Very small deletions within the NESP55 gene in pseudohypoparathyroidism type 1b

Pseudohypoparathyroidism (PHP) is caused by reduced expression of genes within the GNAS cluster, resulting in parathormone resistance. The cluster contains multiple imprinted transcripts, including the stimulatory G protein α subunit (Gs-α) and NESP55 transcript preferentially expressed from the maternal allele, and the paternally expressed XLas, A/B and antisense transcripts. PHP1b can be caused by loss of imprinting affecting GNAS A/B alone (associated with STX16 deletion), or the entire GNAS cluster (associated with deletions of NESP55 in a minority of cases). We performed targeted genomic next-generation sequencing (NGS) of the GNAS cluster to seek variants and indels underlying PHP1b. Seven patients were sequenced by hybridisation-based capture and fourteen more by long-range PCR and transposon-mediated insertion and sequencing. A bioinformatic pipeline was developed for variant and indel detection. In one family with two affected siblings, and in a second family with a single affected individual, we detected maternally inherited deletions of 40 and 33 bp, respectively, within the deletion previously reported in rare families with PHP1b. All three affected individuals presented with atypically severe PHP1b; interestingly, the unaffected mother in one family had the detected deletion on her maternally inherited allele. Targeted NGS can reveal sequence changes undetectable by current diagnostic methods. Identification of genetic mutations underlying epigenetic changes can facilitate accurate diagnosis and counselling, and potentially highlight genetic elements critical for normal imprint settin

Maternal variants in NLRP and other maternal effect proteins are associated with multilocus imprinting disturbance in offspring

Background: Genomic imprinting results from the resistance of germline epigenetic marks to reprogramming in the early embryo for a small number of mammalian genes. Genetic, epigenetic or environmental insults that prevent imprints from evading reprogramming may result in imprinting disorders, which impact growth, development, behaviour and metabolism. We aimed to identify genetic defects causing imprinting disorders, by whole-exome sequencing in families with one or more members affected by multi-locus imprinting disturbance. Methods: Whole-exome sequencing was performed in 38 pedigrees where probands had multi-locus imprinting disturbance, in five of whom, maternal variants in NLRP5 have previously been found. Results: We now report 15 further pedigrees in which offspring had disturbance of imprinting, while their mothers had rare, predicted-deleterious variants in maternal-effect genes, including NLRP2, NLRP7 and PADI6. As well as clinical features of well-recognised imprinting disorders, some offspring had additional features including developmental delay, behavioural problems and discordant monozygotic twinning, while some mothers had reproductive problems including pregnancy loss. Conclusion: The identification of 20 putative maternal-effect variants in 38 families affected by multi-locus imprinting disorders adds to the evidence that maternal genetic factors affect oocyte fitness and thus offspring development. Testing for maternal-effect genetic variants should be considered in families affected by atypical imprinting disorders.<br/

Human imprinting disorders: Principles, practice, problems and progress

Epigenetic regulation orchestrates gene expression with exquisite precision, over a huge dynamic range and across developmental space and time, permitting genomically-homogeneous humans to develop and adapt to their surroundings. Every generation, these epigenetic marks are re-set twice: in the germline, to enable differentiation of sperm and eggs, and at fertilisation, to create the totipotent zygote that then begins growth and differentiation into a new human. A small group of genes evades the second, zygotic wave of epigenetic reprogramming, and these genes retain an epigenetic ‘imprint’ of the parent from whom they were inherited.Imprinted genes are (as a general rule) expressed from one parental allele only. Some imprinted genes are critical regulators of growth and development, and thus disruption of their normal monoallelic expression causes congenital imprinting disorders, with clinical features impacting growth, development, behaviour and metabolism.Imprinting disorders as a group have characteristics that challenge diagnosis and management, including clinical and molecular heterogeneity, overlapping clinical features, somatic mosaicism, and multi-locus involvement. New insights into the biology and epigenomics of the early embryo offers new clues about the origin and importance of imprinting disorders

Transient Neonatal Diabetes Mellitus followed by recurrent asymptomatic hypoglycaemia: a case report

