Duplication of 7p11.2-p13, including GRB10, in Silver-Russell syndrome

Abstract

Silver-Russell syndrome (SRS) is characterized by pre- and postnatal growth failure and other dysmorphic features. The syndrome is genetically heterogeneous, but maternal uniparental disomy of chromosome 7 has been demonstrated in approximately 7% of cases. This suggests that at least one gene on chromosome 7 is imprinted and involved in the pathogenesis of SRS. We have identified a de novo duplication of 7p11.2-p13 in a proband with features characteristic of SRS. FISH confirmed the presence of a tandem duplication encompassing the genes for growth factor receptor-binding protein 10 (GRB10) and insulin-like growth factor-binding proteins 1 and 3 (IGFBP1 and -3) but not that for epidermal growth factor-receptor (EGFR). Microsatellite markers showed that the duplication was of maternal origin. These findings provide the first evidence that SRS may result from overexpression of a maternally expressed imprinted gene, rather than from absent expression of a paternally expressed gene. GRB10 lies within the duplicated region and is a strong candidate, since it is a known growth suppressor. Furthermore, the mouse homologue (Grb10/Meg1) is reported to be maternally expressed and maps to the imprinted region of proximal mouse chromosome 11 that demonstrates prenatal growth failure when it is maternally disomic. We have demonstrated that the GRB10 genomic interval replicates asynchronously in human lymphocytes, suggestive of imprinting. An additional 36 SRS probands were investigated for duplication of GRB10, but none were found. However, it remains possible that GRB10 and/or other genes within 7p11.2-p13 are responsible for some cases of SRS.

Figure 1

Proband with her mother

Figure 2

Partial karyotype with ideogram showing duplication of chromosome 7p11.2-p13, indicated by arrow; the normal homologue is on the left.

Figure 3

A, FISH photomicrograph of nuclei from the DP, hybridized with the GRB10 containing PAC dJ0108E23 (red) and the control dJ0020F22 (green). Three red hybridization signals were clearly visible in all interphase nuclei, indicative of a duplication, compared with the two control signals (green). In the metaphase spreads, an increase in dJ0108E23 signal intensity was observed on one homologue, indicated by the arrow, confirming a duplication. B, Interphase nucleus from the DP, hybridized with dJ0108E23 (red), dJ069I12 (orange), and dJ0570D02 (green). The orange-red-red-green pattern indicates that the dJ069I12 and dJ0570D02 PACs are not contained within the duplication, unlike the double signal from the dJ0108E23 PAC. The adjacent orange, red, and green signals indicate the normal chromosome 7. C, Interphase nucleus from the DP, hybridized with dJ0108E23 (red) and dJ0647J21 (green). The red-green-red-green pattern suggests a duplication in a tandem arrangement. The adjacent red and green signals indicate the normal chromosome 7.

Figure 4

Physical map of 7p11.2-p14, showing the extent of the duplication as detected by interphase FISH. The duplication was analyzed by use of 17 PAC clones from a 16-cM region containing the 10-cM duplication.

Figure 5

Tetranucleotide-repeat markers for loci D7S3069 (A) and D7S1818 (B). Signal intensity was measured by use of a PhosphorImager. The relative intensity of the two bands is expressed as a ratio of upper to lower signals (±SE) in the mother (lanes M), proband (lanes P), and father (lanes F). For both markers, the maternally inherited band in the proband was of increased intensity.

Figure 6

Southern blot hybridization of DNA from normal controls (lanes 1, 4, and 7) and SRS probands (lanes 2, 3, 5, and 6), digested with XbaI. The positions of the upper and lower GRB10 bands and of the A6121-1 band are indicated on the left. DNA from the proband with the 7p11.2-p13 duplication and increased GRB10 dosage is shown in lane 5. DNA from proband 1 with XbaI RFLP (arrow) and with hemizygosity for the upper GRB10 band is shown in lane 6.

Figure 7

Histograms representing the ratio of GRB10 signal to control for 37 probands, as determined by quantitative analysis of Southern blot hybridization. A, Upper GRB10 band. B, Lower GRB10 band. SD and mean for the sample are given, for both sets of ratios. Ratios for DP lie well above the normal range, confirming increased dosage of GRB10. No other SRS probands have similarly increased ratios.