing (36). YTH is a 100- to 150-residue domain present in the mammalian pre-mRNA splicing factor YT521-B (44). This domain is predicted to have a mixed alpha-helix-beta-sheet fold, with four alpha-helices and six beta-strands; the conservation of aromatic residues in the beta-sheets is similar to the RNA recognition motif (RRM) domain, and its predicted biological function is to bind to RNA (36). In two different studies, the gene product of YTHDF2 was shown to be a tumor-associated antigen eliciting high-titer immunoglobulin G antibody responses in cancer patients, and described as renal cell carcinoma–associated ‘‘NY- REN-2 antigen’’ (45) and as chronic lymphocytic leukemia- associated ‘‘KW-14’’ antigen (46). In both studies, the mRNA of the YTHDF2 gene was found to be expressed also in normal tissues, but no quantitative data were reported. We quantitatively evaluated YTHDF2 mRNA expression in 16 different human tissues; the gene resulted to be expressed in all the examined samples, but with variable expression levels and with highest expression in the testis. The relative YTHDF2 mRNA expression in this tissue was 7-fold to 8-fold higher than in small intestine, skeletal muscle, and heart, and 3-fold to 5-fold higher than in the majority of other tissues. A relatively high expression level was also found in placenta. The analysis of the public EST database supports the observation of highest expression in testis. A possible role of the YTHDF2 gene in both fertility and longevity would represent an intriguing hypothesis. In fact, the inverse correlation of longevity and reproductive success is a fundamental assumption of the disposable soma theory (47–49): This prediction has been confirmed in model systems (50), and positive evidence in humans has also been reported (51). The trade-off between fertility and longevity often seems to imply genes involved in carbohydrate homeostasis (52–55). In this regard, it is interesting to note that the YTHDF2 gene is reported to be regulated by high glucose concentration (see GenBank accession no. AF192968). Thus, it could be worthwhile to test the hypothesis of a possible role of YTHDF2 in the insulin/

each allele is similar, and tends to increase when some alleles became highly prevalent. In fact, the frequency comparison of the most abundant alleles in the two age groups confirmed the ‘‘more uneven’’ allele frequency distribution in centenarians, and in particular showed an evident increase of the ‘‘15’’ allele, the most frequent one, and an overall decrease of the less abundant alleles (alleles with frequency lower than 10% in both age groups). Even when the distribution of the most abundant genotypes was analyzed, a significant difference was observed between the two age groups. In particular, odds ratios indicate that carriers of the ‘‘15-15’’ genotype had a doubled chance to become centenarians in comparison with carriers of the less abundant genotypes, i.e., genotypes with frequency less than 10% in both age groups, which on the whole constitute the prevalent genotypic class. These results indicate that, in the Italian population, individuals carrying the most abundant allele (the ‘‘15’’allele) or the corresponding homozygous genotype at YTHDF2 (TG)12-27 locus may have a selective advantage, and an increased probability to reach extremely advanced ages. It is important to note that, differently from what observed in many studies on longevity-associated loci (24,37–39), the data here presented do not suggest any gender-specific association for this locus.

longevity-associated variants of this polymorphism, we analyzed the expression of the YTHDF2 gene in cells having different genotypes. In particular we assessed, by relative quantitative real-time PCR, the YTHDF2 mRNA level in cultured cell lines derived from 10 individuals having the ‘‘15-15’’ genotype, and from 41 individuals having different genotypes. Cell lines were Epstein-Barr Virus-immortalized B lymphocytes, which have been proved to be a reliable model in correlating genotype and gene expression (25,40). The results showed that cell lines with the ‘‘15-15’’ genotype have, on average, more than two times higher levels of YTHDF2 mRNA expression in comparison with cell lines with other genotypes. This finding suggests the presence of different functional variants of the YTHDF2 gene, where the ‘‘15’’ allele is probably associated with a variant characterized by a higher expression level, at least in the described experimental conditions. These data are compatible with the hypothesis that the alleles at this locus could be in linkage disequilibrium with still unidentified genetic variants in the promoter or in other regulatory sites, but a direct influence of YTHDF2 (TG)12-27 polymorphism itself on gene transcription can not be excluded. Indeed, it is not uncommon to find enhancers and regulatory sites in introns, and polymorphic microsatellites can influence gene expression by modifying the distance between specific regulatory sites (41), or by inducing the DNA to assume left-handed helical structures (Z-DNA) (42,43).

data, of the above described peculiar expression pattern in normal tissues, and of an antigenic activity in cancer cells, is typical of a class of genes mainly regulated by epigenetic modifications, the so-called ‘‘cancer-testis antigens’’ (CT antigens) (56,57). They are a heterogeneous group of pro- teins, presumably involved in cell-cycle regulation or tran- scriptional control (58), present as antigens in various cancer cells, and predominantly expressed in testis, placenta, and/or ovary. CT genes are often highly expressed in pancreas (59), which noticeably was the third sample showing a relatively high YTHDF2 expression in our RT–PCR assay.