Background: Transient Neonatal Diabetes Mellitus is the commonest cause of diabetes presenting in the first week of life. Majority of infants recover by 3 months of age but are predisposed to developing type 2 diabetes later on in life. This condition is usually due to genetic aberrations at the 6q24 gene locus, and can be sporadic or inherited. This disorder has three phases: neonatal diabetes, apparent remission, relapse of diabetes.Case Presentation: Our case, a neonate presented with low birth weight and growth retardation along with the metabolic profile consistent with transient diabetes mellitus at birth. We report a novel clinical observation of recurrent asymptomatic hypoglycaemia detected on pre-feed blood glucose level monitoring in our case with transient neonatal diabetes mellitus at 6 weeks of age, 4 weeks after the remission of diabetes mellitus.Conclusion: This case demonstrates that neonates in remission following transient diabetes mellitus can present with recurrent asymptomatic hypoglycaemia without any other obvious congenital malformations seen. This asymptomatic hypoglycaemia may persist for weeks and may be missed if pre-feed blood glucose level monitoring is not done in these infants. Also, these infants may require an aggressive enteral feeding regimen with high glucose delivery rate to maintain normoglycemia.<br/

Bisulphite sequencing of the transient neonatal diabetes mellitus DMR facilitates a novel diagnostic test but reveals no methylation anomalies in patients of unknown aetiology

Transient neonatal diabetes mellitus (TNDM) is associated with overexpression of an imprinted locus on chromosome 6q24; this locus contains a differentially methylated region (DMR) consisting of an imprinted CpG island that normally allows expression only from the paternal allele of genes under its control. Three types of abnormality involving 6q24 are known to cause TNDM: paternal uniparental disomy of chromosome 6 (pUPD6), an isolated methylation defect of the imprinted CpG island at chromosome 6q24 and a duplication of 6q24 of paternal origin. A fourth group of patients has no identifiable anomaly of 6q24. Bisulphite sequencing of the DMR has facilitated the development of a diagnostic test for TNDM based on ratiometric methylation-specific polymerase chain reaction. We have applied this method to 45 cases of TNDM, including 12 with pUPD6, 11 with an isolated methylation mutation at 6q24, 16 with a duplication of 6q24 and six of unknown aetiology, together with 29 normal controls. All were correctly assigned. The method is therefore capable of detecting all known genetic causes of TNDM at 6q24, although pUPD6 and methylation mutation cases are not distinguished from one another. In addition, we have carried out bisulphite sequencing of the DMR to compare its methylation status between six TNDM patients with a known methylation mutation, six patients with no identifiable 6q24 mutation and six normal controls. Whereas methylation mutation patients showed a near-total absence of DNA methylation at the TNDM locus, the patients with no identified molecular anomaly showed no marked methylation variation from controls

Transient neonatal diabetes mellitus in a Turkish patient with three novel homozygous variants in the ZFP57 gene

Neonatal diabetes mellitus (NDM) is a rare form of diabetes that presents within the first six months of life. Nearly 70% of these cases have loss of methylation at the differentially methylated region on chromosome 6q24. To describe the findings in a Turkish male patient with NDM caused by a loss of methylation at chromosome 6q24 and three novel homozygous mutations in the ZFP57 gene, methylation-specific PCR was carried out at 6q24 and mutation analysis of ZFP57 gene was maintained by direct sequencing. Sequencing of ZFP57 gene revealed the hypomethylation of chromosome 6q24 and three novel mutations (chr6:29.641.413 A&gt;T, 29.641.073 C&gt;T, and 29.640.855 G&gt;C), respectively. The latter mutation seems to display the patient’s condition due to a highly conservative amino acid substitution in the protein. We suggest the ZFP57 gene as a causative factor for NDM and it should be considered in genetic testing. Further studies including functional analysis of the detected mutations will provide precise information regarding the effect of the mutations

DNA hypomethylation, transient neonatal diabetes, and prune belly sequence in one of two identical twins

One known genetic mechanism for transient neonatal diabetes is loss of methylation at 6q24. The etiology of prune belly sequence is unknown but a genetic defect, affecting the mesoderm from which the triad abdominal muscle hypoplasia, urinary tract abnormalities, and cryptorchidism develop, has been suggested. We investigated a family, including one twin, with transient neonatal diabetes and prune belly sequence. Autoantibody tests excluded type 1 diabetes. Microsatellite marker analysis confirmed the twins being monozygotic. We identified no mutations in ZFP57, KCNJ11, ABCC8, GCK, HNF1A, HNF1B, HNF3B, IPF1, PAX4, or ZIC3. The proband had loss of methylation at the 6q24 locus TNDM and also at the loci IGF2R, DIRAS3, and PEG1, while the other family members, including the healthy monozygotic twin, had normal findings. The loss of methylation on chromosome 6q24 and elsewhere may indicate a generalized maternal hypomethylation syndrome, which accounts for both transient neonatal diabetes and prune belly sequenc