Heterozygous De Novo UBTF Gain-of-Function Variant Is Associated with Neurodegeneration in Childhood.

Ribosomal RNA (rRNA) is transcribed from rDNA by RNA polymerase I (Pol I) to produce the 45S precursor of the 28S, 5.8S, and 18S rRNA components of the ribosome. Two transcription factors have been defined for Pol I in mammals, the selectivity factor SL1, and the upstream binding transcription factor (UBF), which interacts with the upstream control element to facilitate the assembly of the transcription initiation complex including SL1 and Pol I. In seven unrelated affected individuals, all suffering from developmental regression starting at 2.5-7 years, we identified a heterozygous variant, c.628G\u3eA in UBTF, encoding p.Glu210Lys in UBF, which occurred de novo in all cases. While the levels of UBF, Ser388 phosphorylated UBF, and other Pol I-related components (POLR1E, TAF1A, and TAF1C) remained unchanged in cells of an affected individual, the variant conferred gain of function to UBF, manifesting by markedly increased UBF binding to the rDNA promoter and to the 5\u27- external transcribed spacer. This was associated with significantly increased 18S expression, and enlarged nucleoli which were reduced in number per cell. The data link neurodegeneration in childhood with altered rDNA chromatin status and rRNA metabolism

Genotype and Phenotype in 12 additional individuals with SATB2-Associated Syndrome

SATB2-associated syndrome (SAS) is a multisystemic disorder caused by alterations of the SATB2 gene. We describe the phenotype and genotype of 12 individuals with 10 unique (de novo in 11 of 11 tested) pathogenic variants (1 splice site, 5 frameshift, 3 nonsense, and 2 missense) in SATB2 and review all cases reported in the published literature caused by point alterations thus far. In the cohort here described, developmental delay (DD) with severe speech compromise, facial dysmorphism, and dental anomalies were present in all cases. We also present the third case of tibial bowing in an individual who, just as in the previous 2 individuals in the literature, also had a truncating pathogenic variant of SATB2. We explore early genotype-phenotype correlations and reaffirm the main clinical features of this recognizable syndrome: universal DD with severe speech impediment, mild facial dysmorphism, and high frequency of craniofacial anomalies, behavioral issues, and brain neuroradiographic changes. As the recently proposed surveillance guidelines for individuals with SAS are adopted by providers, further delineation of the frequency and impact of other phenotypic traits will become available. Similarly, as new cases of SAS are identified, further exploration of genotype-phenotype correlations will be possible

WNT activation by lithium abrogates TP53 mutation associated radiation resistance in medulloblastoma

TP53 mutations confer subgroup specific poor survival for children with medulloblastoma. We hypothesized that WNT activation which is associated with improved survival for such children abrogates TP53 related radioresistance and can be used to sensitize TP53 mutant tumors for radiation. We examined the subgroup-specific role of TP53 mutations in a cohort of 314 patients treated with radiation. TP53 wild-type or mutant human medulloblastoma cell-lines and normal neural stem cells were used to test radioresistance of TP53 mutations and the radiosensitizing effect of WNT activation on tumors and the developing brain. Children with WNT/TP53 mutant medulloblastoma had higher 5-year survival than those with SHH/TP53 mutant tumours (100% and 36.6% +/- 8.7%, respectively (p < 0.001)). Introduction of TP53 mutation into medulloblastoma cells induced radioresistance (survival fractions at 2Gy (SF2) of 89% +/- 2% vs. 57.4% +/- 1.8% (p < 0.01)). In contrast, beta-catenin mutation sensitized TP53 mutant cells to radiation (p < 0.05). Lithium, an activator of the WNT pathway, sensitized TP53 mutant medulloblastoma to radiation (SF2 of 43.5% +/- 1.5% in lithium treated cells vs. 56.6 +/- 3% (p < 0.01)) accompanied by increased number of.H2AX foci. Normal neural stem cells were protected from lithium induced radiation damage (SF2 of 33% +/- 8% for lithium treated cells vs. 27% +/- 3% for untreated controls (p = 0.05). Poor survival of patients with TP53 mutant medulloblastoma may be related to radiation resistance. Since constitutive activation of the WNT pathway by lithium sensitizes TP53 mutant medulloblastoma cells and protect normal neural stem cells from radiation, this oral drug may represent an attractive novel therapy for high-risk medulloblastomas.B.R.A.I.N Child Canada; Cancer Research UK; Brain Tumour Charity; Hungarian Brain Research Program [KTIA_13_NAP-A-V/3]; Janos Bolyai Scholarship of the Hungarian Academy of Sciences [TAMOP-4.2.2. A-11/1/KONV-2012-0025]; German Cancer Aid/Dr. Mildred Scheel Foundation for Cancer Research; Cure Childhood Cancer Foundation; St. Baldrick's Foundation; Southeastern Brain Tumor Foundation; Action Medical Research; [CZ.1.05/2.1.00/03.0101]; [CZ.1.07/2.3.00/20.0183

