ABSTRACT
We encountered a Chinese girl with pseudohypoparathyroidism type 1A (PHP1A) and her mother with pseudopseudohypoparathyroidism (PPHP). Sequencing analysis of GNAS-Gsα revealed a heterozygous c.212+2T>C variant (NM_000516.4) affecting the canonical splice donor site of intron 2 in the girl and her mother. RT-PCR performed on mRNA samples obtained from cycloheximide-treated and cycloheximide-untreated lymphoblastoid cell lines of this girl revealed the utilization of an alternative splice donor site at 33-34 bp from the boundary between exon 2 and intron 2 and the production of an aberrant mRNA with a retention of a 32 bp intronic sequence between exon 2 and exon 3 (p.(Gly72Lysfs*39)), which satisfied the condition for the occurrence of nonsense-mediated mRNA decay, as predicted by SpliceAI. This study revealed the molecular consequences of disruption of the canonical splice donor site and confirmed the clinical utility of SpliceAI.
ABSTRACT
OBJECTIVE: Pseudohypoparathyroidism type 1B (PHP1B) caused by methylation defects of differentially methylated regions (DMRs) on the GNAS locus can be categorized into groups according to etiologies and methylation defect patterns of the DMRs. The aim of this study was to clarify the clinical characteristics of each group. DESIGN: Comprehensive molecular analyses consisting of methylation, copy number, and microsatellite analyses. METHODS: Eighty-four patients with PHP1B were included in this study. We classified them into 5 groups, namely, autosomal dominant inheritance-PHP1B (Group 1, G1), sporadic-PHP1B (G2), and atypical-PHP1B (G3-G5), based on the methylation defect patterns in 4 DMRs on the GNAS locus and etiologies and evaluated the clinical findings in each group and compared them among the groups. RESULTS: G2 had the youngest age and the highest serum intact parathyroid hormone levels among the 5 groups at the time of diagnosis. The most common symptoms at the time of diagnosis were tetany in G1, and seizures or loss of consciousness in G2. Albright's hereditary osteodystrophy and PHP-suggestive features were most frequently observed in the G2 proband. Nine patients had neurodevelopmental disorders (NDs) consisting of mild to borderline intellectual disability and/or developmental delay. There were no significant correlations between the average methylation ratios of 7 CpG sites in the GNAS-A/B:TSS-DMR and hormonal and biochemical findings. CONCLUSION: This study revealed the differences in some clinical characteristics, particularly clinical features, and ages at the time of diagnosis between G2 and other groups and detailed NDs observed in some patients with PHP1B.
Subject(s)
GTP-Binding Protein alpha Subunits, Gs , Pseudohypoparathyroidism , Humans , GTP-Binding Protein alpha Subunits, Gs/genetics , Chromogranins/genetics , Pseudohypoparathyroidism/genetics , Pseudohypoparathyroidism/diagnosis , Family , DNA Methylation/geneticsABSTRACT
Although 17ß-hydroxysteroid dehydrogenase type 3 (HSD17B3) deficiency is diagnosed when a testosterone/androstenedione (T/A-dione) ratio after human chorionic gonadotropin (hCG) stimulation is below 0.8, this cut-off value is primarily based on hormonal data measured by conventional immunoassay (IA) in patients with feminized or ambiguous genitalia. We examined two 46,XY Japanese patients with undermasculinized genitalia including hypospadias (patient 1 and patient 2). Endocrine studies by IA showed well increased serum T value after hCG stimulation (2.91 ng/mL) and a high T/A-dione ratio (4.04) in patient 1 at 2 weeks of age and sufficiently elevated basal serum T value (2.60 ng/mL) in patient 2 at 1.5 months of age. Despite such partial androgen insensitivity syndrome-like findings, whole exome sequencing identified biallelic â³pathogenicâ³ or â³likely pathogenicâ³ variants in HSD17B3 (c .188 C>T:p.(Ala63Val) and c .194 C>T:p.(Ser65Leu) in patient 1, and c.139 A>G:p.(Met47Val) and c.672 + 1 G>A in patient 2) (NM_000197.2), and functional analysis revealed reduced HSD17B3 activities of the missense variants (â¼ 43% for p.Met47Val, â¼ 14% for p.Ala63Val, and â¼ 0% for p.Ser65Leu). Thus, we investigated hCG-stimulated serum steroid metabolite profiles by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in patient 1 at 7 months of age and in patient 2 at 11 months of age as well as in five control males with idiopathic micropenis aged 1 - 8 years, and found markedly high T/A-dione ratios (12.3 in patient 1 and 5.4 in patient 2) which were, however, obviously lower than those in the control boys (25.3 - 56.1) and sufficiently increased T values comparable to those of control males. The elevated T/A-dione ratios are considered be due to the residual HSD17B3 function and the measurement by LC-MS/MS. Thus, it is recommended to establish the cut-off value for the T/A-dione ratio according to the phenotypic sex reflecting the residual function and the measurement method.
