Exclusion of a disease relevant role of PAX4 in the aetiology of Silver-Russell syndrome: screening for mutations and determination of imprinting status.

EDITOR—Silver-Russell syndrome (SRS) describes a uniform malformation syndrome characterised by intrauterine and postnatal growth retardation (IUGR/PGR), asymmetry of the head and limbs, a small triangular face, and other less constant features. The majority of the 400 cases described so far occurred sporadically, but some familial cases indicate a genetic cause of the disease. In rare cases, chromosomal aberrations have been found. Although no uniform pattern is apparent, five cases have recently been reported involving chromosome 7. A subset of 7-10% of SRS patients shows maternal uniparental disomy (mUPD) for the whole of chromosome 7, thus indicating the involvement of at least one imprinted gene on this chromosome. Mutations in this gene or imprinting mutations may contribute to the SRS phenotype. So far, two imprinted genes localised in 7q32 (MEST/PEG1, COP-G2) have been excluded as candidate genes for SRS. 9 PAX4 is a member of a highly conserved gene family, and has been mapped to 7q32. Evidence of the crucial role of PAX genes in organogenesis and in diVerentiation is provided by mouse developmental mutants as well as by human diseases (PAX2, kidney abnormalities, optic nerve coloboma; PAX3, Waardenburg syndrome types I and III; PAX5, small lymphotic lymphomas; PAX6, aniridia; PAX9, optic nerve coloboma). 13 14 A role for PAX4 in the aetiology of diabetes type II has recently been dismissed by investigations in a group of French patients as well as in a population of 116 unrelated Ashkenazi Jews aVected by the disease. 16 Furthermore, PAX4 has been ruled out as a possible candidate gene for the Wolcott-Rallison syndrome. The PAX4 gene product contains two DNA binding motifs, a paired domain which consists of three alpha helices, and a paired type homeodomain with a helix turn helix motif; owing to its localisation in the nucleus and its ability to bind DNA, it might serve as a transcription factor. PAX4 is expressed predominantly in placenta and skeletal muscle; fewer transcripts have been observed in the heart. Detection of Pax4 transcripts in the early pancreas of mice in combination with results of knock out experiments indicate an important role for Pax4 during development: mice homozygously deficient for Pax4 suVer from growth retardation and dehydration at birth and die within three days. Its expression in pancreatic island progenitor cells, which are the common progenitor for beta and delta cells, controls the initial processes of endocrine diVerentiation. In the mature endocrine pancreas, it might contribute to the maintenance of the diVerentiated state of beta cells, since then Pax4 is restricted to this cell population. So far, there is no information available about the imprinting status of PAX4. Since imprinted genes tend to be clustered, the close proximity of PAX4 to MEST/PEG1 and COP-G2, which have recently been shown to be paternally expressed, renders its imprinting possible. Taking into account its function, the very restricted spatial and temporary pattern of expression during embryogenesis, and its localisation in 7q32, PAX4 was considered to be a good candidate gene for SRS. Our study population consisted of 50 SRS patients in which diagnosis was ascertained according to Wollmann et al. Cytogenetic analyses by G banding on peripheral blood lymphocyte cultures of these patients did not show any abnormalities. A maximum of 53 unrelated, healthy probands of German origin were investigated as controls. Genomic DNA from patients and controls was isolated