Novel 14q32.2 paternal deletion encompassing the whole DLK1 gene associated with Temple syndrome

Abstract

Background: Temple syndrome (TS14) is a rare imprinting disorder caused by maternal UPD14, imprinting defects or paternal microdeletions which lead to an increase in the maternal expressed genes and a silencing the paternally expressed genes in the 14q32 imprinted domain. Classical TS14 phenotypic features include pre- and postnatal short stature, small hands and feet, muscular hypotonia, motor delay, feeding difficulties, weight gain, premature puberty along and precocious puberty.

Methods: An exon array comparative genomic hybridization was performed on a patient affected by psychomotor and language delay, muscular hypotonia, relative macrocephaly, and small hand and feet at two years old. At 6 years of age, the proband presented with precocious thelarche. Genes dosage and methylation within the 14q32 region were analyzed by MS-MLPA. Bisulfite PCR and pyrosequencing were employed to quantification methylation at the four known imprinted differentially methylated regions (DMR) within the 14q32 domain: DLK1 DMR, IG-DMR, MEG3 DMR and MEG8 DMR.

Results: The patient had inherited a 69 Kb deletion, encompassing the entire DLK1 gene, on the paternal allele. Relative hypermethylation of the two maternally methylated intervals, DLK1 and MEG8 DMRs, was observed along with normal methylation level at IG-DMR and MEG3 DMR, resulting in a phenotype consistent with TS14. Additional family members with the deletion showed modest methylation changes at both the DLK1 and MEG8 DMRs consistent with parental transmission.

Conclusion: We describe a girl with clinical presentation suggestive of Temple syndrome resulting from a small paternal 14q32 deletion that led to DLK1 whole-gene deletion, as well as hypermethylation of the maternally methylated DLK1-DMR.

Keywords: DLK1; DMR; Deletion; Methylation; Temple syndrome (TS14).

Fig. 1

Clinical traits present in our case. The photographs show prominent forehead, almond-shaped eyes, broad nasal tip small hand and attached lobe in ear

Fig. 2

Deletions involving the DLK1 gene identified in this study and those reported in the literature

Fig. 3

Family tree. III.2 is the TS14 index case. The remaining family members have normal phenotypes, with I.2, II.2, II.3 carrying the DLK1 microdeletion on the maternal allele

Fig. 4

Methylation profiling of the TS14 index case and familial members. A Pyrosequencing was used to quantify methylation of CpG dinucleotides in the DLK DMR (maternally methylated), IG-DMR (paternally methylated), MEG DMR (paternally methylated) and the MEG8 DMR (maternally methylated). Violin plots represent the average methylation profiles of 19 control individuals, with data points for family members located alongside. The black and white boxes above and below the violin plots represent methylation values for UPD(14)pat and UPD(14)mat control samples respectively. B Bisulfite PCR followed sub-cloning for amplicons from proband III.1 and a control leukocyte DNA sample. Each circle represent a single dinucleotide on the DNA stand. (•) methylated cytosine, (o) unmethylated cytosine. Each row corresponds to an individual cloned sequence. CpG dinucleotides in red box indicated those quantified by pyrosequencing

Fig. 5

Structure of the imprinted 14q32 region, paternal and maternal DLK1 gene deletion. Maternally expressed genes are indicated in red boxes and paternally expressed genes are indicated by blue boxes. The methylation is indicated by black lollipops. The DLK1-DMR is maternally methylated (Mat), MEG3/DLK1:IG-DMR and MEG3:TSS-DMR are paternally methylated (Pat) and MEG8: Int2-DMR is maternally methylated (Mat)