Subject(s)
Androgen-Insensitivity Syndrome , Disorders of Sex Development , Humans , Male , Chromatography, Liquid , Tandem Mass Spectrometry , Testosterone , 17-Hydroxysteroid Dehydrogenases/geneticsABSTRACT
BACKGROUND: Our previous study suggested that assisted reproductive technology (ART) may be a possible risk factor for the development of epimutation-mediated imprinting disorders (epi-IDs) for mothers aged ≥ 30 years. However, whether ART or advanced parental age facilitates the development of uniparental disomy-mediated IDs (UPD-IDs) has not yet been investigated. RESULTS: We enrolled 130 patients with aneuploid UPD-IDs including various IDs confirmed by molecular studies and obtained ART data of the general population and patients with epi-IDs from a robust nationwide database and our previous report, respectively. We compared the proportion of ART-conceived livebirths and maternal childbearing age between patients with UPD-IDs and the general population or patients with epi-IDs. The proportion of ART-conceived livebirths in patients with aneuploid UPD-IDs was consistent with that in the general population of maternal age ≥ 30 years and was lower than that in the patients with epi-IDs, although there was no significant difference. The maternal childbearing age of patients with aneuploid UPD-IDs was skewed to the increased ages with several cases exceeding the 97.5th percentile of maternal childbearing age of the general population and significantly higher than that of patients with epi-IDs (P < 0.001). In addition, we compared the proportion of ART-conceived livebirths and parental age at childbirth between patients with UPD-IDs caused by aneuploid oocytes (oUPD-IDs) and that by aneuploid sperm (sUPD-IDs). Almost all ART-conceived livebirths were identified in patients with oUPD-IDs, and both maternal age and paternal age at childbirth were significantly higher in patients with oUPD-IDs than in patients with sUPD-IDs. Because maternal age and paternal age were strongly correlated (rs = 0.637, P < 0.001), higher paternal age in oUPD-IDs was explained by the higher maternal age in this group. CONCLUSIONS: Different from the case of epi-IDs, ART itself is not likely to facilitate the development of aneuploid UPD-IDs. We demonstrated that advanced maternal age can be a risk factor for the development of aneuploid UPD-IDs, particularly oUPD-IDs.
Subject(s)
Genomic Imprinting , Uniparental Disomy , Female , Humans , Male , Pregnancy , Uniparental Disomy/genetics , DNA Methylation , Semen , Aneuploidy , Risk Assessment , Mothers , Oocytes , Reproductive Techniques, Assisted/adverse effectsABSTRACT
Recent studies have indicated that heterozygous loss-of-function variants in fibroblast growth factor receptor 1 (FGFR1) are involved in the development of congenital hypogonadotropic hypogonadism and combined pituitary hormone deficiency (CPHD). We encountered a Japanese boy with short stature and pubertal failure. Endocrine studies showed GH, TSH, and LH/FSH deficiencies, and brain magnetic resonance imaging delineated hypoplastic anterior pituitary and ectopic posterior pituitary. The patient was treated with GH, l-thyroxine, and hCG/rFSH. Next-generation sequencing panel for pituitary dysfunction identified a probably weak disease-associated heterozygous missense variant in FGFR1 (NM_023110.3:c.176A>T:p.(Asp59Val)), together with a probably non-deleterious heterozygous missense variant in KISS1R (NM_032551.5:c.769G>C:p.(Val257Leu)). We also review six previously reported CHPD patients with probably deleterious FGFR1 variants. The data, in conjunction with the previously reported cases, argue for the relevance of FGFR1 variants to the development of CPHD.
