Bernard Conrad

Recurrent rearrangements of chromosome 1q21.1 and variable pediatric phenotypes

BACKGROUND Duplications and deletions in the human genome can cause disease or predispose persons to disease. Advances in technologies to detect these changes allow for the routine identification of submicroscopic imbalances in large numbers of patients. METHODS We tested for the presence of microdeletions and microduplications at a specific region of chromosome 1q21.1 in two groups of patients with unexplained mental retardation, autism, or congenital anomalies and in unaffected persons. RESULTS We identified 25 persons with a recurrent 1.35-Mb deletion within 1q21.1 from screening 5218 patients. The microdeletions had arisen de novo in eight patients, were inherited from a mildly affected parent in three patients, were inherited from an apparently unaffected parent in six patients, and were of unknown inheritance in eight patients. The deletion was absent in a series of 4737 control persons (P=1.1x10(-7)). We found considerable variability in the level of phenotypic expression of the microdeletion; phenotypes included mild-to-moderate mental retardation, microcephaly, cardiac abnormalities, and cataracts. The reciprocal duplication was enriched in nine children with mental retardation or autism spectrum disorder and other variable features (P=0.02). We identified three deletions and three duplications of the 1q21.1 region in an independent sample of 788 patients with mental retardation and congenital anomalies. CONCLUSIONS We have identified recurrent molecular lesions that elude syndromic classification and whose disease manifestations must be considered in a broader context of development as opposed to being assigned to a specific disease. Clinical diagnosis in patients with these lesions may be most readily achieved on the basis of genotype rather than phenotype.

A Human Endogenous Retroviral Superantigen as Candidate Autoimmune Gene in Type I Diabetes

Microbial superantigens (SAGs) have been implicated in the pathogenesis of human autoimmune diseases. Preferential expansion of the Vveta7 T cell receptor positive T cell subset in patients suffering from acute-onset type I diabetes has indicated the presence of a surface membrane-bound SAG. Here, we have isolated a novel mouse mammary tumor virus-related human endogenous retrovirus. We further show that the N-terminal moiety of the envelope gene encodes an MHC class II-dependent SAG. We propose that expression of this SAG, induced in extrapancreatic and professional antigen-presenting cells, leads to beta-cell destruction via the systemic activation of autoreactive T cells. The SAG encoded by this novel retrovirus thus constitutes a candidate autoimmune gene in type I diabetes.

A gene encoding a novel RFX-associated transactivator is mutated in the majority of MHC class II deficiency patients

Major histocompatibility class II (MHC-II) molecules are transmembrane proteins that have a central role in development and control of the immune system. They are encoded by a multi-gene family and their expression is tightly regulated. MHC-II deficiency (OMIM 209920) is an autosomal recessive immunodeficiency syndrome resulting from defects in trans-acting factors essential for transcription of MHC-II genes. There are four genetic complementation groups (A, B, C and D), reflecting the existence of four MHC-II regulators. The factors defective in groups A (CIITA), C (RFX5) and D (RFXAP) have been identified. CIITA is a non-DNA-binding co-activator that controls the cell-type specificity and inducibility of MHC-II expression. RFX5 and RFXAP are two subunits of RFX, a multi-protein complex that binds the X box motif of MHC-II promoters. Mutations in the genes encoding RFX5 (RFX5) or RFXAP (RFXAP) abolish binding of RFX (Refs 7,12). Similar to groups C and D, group B is characterized by a defect in RFX binding, and although it accounts for the majority of patients, the factor defective in group B has remained unknown. We report here the isolation of RFX by a novel single-step DNA-affinity purification approach and the identification of RFXANK, the gene encoding a third subunit of RFX. RFXANK restores MHC-II expression in cell lines from patients in group B and is mutated in these patients. RFXANK contains a protein-protein interaction region consisting of three ankyrin repeats. Its interaction with RFX5 and RFXAP is essential for binding of the RFX complex to MHC-II promoters.

Interferon-α-Induced Endogenous Superantigen: A Model Linking Environment and Autoimmunity

Abstract We earlier proposed that a human endogenous retroviral (HERV) superantigen (SAg) IDDMK 1,2 22 may cause type I diabetes by activating autoreactive T cells. Viral infections and induction of interferon-α (IFN-α) are tightly associated with the onset of autoimmunity. Here we establish a link between viral infections and IFN-α-regulated SAg expression of the polymorphic and defective HERV-K18 provirus. HERV-K18 has three alleles, IDDMK 1,2 22 and two full-length envelope genes, that all encode SAgs. Expression of HERV-K18 SAgs is inducible by IFN-α and this is sufficient to stimulate Vβ7 T cells to levels comparable to transfectants constitutively expressing HERV-K18 SAgs. Endogenous SAgs induced via IFN-α by viral infections is a novel mechanism through which environmental factors may cause disease in genetically susceptible individuals.

Potential Mechanisms of Interferon-α Induced Autoimmunity

The type I interferons (IFN) are cytokines encoded by a multigene family comprising 13 closely related IFN-A genes, and a single IFN-B gene. These factors are rapidly induced upon viral infection, and have pleiotropic effects. Historically, the induction of a cell-autonomous state of antiviral resistance, the inhibition of cell growth, and the regulation of apoptosis were appreciated first. More recently, it became generally accepted that they can regulate immune effector functions. This latter feature led them to be reconsidered as signals linking innate and adaptive immunity, and potentially orchestrating autoimmunity associated with viral infection and IFN-α therapy. Common to almost all autoimmune diseases is their polygenic inheritance, incomplete penetrance, and evidence for the role of environmental factors, particularly viral infection. In addition, they are characterized by increased numbers of circulating autoreactive T- and B-cells. Endogenously produced or therapeutically applied IFN-α can tilt the usually tightly controlled balance towards activation of these autoreactive cells via a vast array of mechanisms. The genetic susceptibility factors determine which type of autoimmunity will develop. IFN-α induces numerous target genes in antigen presenting cells (APC), such that APC are stimulated and enhance humoral autoimmunity, promote isotype switching, and potently activate autoreactive T cells. Moreover, IFN-α can synergistically amplify T cell autoreactivity by directly promoting T cell activation and keeping activated T cells alive. In essence, type I IFNs may constitute one example of genes that have been conserved because they confer dominant disease resistance, but at the same time they can trigger autoimmunity in genetically susceptible individuals.