Immunoglobulin isotype switching in health and disease

Immunoglobulin isotype switching is the process where the Cmy region, initially located downstream of the VDJ region is replaced by another constant region gene. Class switch recombination takes place between switch (S) regions, composed of tandem repeats of short unit sequences, located 5 of each CH gene, except for Cdelta. The class switch machinery works by looping out the sequence between a donor and an acceptor S region and thus allows a single B cell to alter its Ig effector function while retaining the same antigen specificity. This study investigated molecular aspects of immunoglobulin isotype switching in health and disease. In vivo expression of germ-line mRNA in various organs, during B cell development and in immune diseases demonstrated a good correlation between the Ig levels and the expression of germ-line transcription. This findings suggest, that in vivo, a functional correlation exists between germ- line transcriptional activity and isotype switching. Analysis of the molecular basis of IgA deficiency demonstrated a significant decrease in the switch frequency to Salpha1 in these patients. The decrease in the number of switch fragments is consistent with a profound decrease in the Calpha membrane mRNA expression in unstimulated PBMC, as well as in the Calpha mRNA levels and IgA production in PWM-stimulated PBMC. The data indicate that the failure to switch to IgA producing B cells may be an important molecular mechanism in IgA deficiency. Molecular mechanisms leading to IgA production in IgA nephropathy patients showed an increase of both IgA1 and IgA2 subclass specific RNA with a preference for IgA1 and also an increased number of IgA1 and IgA2 producing cells. There was no expression of Ia transcripts in unstimulated PBMC from IgAN patients, but Ialpha RNA could be induced with TGF-beta1. This finding together with the elevated CD40L expression and cytokine production indicate that defect(s) in the immune regulation may play an important role in the pathogenesis of IgA nephropathy. Patients infected with Schistosomiasis mansoni and patients with atopic dermatitis showed a higher expression of Ie germ-line transcripts, which correlated with the high IgE production in these patients. Further analysis showed both mono sequential and double sequential switching to IgE in both patient groups via y isotypes. These in vivo data in humans demonstrate that switching to IgE can occur from sequential rearrangements. Immunoglobulin heavy chain transcripts of my, alpha and y isotypes were expressed, starting from 8 weeks of gestation. Characterization of the CDR3 region from clones of four different fetal livers suggested that any of the D and JH family members could be used during the early stages of B cell development. Identification of switch fragments provide direct evidence that immunoglobulin isotype switching begins already at 8 weeks of gestation in fetal liver. ISBN-91-628-2236-5 REPRO PRINT AB STOCKHOLM 199

PreBIND and Textomy – mining the biomedical literature for protein-protein interactions using a support vector machine