ABSTRACT
Loss of methylation (LOM) at GNAS-A/B:TSS-differentially methylated regions (DMRs) in the GNAS locus is observed in pseudohypoparathyroidism type 1B (PHP1B). Many PHP1B cases are sporadic, but autosomal dominant-PHP1B has a deletion involving NESP55 expressed from the maternal allele or STX16 located upstream of the GNAS locus on the maternal allele. We report the possible first familial PHP1B cases with retrotransposon insertion in the GNAS locus on the maternal allele. To our knowledge, they are the possible first cases with imprinting disorders caused by retrotransposon insertion. The two sibling cases experienced tetany and/or cramps from school age and had hypocalcemia and an increased serum intact parathyroid hormone (PTH) level together with overweight, round face, and normal intellectual levels. Methylation analysis for DMRs in the GNAS locus showed only LOM of the GNAS-A/B:TSS-DMR. Copy number abnormalities at STX16 and the GNAS locus were not detected by array comparative genomic hybridization. Whole-genome sequencing and Sanger sequencing revealed an approximately 1000-bp SVA retrotransposon insertion upstream of the first exon of A/B on the GNAS locus in these siblings. Whole-genome methylome analysis by Enzymatic Methyl-Seq in the siblings showed normal methylation status in the region surrounding the insertion site and mild LOM of the GNAS-A/B:TSS-DMR. We conducted transcriptome analysis using mRNA from skin fibroblasts and induced pluripotent stem cells (iPSCs) derived from the siblings and detected no aberrant NESP55 transcripts. Quantitative reverse-transcriptase PCR (qRT-PCR) analysis in skin fibroblasts showed increased A/B expression in the patients and no NESP55 expression, even in a control. qRT-PCR analysis in iPSCs showed decreased NESP55 expression with normal methylation status of the GNAS-NESP:TSS-DMR in the patients. The retrotransposon insertion in the siblings likely caused decreased NESP55 expression that could lead to increased A/B expression via LOM of the GNAS-A/B:TSS-DMR, subsequent reduced Gsα expression, and finally, PHP1B development. © 2022 American Society for Bone and Mineral Research (ASBMR).
Subject(s)
Pseudohypoparathyroidism , Retroelements , Humans , Chromogranins/genetics , Chromogranins/metabolism , Comparative Genomic Hybridization , Pseudohypoparathyroidism/genetics , GTP-Binding Protein alpha Subunits, Gs/genetics , GTP-Binding Protein alpha Subunits, Gs/metabolism , RNA, Messenger/metabolism , Parathyroid Hormone/genetics , DNA-Directed RNA Polymerases/genetics , DNA-Directed RNA Polymerases/metabolism , DNA Methylation/genetics , PseudohypoparathyroidismABSTRACT
CONTEXT: Children born small-for-gestational-age with short stature (SGA-SS) is associated with (epi)genetic defects, including imprinting disorders (IDs), pathogenic copy number variants (PCNVs), and pathogenic variants of genes involved in growth. However, comprehensive studies evaluating these 3 factors are very limited. OBJECTIVE: To clarify the contribution of PCNVs and candidate pathogenic variants to SGA-SS. DESIGN: Comprehensive molecular analyses consisting of methylation analysis, copy number analysis, and multigene sequencing. METHODS: We enrolled 140 patients referred to us for genetic testing for SGA-SS. Among them, we excluded 42 patients meeting Netchine-Harbison clinical scoring system criteria for Silver-Russell syndrome and 4 patients with abnormal methylation levels of the IDs-related differentially methylated regions. Consequently, we conducted copy number analysis and multigene sequencing for 86 SGA-SS patients with sufficient sample volume. We also evaluated clinical phenotypes of patients with PCNVs or candidate pathogenic variants. RESULTS: We identified 8 (9.3%) and 11 (12.8%) patients with PCNVs and candidate pathogenic variants, respectively. According to the American College of Medical Genetics standards and guidelines, 5 variants were classified as pathogenic and the remaining 6 variants were classified as variants of unknown significance. Genetic diagnosis was made in 12 patients. All patients with PCNVs or candidate pathogenic variants did not correspond perfectly to characteristic clinical features of each specific genetic cause. CONCLUSION: We clarified the contribution of PCNVs and pathogenic variants to SGA-SS without IDs. Comprehensive molecular analyses, including copy number analysis and multigene sequencing, should be considered for patients with unknown SGA-SS etiology.