Neuropsychological impairments and the impact of dystrophin mutations on general cognitive functioning of patients with Duchenne muscular dystrophy

Mutations in the dystrophin gene have long been recognised as a cause of mental retardation. However, for reasons that are unclear, some boys with dystrophin mutations do not show general cognitive deficits. To investigate the relationship between dystrophin mutations and cognition, the general intellectual abilities of a group of 25 boys with genetically confirmed Duchenne muscular dystrophy were evaluated. Furthermore, a subgroup underwent additional detailed neuropsychological assessment. The results showed a mean full scale intelligence quotient (IQ) of 88 (standard deviation 24). Patients performed very poorly on various neuropsychological tests, including arithmetics, digit span tests and verbal fluency. No simple relationship between dystrophin mutations and cognitive functioning could be detected. However, our analysis revealed that patients who lack the dystrophin isoform Dp140 have significantly greater cognitive problems.

Targeted Sequence Capture and High-Throughput Sequencing in the Molecular Diagnosis of Ichthyosis and Other Skin Diseases

cated that MBTPS2 is mainly expressed in the upper granular layer in normal skin, as previously shown (Aten et al., 2010); however, in OS skin, MBTPS2 was expressed throughout the epidermis (Figure 2c). There was no apparent difference in MBTPS2 localization in the skin of a KFSD patient with the p.N508S mutation (Aten et al., 2010). It is unclear why this is but it may be because of differences in processing of the mutants in the two diseases. In summary, we demonstrate a novel association between an MBTPS2 mutation and an X-linked form of OS. This expands the number of disorders linked to MBTPS2 mutations and reveals clinical heterogeneity associated with different MBTPS2 mutations. Written, informed consent was obtained from all family members or their legal guardians. This study was approved by the South East NHS Research Ethics Committee and was performed according to the Declaration of Helsinki Principles.

Unilateral focal polymicrogyria in a patient with classical Aarskog–Scott syndrome due to a novel missense mutation in an evolutionary conserved RhoGEF domain of the faciogenital dysplasia gene FGD1†

Faciogenital dysplasia or Aarskog–Scott syndrome (AAS) is an X‐linked disorder characterized by craniofacial, skeletal, and urogenital malformations and short stature. Mutations in the only known causative gene FGD1 are found in about one‐fifth of the cases with the clinical diagnosis of AAS. FGD1 is a guanine nucleotide exchange factor (GEF) that specifically activates the Rho GTPase Cdc42 via its RhoGEF domain. The Cdc42 pathway is involved in skeletal formation and multiple aspects of neuronal development. We describe a boy with typical AAS and, in addition, unilateral focal polymicrogyria (PMG), a feature hitherto unreported in AAS. Sequencing of the FGD1 gene in the index case and his mother revealed the presence of a novel mutation (1396A>G; M466V), located in the evolutionary conserved α‐helix 4 of the RhoGEF domain. M466V was not found in healthy family members, in >300 healthy controls and AAS patients, and has not been reported in the literature or mutation databases to date, indicating that this novel missense mutation causes AAS, and possibly PMG. Brain cortex malformations such as PMG could be initiated by mutations in the evolutionary conserved RhoGEF domain of FGD1, by perturbing the signaling via Rho GTPases such as Cdc42 known to cause brain malformation.

Transcriptional activation by bidirectional RNA polymerase II elongation over a silent promoter

Transcriptional interference denotes negative cis effects between promoters. Here, we show that promoters can also interact positively. Bidirectional RNA polymerase II (Pol II) elongation over the silent human endogenous retrovirus (HERV)‐K18 promoter (representative of 2.5 × 103 similar promoters genomewide) activates transcription. In tandem constructs, an upstream promoter activates HERV‐K18 transcription. This is abolished by inversion of the upstream promoter, or by insertion of a poly(A) signal between the promoters; transcription is restored by poly(A) signal mutants. TATA‐box mutants in the upstream promoter reduce HERV‐K18 transcription. Experiments with the same promoters in a convergent orientation produce similar effects. A small promoter deletion partially restores HERV‐K18 activity, consistent with activation resulting from repressor repulsion by the elongating Pol II. Transcriptional elongation over this class of intragenic promoters will generate co‐regulated sense–antisense transcripts, or, alternatively initiating transcripts, thus expanding the diversity and complexity of the human transcriptome.

Contiguous ∼16 Mb 1p36 deletion: Dominant features of classical distal 1p36 monosomy with haplo-lethality†

Monosomy 1p36 results from heterozygous deletions of the terminal short chromosome 1 arm, the most common terminal deletion in humans. The microdeletion is split in two usually non‐overlapping and clinically distinct classical distal and proximal 1p36 monosomy syndromes. Using comparative genome hybridization, MLPA and qPCR we identified the largest contiguous ∼16 Mb terminal 1p36 deletion reported to date. It covers both distal and proximal regions, causes a neonatally lethal variant with virtually exclusive features of distal 1p36 monosomy, highlighting the key importance of the gene‐rich distal region for the “compound” 1p36 phenotype and a threshold deletion‐size effect for haplo‐lethality.