<p>Abstract</p> <p>Background</p> <p>The majority of experimentally verified molecular interaction and biological pathway data are present in the unstructured text of biomedical journal articles where they are inaccessible to computational methods. The Biomolecular interaction network database (BIND) seeks to capture these data in a machine-readable format. We hypothesized that the formidable task-size of backfilling the database could be reduced by using Support Vector Machine technology to first locate interaction information in the literature. We present an information extraction system that was designed to locate protein-protein interaction data in the literature and present these data to curators and the public for review and entry into BIND.</p> <p>Results</p> <p>Cross-validation estimated the support vector machine's test-set precision, accuracy and recall for classifying abstracts describing interaction information was 92%, 90% and 92% respectively. We estimated that the system would be able to recall up to 60% of all non-high throughput interactions present in another yeast-protein interaction database. Finally, this system was applied to a real-world curation problem and its use was found to reduce the task duration by 70% thus saving 176 days.</p> <p>Conclusions</p> <p>Machine learning methods are useful as tools to direct interaction and pathway database back-filling; however, this potential can only be realized if these techniques are coupled with human review and entry into a factual database such as BIND. The PreBIND system described here is available to the public at <url>http://bind.ca</url>. Current capabilities allow searching for human, mouse and yeast protein-interaction information.</p

Complex genomic rearrangements in the dystrophin gene due to replication-based mechanisms

Genomic rearrangements such as intragenic deletions and duplications are the most prevalent type of mutations in the dystrophin gene resulting in Duchenne and Becker muscular dystrophy (D/BMD). These copy number variations (CNVs) are nonrecurrent and can result from either nonhomologous end joining (NHEJ) or microhomology-mediated replication-dependent recombination (MMRDR). We characterized five DMD patients with complex genomic rearrangements using a combination of MLPA/mRNA transcript analysis/custom array comparative hybridization arrays (CGH) and breakpoint sequence analysis to investigate the mechanisms for these rearrangements. Two patients had complex rearrangements that involved microhomologies at breakpoints. One patient had a noncontiguous insertion of 89.7 kb chromosome 4 into intron 43 of DMD involving three breakpoints with 2–5 bp microhomology at the junctions. A second patient had an inversion of exon 44 flanked by intronic deletions with two breakpoint junctions each showing 2 bp microhomology. The third patient was a female with an inherited deletion of exon 47 in DMD on the maternal allele and a de novo noncontiguous duplication of exons 45–49 in DMD and MID1 on the paternal allele. The other two patients harbored complex noncontiguous duplications within the dystrophin gene. We propose a replication-based mechanisms for all five complex DMD rearrangements. This study identifies additional underlying mechanisms in DMD, and provides insight into the molecular bases of these genomic rearrangements

De novo variants in POLR3B cause ataxia, spasticity, and demyelinating neuropathy

POLR3B encodes the second-largest catalytic subunit of RNA polymerase III, an enzyme involved in transcription. Bi-allelic pathogenic variants in POLR3B are a well-established cause of hypomyelinating leukodystrophy. We describe six unrelated individuals with de novo missense variants in POLR3B and a clinical presentation substantially different from POLR3-related leukodystrophy. These individuals had afferent ataxia, spasticity, variable intellectual disability and epilepsy, and predominantly demyelinating sensory motor peripheral neuropathy. Protein modeling and proteomic analysis revealed a distinct mechanism of pathogenicity; the de novo POLR3B variants caused aberrant association of individual enzyme subunits rather than affecting overall enzyme assembly or stability. We expand the spectrum of disorders associated with pathogenic variants in POLR3B to include a de novo heterozygous POLR3B-related disorder

Erratum: De novo variants in POLR3B cause ataxia, spasticity, and demyelinating neuropathy (The American Journal of Human Genetics (2021) 108(1) (186–193), (S0002929720304389), (10.1016/j.ajhg.2020.12.002))

(The American Journal of Human Genetics 108, 186–193; January 7, 2021) In the originally published version of this article, there is a typographical error for the variant c.3137G>A (p.Arg1046His) present in subject 3. This was inadvertently described as p.Arg1064His in Figures 2, 3, and S3, which appear corrected here and have been corrected in the online version of the article. The authors apologize for the error. [Figure presented] [Figure presented] [Figure presented] [Figure presented] [Figure presented] [Figure presented