Subject(s)
Dwarfism , Silver-Russell Syndrome , DNA Copy Number Variations , Dwarfism/genetics , Genetic Testing , Humans , Infant, Newborn , Infant, Small for Gestational Age , Silver-Russell Syndrome/geneticsABSTRACT
Silver-Russel syndrome (SRS) is a representative imprinting disorder (ID) characterized by growth failure and diagnosed by clinical features. Recently, international consensus has recommended using the Netchine-Harbison clinical scoring system (NH-CSS) as clinical diagnostic criteria. Loss of methylation of H19/IGF2:intergenic differentially methylated region (H19LOM) and maternal uniparental disomy chromosome 7 (UPD(7)mat) are common etiologies of SRS; however, other IDs, pathogenic variants (PVs) of genes, and pathogenic copy number variants (PCNVs) have been reported in patients meeting NH-CSS. To clarify the frequency and clinical characteristics of each etiology, we conducted (epi)genetic analysis in 173 patients satisfying NH-CSS. H19LOM and UPD(7)mat were identified in 34.1%. PCNVs, other IDs, and PVs were in 15.0%. Patients with all six NH-CSS items were most frequently observed with H19LOM and UPD(7)mat. This study confirmed the suitability of NH-CSS as clinical diagnostic criteria, the (epi)genetic heterogeneity of SRS, and showed the necessity of further discussion regarding the "SRS spectrum".
Subject(s)
Silver-Russell Syndrome , DNA Copy Number Variations , DNA Methylation , Genomic Imprinting , Humans , Phenotype , Silver-Russell Syndrome/diagnosis , Silver-Russell Syndrome/genetics , Uniparental Disomy/geneticsABSTRACT
Infantile hemangioma (IH) is a common tumor in infants that gradually resolves and is often untreated. However, for cosmetic reasons, parents often opt for treatment. Oral propranolol, the first-line therapy for IH, is sometimes associated with several side effects, including hypotension, bradycardia, and hypoglycemia. No clinical studies on topical propranolol have been conducted using standardized procedures. We evaluated the efficacy and safety of topical propranolol in patients with IH. This multicenter, prospective pilot study was conducted from June 2019 to October 2020 and involved eight Japanese infants aged 35-150 days with proliferating IH. Patients were treated with 5% propranolol cream twice daily. We examined the efficacy rate based on central evaluation (complete or near-complete healing of the target hemangioma) at weeks 24 and 12, respectively, compared to baseline values. The efficacy rate at week 24 was 68.8% (95% confidence interval: 44.1-85.9%). The surface area, maximum diameter, and color intensity of the target IH decreased over time. Adverse event and drug-related adverse event rates were 87.5% and 0%, respectively. Propranolol cream may be effective and safe in Japanese patients with IH and may be considered a first-choice treatment for small and superficial IHs in cosmetically problematic areas.
Subject(s)
Hemangioma, Capillary , Hemangioma , Skin Neoplasms , Administration, Oral , Adrenergic beta-Antagonists/adverse effects , Hemangioma/chemically induced , Hemangioma/drug therapy , Hemangioma/pathology , Hemangioma, Capillary/chemically induced , Hemangioma, Capillary/drug therapy , Humans , Infant , Pilot Projects , Propranolol/adverse effects , Prospective Studies , Skin Neoplasms/chemically induced , Skin Neoplasms/drug therapy , Treatment OutcomeABSTRACT
BACKGROUND AND OBJECTIVES: Intravenous immunoglobulin (IVIG) therapy for acute-stage Kawasaki disease (KD) is the first-line treatment for preventing the development of coronary artery aneurysms (CAA). Corticosteroids (prednisolone) and infliximab are often used in patients at a high risk of CAA or those with CAA at diagnosis; however, there are only a few reports of non-responders to corticosteroids as an adjuvant therapy or rescue alternative to IVIG. In this study, we compared the therapeutic effects of primary and secondary prednisolone with IVIG for KD. METHODS: We established the following three protocols: A was a secondary rescue prednisolone protocol; B was no prednisolone and second-line infliximab protocol, and C was the primary prednisolone protocol. The indication for prednisolone administration was based on the following: primary prednisolone administration, Kobayashi score; and secondary administration, Shizuoka score. RESULTS: Four hundred and sixty-nine patients were enrolled in the three protocols. A comparison between primary and secondary prednisolone and IVIG, as the first-line therapy revealed that the number of first non-responders in C group was 7 (8.3%), which was significantly lower than the 50 (20.9%) in A group. There was a significant difference in the first and second non-responders among the three groups, and the number of non-responders in A group was 6 (2.5%), which was significantly lower than the 13 (9.9%) in B group (p < 0.001, by Bonferroni test). The multivariate logistic regression analysis showed that IVIG non-responders among the protocol groups had an adjusted odds ratio of 6.47. Fifteen IVIG non-responders were administered infliximab as a second-line therapy, and of them, 9 (60%) showed therapy resistance. CAA occurred in 21 patients (4.6%). There was no significant difference among each protocol group. CONCLUSIONS: The number of IVIG non-responders in the group with prednisolone administration was lower than that in the group without prednisolone administration. Secondary rescue infliximab therapy for IVIG non-responders resulted in a lower defervescence effect than the secondary rescue IVIG with prednisolone administration. Further prospective randomized studies are needed to identify factors useful for preventing IVIG non-responders and determine the optimal rescue therapy for preventing CAA.
