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Blastocyst-derived stem cell lines were shown to self-organize into embryo-like structures in 3D cell culture environments. Here, we provide evidence that embryo-like structures can be generated solely based on transcription factor-mediated reprogramming of embryonic stem cells in a simple 3D co-culture system. Embryonic stem cells in these cultures self-organize into elongated, compartmentalized embryo-like structures reflecting aspects of the inner regions of the early post-implantation embryo. Single-cell RNA-sequencing reveals transcriptional profiles resembling epiblast, primitive-/visceral endoderm, and extraembryonic ectoderm of early murine embryos around E4.5–E5.5. In this stem cell-based embryo model, progression from rosette formation to lumenogenesis accompanied by progression from naïve- to primed pluripotency was observed within Epi-like cells. Additionally, lineage specification of primordial germ cells and distal/anterior visceral endoderm-like cells was observed in epiblast- or visceral endoderm-like compartments, respectively. The system presented in this study allows for fast and reproducible generation of embryo-like structures, providing an additional tool to study aspects of early embryogenesis. Synthetic embryo models have arisen as an approach to probe early development in vitro, facilitating the study of difficult to access stages. Here the authors present a simple system for generating embryo-like structures that resemble peri-implantation mouse embryos.

The existence of transgenerational effects of radiation exposure on the human germline remains controversial. Evidence for transgenerational biomarkers are of particular interest for populations, who have been exposed to higher than average levels of ionizing radiation (IR). This study investigated signatures of parental exposure to IR in offspring of former German radar operators and Chernobyl cleanup workers, focusing on clustered de novo mutations (cDNMs), defined as multiple de novo mutations (DNMs) within 20 bp. We recruited 110 offspring of former German radar operators, who were likely to have been exposed to IR (Radar cohort, exposure = 0–353 mGy), and reanalyzed sequencing data of 130 offspring of Chernobyl cleanup workers (CRU, exposure = 0–4080 mGy) from Yeager, et al. In addition, we analyzed whole genome trio data of 1275 offspring from unexposed families (Inova cohort). We observed on average 2.65 cDNMs (0.61 adjusted for the positive predictive value (PPV)) per offspring in the CRU cohort, 1.48 (0.34 PPV) in the Radar cohort and 0.88 (0.20 PPV) in the Inova cohort. Although under the condition that the proportion of true mutations is low in this analysis, this represented a significant increase ( ) of cDNMs counts, that scaled with paternal exposure to IR ( ). Our findings corroborate that cDNMs are a potential transgenerational biomarker of paternal IR exposure.

Among children, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections are typically mild. Here, we describe the case of a 3.5-year-old girl with an unusually severe presentation of coronavirus disease (COVID-19). The child had an autoinflammatory disorder of unknown etiology, which had been treated using prednisolone and methotrexate, and her parents were half cousins of Turkish descent. After 5 days of nonspecific viral infection symptoms, tonic-clonic seizures occurred followed by acute cardiac insufficiency, multi-organ insufficiency, and ultimate death. Trio exome sequencing identified a homozygous splice-variant in the gene TBK1, and a homozygous missense variant in the gene TNFRSF13B. Heterozygous deleterious variants in the TBK1 gene have been associated with severe COVID-19, and the variant in the TNFRSF13B gene has been associated with common variable immunodeficiency (CVID). We suggest that the identified variants, the autoinflammatory disorder and its treatment, or a combination of these factors probably predisposed to lethal COVID-19 in the present case.

Purpose To provide a detailed electroclinical description and expand the phenotype of PIGT-CDG, to perform genotype–phenotype correlation, and to investigate the onset and severity of the epilepsy associated with the different genetic subtypes of this rare disorder. Furthermore, to use computer-assisted facial gestalt analysis in PIGT-CDG and to the compare findings with other glycosylphosphatidylinositol (GPI) anchor deficiencies. Methods We evaluated 13 children from eight unrelated families with homozygous or compound heterozygous pathogenic variants in PIGT . Results All patients had hypotonia, severe developmental delay, and epilepsy. Epilepsy onset ranged from first day of life to two years of age. Severity of the seizure disorder varied from treatable seizures to severe neonatal onset epileptic encephalopathies. The facial gestalt of patients resembled that of previously published PIGT patients as they were closest to the center of the PIGT cluster in the clinical face phenotype space and were distinguishable from other gene-specific phenotypes. Conclusion We expand our knowledge of PIGT . Our cases reaffirm that the use of genetic testing is essential for diagnosis in this group of disorders. Finally, we show that computer-assisted facial gestalt analysis accurately assigned PIGT cases to the multiple congenital anomalies–hypotonia–seizures syndrome phenotypic series advocating the additional use of next-generation phenotyping technology.