Subject(s)
Immunoglobulins, Intravenous/administration & dosage , Infliximab/administration & dosage , Mucocutaneous Lymph Node Syndrome/drug therapy , Prednisolone/administration & dosage , Child, Preschool , Cohort Studies , Female , Humans , Immunoglobulins, Intravenous/therapeutic use , Infant , Male , Prospective StudiesABSTRACT
BACKGROUND: (Epi)genetic disorders associated with small-for-gestational-age with short stature (SGA-SS) include imprinting disorders (IDs). Silver-Russell syndrome (SRS) is a representative ID in SGA-SS and has heterogenous (epi)genetic causes. SUBJECTS AND METHODS: To clarify the contribution of IDs to SGA-SS and the molecular and phenotypic spectrum of SRS, we recruited 269 patients with SGA-SS, consisting of 103 and 166 patients referred to us for genetic testing for SGA-SS and SRS, respectively. After excluding 20 patients with structural abnormalities detected by comparative genomic hybridization analysis using catalog array, 249 patients were classified into 3 subgroups based on the Netchine-Harbison clinical scoring system (NH-CSS), SRS diagnostic criteria. We screened various IDs by methylation analysis for differentially methylated regions (DMRs) related to known IDs. We also performed clinical analysis. RESULTS: These 249 patients with SGA-SS were classified into the "SRS-compatible group" (n = 148), the "non-SRS with normocephaly or relative macrocephaly at birth group" (non-SRS group) (n = 94), or the "non-SRS with relative microcephaly at birth group" (non-SRS with microcephaly group) (n = 7). The 44.6% of patients in the "SRS-compatible group," 21.3% of patients in the "non-SRS group," and 14.3% in the "non-SRS with microcephaly group" had various IDs. Loss of methylation of the H19/IGF2:intergenic-DMR and uniparental disomy chromosome 7, being major genetic causes of SRS, was detected in 30.4% of patients in the "SRS-compatible group" and in 13.8% of patients in the "non-SRS group." CONCLUSION: We clarified the contribution of IDs as (epi)genetic causes of SGA-SS and the molecular and phenotypic spectrum of SRS. Various IDs constitute underlying factors for SGA-SS, including SRS.
Subject(s)
Dwarfism/genetics , Genomic Imprinting/genetics , Infant, Small for Gestational Age , Silver-Russell Syndrome/genetics , Abnormalities, Multiple/epidemiology , Abnormalities, Multiple/genetics , Case-Control Studies , Child, Preschool , Comparative Genomic Hybridization , DNA Mutational Analysis , Dwarfism/drug therapy , Dwarfism/epidemiology , Female , Genetic Diseases, Inborn/epidemiology , Genetic Diseases, Inborn/genetics , Human Growth Hormone/therapeutic use , Humans , Infant, Newborn , Infant, Small for Gestational Age/growth & development , Japan/epidemiology , Male , Microcephaly/complications , Microcephaly/epidemiology , Microcephaly/genetics , Phenotype , Silver-Russell Syndrome/classification , Silver-Russell Syndrome/drug therapy , Silver-Russell Syndrome/epidemiologySubject(s)
Chediak-Higashi Syndrome/diagnosis , Genetic Testing , Hyperpigmentation/diagnosis , Vesicular Transport Proteins/genetics , Biopsy , Chediak-Higashi Syndrome/genetics , Chediak-Higashi Syndrome/pathology , DNA Mutational Analysis , Epidermis/pathology , Female , Heterozygote , Humans , Hyperpigmentation/genetics , Hyperpigmentation/pathology , Infant , MutationABSTRACT
We sequenced MKRN3, the major causative gene of central precocious puberty in Western countries, in 24 Japanese or Chinese patients and examined the DNA methylation and copy-number statuses of this gene in 19 patients. We identified no (epi)genetic defects except for one previously reported mutation. These results, together with reports from Korea, indicate that MKRN3 defects are rare in Asian populations. The ethnic differences likely reflect Western country-specific founder mutations and the rarity of de novo mutations.