A genome-wide evaluation of the effects of ionizing radiation on mutation induction in the mouse germline has identified multisite de novo mutations (MSDNs) as marker for previous exposure. Here we present the results of a small pilot study of whole genome sequencing in offspring of soldiers who served in radar units on weapon systems that were emitting high-frequency radiation. We found cases of exceptionally high MSDN rates as well as an increased mean in our cohort: While a MSDN mutation is detected in average in 1 out of 5 offspring of unexposed controls, we observed 12 MSDNs in altogether 18 offspring, including a family with 6 MSDNs in 3 offspring. Moreover, we found two translocations, also resulting from neighboring mutations. Our findings indicate that MSDNs might be suited in principle for the assessment of DNA damage from ionizing radiation also in humans. However, as exact person-related dose values in risk groups are usually not available, the interpretation of MSDNs in single families would benefit from larger molecular epidemiologic studies on this new biomarker.

Pathogenic germline mutations in PIGV lead to glycosylphosphatidylinositol biosynthesis deficiency (GPIBD). Individuals with pathogenic biallelic mutations in genes of the glycosylphosphatidylinositol (GPI)-anchor pathway exhibit cognitive impairments, motor delay, and often epilepsy. Thus far, the pathophysiology underlying the disease remains unclear, and suitable rodent models that mirror all symptoms observed in human patients have not been available. Therefore, we used CRISPR-Cas9 to introduce the most prevalent hypomorphic missense mutation in European patients, Pigv:c.1022C > A (p.A341E), at a site that is conserved in mice. Mirroring the human pathology, mutant Pigv 341E mice exhibited deficits in motor coordination, cognitive impairments, and alterations in sociability and sleep patterns, as well as increased seizure susceptibility. Furthermore, immunohistochemistry revealed reduced synaptophysin immunoreactivity in Pigv341E mice, and electrophysiology recordings showed decreased hippocampal synaptic transmission that could underlie impaired memory formation. In single-cell RNA sequencing, Pigv 341E-hippocampal cells exhibited changes in gene expression, most prominently in a subtype of microglia and subicular neurons. A significant reduction in Abl1 transcript levels in several cell clusters suggested a link to the signaling pathway of GPI-anchored ephrins. We also observed elevated levels of Hdc transcripts, which might affect histamine metabolism with consequences for circadian rhythm. This mouse model will not only open the doors to further investigation into the pathophysiology of GPIBD, but will also deepen our understanding of the role of GPI-anchor–related pathways in brain development.

Background Two interstitial microdeletions Xp11.22 including the CLCN5 and SHROOM4 genes were recently reported in a male individual affected with Dent disease, short stature, psychomotor delay and minor facial anomalies. Dent disease, characterized by a specific renal phenotype, is caused by truncating mutations of CLCN5 in the majority of affected cases. Case presentation Here, we present clinical and molecular findings in a male patient with clinical signs of Dent disease, developmental delay, short stature, microcephaly, and facial dysmorphism. Using molecular karyotyping we identified a hemizygous interstitial microdeletion Xp11.23p.11.22 of about 700 kb, which was inherited from his asymptomatic mother. Among the six deleted genes is CLCN5, which explains the renal phenotype in our patient. SHROOM4, which is partially deleted in this patient, is involved in neuronal development and was shown to be associated with X-linked intellectual disability. This is a candidate gene, the loss of which is thought to be associated with his further clinical manifestations. To rule out mutations in other genes related to intellectual disability, whole exome sequencing was performed. No other pathogenic variants that could explain the phenotypic features, were found. Conclusion We compared the clinical findings of the patient presented here with the reported case with an Xp11.22 microdeletion including CLCN5 and SHROOM4 and re-defined the phenotypic spectrum associated with this microdeletion.