ABSTRACT
CONTEXT: Pseudohypoparathyroidism type I (PHP-I) is divided into PHP-Ia with Albright hereditary osteodystrophy and PHP-Ib, which usually shows no Albright hereditary osteodystrophy features. Although PHP-Ia and PHP-Ib are typically caused by genetic defects involving α subunit of the stimulatory G protein (Gsα)-coding GNAS exons and methylation defects of the GNAS differentially methylated regions (DMRs) on the maternal allele, respectively, detailed phenotypic characteristics still remains to be examined. OBJECTIVE: To clarify phenotypic characteristics according to underlying (epi)genetic causes. PATIENTS AND METHODS: We performed (epi)genotype-phenotype analysis in 69 Japanese patients with PHP-I; that is, 28 patients with genetic defects involving Gsα-coding GNAS exons (group 1) consisting of 12 patients with missense variants (subgroup A) and 16 patients with null variants (subgroup B), as well as 41 patients with methylation defects (group 2) consisting of 21 patients with broad methylation defects of the GNAS-DMRs (subgroup C) and 20 patients with an isolated A/B-DMR methylation defect accompanied by the common STX16 microdeletion (subgroup D). RESULTS: Although (epi)genotype-phenotype findings were grossly similar to those reported previously, several important findings were identified, including younger age at hypocalcemic symptoms and higher frequencies of hyperphosphatemia in subgroup C than in subgroup D, development of brachydactyly in four patients of subgroup C, predominant manifestation of subcutaneous ossification in subgroup B, higher frequency of thyrotropin resistance in group 1 than in group 2, and relatively low thyrotropin values in four patients with low T4 values and relatively low luteinizing hormone/follicle-stimulating hormone values in five adult females with ovarian dysfunction. CONCLUSION: The results imply the presence of clinical findings characteristic of each underlying cause and provide useful information on the imprinting status of Gsα.
ABSTRACT
BACKGROUND: Silver-Russell syndrome (SRS) is a rare congenital disorder characterized by pre- and postnatal growth failure and dysmorphic features. Recently, pathogenic copy number variations (PCNVs) and imprinting defects other than hypomethylation of the H19-differentially methylated region (DMR) and maternal uniparental disomy chromosome 7 have been reported in patients with the SRS phenotype. This study aimed to clarify the frequency and clinical features of patients with SRS phenotype caused by PCNVs. METHODS: We performed array comparative genomic hybridization analysis using a catalog array for 54 patients satisfying the Netchine-Harbison clinical scoring system (NH-CSS) (SRS-compatible) and for 28 patients presenting with three NH-CSS items together with triangular face and/or fifth finger clinodactyly and/or brachydactyly (SRS-like) without abnormal methylation levels of 9 DMRs related to known imprinting disorders. We then investigated the clinical features of patients with PCNVs. RESULTS: Three of the 54 SRS-compatible patients (5.6%) and 2 of the 28 SRS-like patients (7.1%) had PCNVs. We detected 3.5 Mb deletion in 4p16.3, mosaic trisomy 18, and 3.77-4.00 Mb deletion in 19q13.11-12 in SRS-compatible patients, and 1.41-1.97 Mb deletion in 7q11.23 in both SRS-like patients. Congenital heart diseases (CHDs) were identified in two patients and moderate to severe global developmental delay was observed in four patients. CONCLUSIONS: Of the patients in our study, 5.6% of SRS-compatible and 7.1% of SRS-like patients had PCNVs. All PCNVs have been previously reported for genetic causes of contiguous deletion syndromes or mosaic trisomy 18. Our study suggests patients with PCNVs, who have a phenotype resembling SRS, show a high tendency towards CHDs and/or apparent developmental delay.
Subject(s)
Comparative Genomic Hybridization/methods , DNA Copy Number Variations , Developmental Disabilities/genetics , Heart Diseases/congenital , Silver-Russell Syndrome/genetics , Child , Child, Preschool , DNA Methylation , Developmental Disabilities/diagnosis , Epigenesis, Genetic , Female , Genetic Heterogeneity , Genomic Imprinting , Heart Diseases/diagnosis , Heart Diseases/genetics , Humans , Infant , Male , Young AdultABSTRACT
PURPOSE: Recent studies have identified multilocus imprinting disturbances (MLIDs) in a subset of patients with imprinting diseases (IDs) caused by epimutations. We examined MLIDs in patients with Temple syndrome (TS14) and Kagami-Ogata syndrome (KOS14). METHODS: We studied four TS14 patients (patients 1-4) and five KOS14 patients (patients 5-9) with epimutations. We performed HumanMethylation450 BeadChip (HM450k) analysis for 43 differentially methylated regions (DMRs) (753 CpG sites) and pyrosequencing for 12 DMRs (62 CpG sites) using leukocyte genomic DNA (Leu-gDNA) of patients 1-9, and performed HM450k analysis for 43 DMRs (a slightly different set of 753 CpG sites) using buccal cell gDNA (Buc-gDNA) of patients 1, 3, and 4. We also performed mutation analysis for six causative and candidate genes for MLIDs and quantitative expression analysis using immortalized lymphocytes in MLID-positive patients. RESULTS: Methylation analysis showed hypermethylated ZDBF2-DMR and ZNF597/NAA60-DMR, hypomethylated ZNF597-DMR in both Leu-gDNA and Buc-gDNA, and hypomethylated PPIEL-DMR in Buc-gDNA of patient 1, and hypermethylated GNAS-A/B-DMR in Leu-gDNA of patient 3. No mutations were detected in the six genes for MLIDs. Expression patterns of ZDBF2, ZNF597, and GNAS-A/B were consistent with the identified MLIDs. CONCLUSION: This study indicates the presence of MLIDs in TS14 patients but not in KOS14 patients.Genet Med 19 4, 476-482.
Subject(s)
Chromosomes, Human, Pair 14/genetics , DNA Methylation , Genetic Testing/methods , Uniparental Disomy/genetics , CpG Islands , DNA-Binding Proteins/genetics , Genomic Imprinting , Humans , Mutation , Transcription Factors/geneticsABSTRACT
Germline intragenic mutations in the GNAS locus result in pseudohypoparathyroidism type 1a (PHP1a) and related conditions. Nearly half of the previously reported GNAS intragenic mutations were structural variants, including 3 tandem duplications of 12-25 bp. However, the precise mutation spectrum and the genomic basis of GNAS structural variants remain to be clarified. Here, we report a de novo 50-bp tandem duplication in GNAS (c.723_772dup50, p.Glu259Leufs*29) identified in a patient with typical clinical features of PHP1a. The mutant transcript was predicted to undergo mRNA decay or encode a nonfunctional protein. The 2 breakpoints of the duplication shared a 1-bp microhomology but were not associated with long homology or nucleotide stretches. We also examined the breakpoint structures of 3 previously reported GNAS duplications and found that 1 had a structure similar to that of our case, while the remaining 2 had blunt-ended breakpoints without microhomologies. In silico analyses revealed that the GNAS-flanking region was not enriched with repeats, palindromes, noncanonical DNA motifs, or GC content. This study expands the mutation spectrum of GNAS and provides the first indication that GNAS intragenic structural variants are induced by multiple processes, including nonhomologous end-joining and/or microhomology-mediated break-induced replication, independently of known rearrangement-inducing DNA features.