Background Acromesomelic chondrodysplasias are a rare subgroup of the clinically and genetically heterogeneous osteochondrodysplasias that are characterised by abnormalities in the limb development and short stature. Here, we report a 2‐year‐old boy, offspring of consanguineous parents, with acromesomelic dysplasia and postaxial polydactyly in which exome sequencing identified a novel homozygous missense variant in BMPR1B. The patient showed skeletal malformation of both hands and feet that included complex brachydactyly with the thumbs most severely affected, postaxial polydactyly of both hands, shortened toes as well as a bilateral hypoplasia of the fibula. Methods Whole trio exome sequencing was conducted to identify potential genetic variants in the patient. Results The analysis identified the biallelic variant NM_001203.3:c.821A > G;p.(Gln274Arg) in BMPR1B, a gene encoding bone morphogenetic protein receptor 1B. Conclusion The skeletal phenotype can be brought in line with the phenotypes of previously reported cases of BMPR1B‐associated chondrodysplasias. However, the postaxial polydactyly described here is a novel clinical finding in a BMPR1B‐related case; notably, it has previously been reported in other acromesomelic dysplasia cases caused by homozygous pathogenic variants in GDF5—a gene which encodes for growth differentiation factor 5, a high‐affinity ligand to BMPR1B. Postaxial polydactyly—a novel clinical finding in a patient with a biallelic BMPR1B variant.

The case report by Mabry et al. (1970) of a family with four children with elevated tissue non-specific alkaline phosphatase, seizures and profound developmental disability, became the basis for phenotyping children with the features that became known as Mabry syndrome. Aside from improvements in the services available to patients and families, however, the diagnosis and treatment of this, and many other developmental disabilities, did not change significantly until the advent of massively parallel sequencing. As more patients with features of the Mabry syndrome were identified, exome and genome sequencing were used to identify the glycophosphatidylinositol (GPI) biosynthesis disorders (GPIBDs) as a group of congenital disorders of glycosylation (CDG). Biallelic variants of the phosphatidylinositol glycan (PIG) biosynthesis, type V (PIGV) gene identified in Mabry syndrome became evidence of the first in a phenotypic series that is numbered HPMRS1-6 in the order of discovery. HPMRS1 [MIM: 239300] is the phenotype resulting from inheritance of biallelic PIGV variants. Similarly, HPMRS2 (MIM 614749), HPMRS5 (MIM 616025) and HPMRS6 (MIM 616809) result from disruption of the PIGO, PIGW and PIGY genes expressed in the endoplasmic reticulum. By contrast, HPMRS3 (MIM 614207) and HPMRS4 (MIM 615716) result from disruption of post attachment to proteins PGAP2 (HPMRS3) and PGAP3 (HPMRS4). The GPI biosynthesis disorders (GPIBDs) are currently numbered GPIBD1-21. Working with Dr. Mabry, in 2020, we were able to use improved laboratory diagnostics to complete the molecular diagnosis of patients he had originally described in 1970. We identified biallelic variants of the PGAP2 gene in the first reported HPMRS patients. We discuss the longevity of the Mabry syndrome index patients in the context of the utility of pyridoxine treatment of seizures and evidence for putative glycolipid storage in patients with HPMRS3. From the perspective of the laboratory innovations made that enabled the identification of the HPMRS phenotype in Dr. Mabry’s patients, the need for treatment innovations that will benefit patients and families affected by developmental disabilities is clear.

Developmental and epileptic encephalopathies are characterized by infantile seizures and psychomotor delay. Glycosylphosphatidylinositol biosynthesis defects, resulting in impaired tethering of various proteins to the cell surface, represent the underlying pathology in some patients. One of the genes involved, PIGP, has recently been associated with infantile seizures and developmental delay in two siblings. Here, we report the second family with a markedly overlapping phenotype due to a homozygous frameshift mutation (c.456delA;p.Glu153Asnfs*34) in PIGP. Flow cytometry of patient granulocytes confirmed reduced expression of glycosylphosphatidylinositol‐anchored proteins as functional consequence. Our findings corroborate PIGP as a monogenic disease gene for developmental and epileptic encephalopathy.