Subject(s)
Chromogranins/genetics , GTP-Binding Protein alpha Subunits, Gs/genetics , Mutation , Pseudohypoparathyroidism/genetics , Tandem Repeat Sequences , Base Sequence , Child, Preschool , DNA Mutational Analysis/methods , Humans , MaleABSTRACT
Pseudohypoparathyroidism type 1b (PHP-1b) is usually diagnosed on various symptoms of hypocalcemia. Previous studies reported a few cases of autosomal dominant pattern PHP-1b identified on familial analysis with asymptomatic hypocalcemia. Herein we report the case of a 6-year-old male patient with sporadic PHP-1b incidentally detected on preoperative examination. He had neither characteristic findings of Albright hereditary osteodystrophy nor evidence of tetany. Sporadic PHP-1b was diagnosed on the basis of clinical observation and laboratory examination. In addition, genetic testing using methylation-specific multiplex ligation-dependent probe amplification indicated broad methylation abnormalities and confirmed the sporadic form of PHP-1b. Sporadic PHP-1b might often be overlooked when diagnosis is done simply on definitive clinical features. To avoid this, DNA sequencing and methylation analysis should be performed even in the absence of definitive clinical features.
Subject(s)
Chromogranins/genetics , DNA/genetics , GTP-Binding Protein alpha Subunits, Gs/genetics , Hypocalcemia/etiology , Pseudohypoparathyroidism/diagnosis , Child , Chromogranins/metabolism , DNA Mutational Analysis , GTP-Binding Protein alpha Subunits, Gs/metabolism , Genetic Testing , Humans , Hypocalcemia/blood , Hypocalcemia/diagnosis , Male , Pseudohypoparathyroidism/complications , Pseudohypoparathyroidism/genetics , PseudohypoparathyroidismABSTRACT
NR0B1 is the causative gene for X-linked adrenal hypoplasia congenita, characterized by adrenal insufficiency, hypogonadotropic hypogonadism, and infertility. We identified an NR0B1 frameshift mutation in a boy with precocious puberty who had no signs of adrenal insufficiency. Blood examination revealed elevated testosterone levels and gonadotropin hyperresponses to gonadotropin releasing hormone (GnRH) stimulation, together with normal adrenal hormone levels. GnRH analog treatment partially ameliorated his clinical features. Molecular analysis identified a p.Glu3fsAla*16 in NR0B1. These results expand the clinical manifestations of NR0B1 mutations to include central precocious puberty without adrenal insufficiency. NR0B1 mutations likely underlie androgen overproduction via GnRH-dependent and -independent mechanisms.
Subject(s)
DAX-1 Orphan Nuclear Receptor/genetics , Frameshift Mutation/genetics , Puberty, Precocious/genetics , Androgens/blood , Child, Preschool , Gonadotropin-Releasing Hormone/analogs & derivatives , Gonadotropin-Releasing Hormone/therapeutic use , Gonadotropins/blood , Humans , Male , Puberty, Precocious/blood , Puberty, Precocious/drug therapy , Testosterone/bloodABSTRACT
CONTEXT: Pseudohypoparathyroidism type 1b (PHP-1b) results from methylation defects at the G protein stimulatory α subunit (GNAS) exon A/B-differentially methylated region (DMR). Although microduplications in the GNAS region were recently identified in two PHP-1b patients, genetic information on these patients remained fragmentary. CASE DESCRIPTION: A 20-year-old Japanese male and his mother presented with hypocalcemia and elevated blood levels of intact PTH. The proband had a maternal uncle who was previously diagnosed with PHP-1b. Methylation-specific multiplex ligation-dependent probe amplification, array-based comparative genomic hybridization, pyrosequencing, fluorescence in situ hybridization, and whole-genome sequencing were performed for this family. The proband, mother, and uncle carried maternally derived approximately 133-kb duplication-triplication-duplication rearrangements at 20q13.32 involving NESP55, NESPAS, XLαs, and exon A/B-DMR but not STX16 or the Gsα coding region. These individuals exhibited partial methylation defects of NESP55-, NESPAS-, and XLαs-DMRs, which were ascribable to the increased copy numbers of these regions retaining the maternally derived methylation pattern and loss of methylation of exon A/B-DMR, which was inexplicable by the copy-number alterations. Fusion junctions of the rearrangement resided within non-repeat sequences and were accompanied by short-templated insertions. CONCLUSIONS: Our results indicate that maternally derived copy-number gains in the GNAS region mediated by nonhomologous end-joining and/or by break-induced replication can underlie autosomal dominant PHP-1b. These rearrangements likely affect methylation of exon A/B-DMR by disconnecting or disrupting its cis-acting regulator(s). This study provides a novel example of human disorders resulting from functional disturbance in the cis-regulatory machinery of DNA methylation.