Myofibrillar myopathy (MFM) is a human disease that is characterized by focal myofibrillar destruction and pathological cytoplasmic protein aggregations. In an extended German pedigree with a novel form of MFM characterized by clinical features of a limb-girdle myopathy and morphological features of MFM, we identified a co-segregating, heterozygous nonsense mutation (8130G-->A; W2710X) in the filamin c gene (FLNC) on chromosome 7q32.1. The mutation is the first found in FLNC and is localized in the dimerization domain of filamin c. Functional studies showed that, in the truncated mutant protein, this domain has a disturbed secondary structure that leads to the inability to dimerize properly. As a consequence of this malfunction, the muscle fibers of our patients display massive cytoplasmic aggregates containing filamin c and several Z-disk-associated and sarcolemmal proteins.

Recombination rates seem to vary extensively along the human genome. Pedigree analysis suggests that rates vary by an order of magnitude when measured at the megabase scale, and at a finer scale, sperm typing studies point to the existence of recombination hotspots. These are short regions (1-2 kb) in which recombination rates are 10-1,000 times higher than the background rate. Less is known about how recombination rates change over time. Here we determined to what degree recombination rates are conserved among closely related species by estimating recombination rates from 14 Mb of linkage disequilibrium data in central chimpanzee and human populations. The results suggest that recombination hotspots are not conserved between the two species and that recombination rates in larger (50 kb) genomic regions are only weakly conserved. Therefore, the recombination landscape has changed markedly between the two species.

The limb-girdle muscular dystrophies are a group of disorders with wide genetic and clinical heterogeneity. Recently, mutations in the ANO5 gene, which encodes a putative calcium-activated chloride channel belonging to the Anoctamin family of proteins, were identified in five families with one of two previously identified disorders, limb-girdle muscular dystrophy 2L and non-dysferlin Miyoshi muscular dystrophy. We screened a candidate group of 64 patients from 59 British and German kindreds and found the truncating mutation, c.191dupA in exon 5 of ANO5 in 20 patients, homozygously in 15 and in compound heterozygosity with other ANO5 variants in the rest. An intragenic single nucleotide polymorphism and an extragenic microsatellite marker are in linkage disequilibrium with the mutation, suggesting a founder effect in the Northern European population. We have further defined the clinical phenotype of ANO5-associated muscular dystrophy. Patients show adult onset proximal lower limb weakness with highly raised serum creatine kinase values (average 4500 IU/l) and frequent muscle atrophy and asymmetry of muscle involvement. Onset varies from the early 20 s to 50 s and the weakness is generally slowly progressive, with most patients remaining ambulant for several decades. Distal presentation is much less common but a milder degree of distal lower limb weakness is often observed. Upper limb strength is only mildly affected and cardiac and respiratory function is normal. Females appear less frequently affected. In the North of England population we have identified eight patients with ANO5 mutations, suggesting a minimum prevalence of 0.27/100,000, twice as common as dysferlinopathy. We suggest that mutations in ANO5 represent a relatively common cause of adult onset muscular dystrophy with high serum creatine kinase and that mutation screening, particularly of the common mutation c.191dupA, should be an early step in the diagnostic algorithm of adult limb-girdle muscular dystrophy patients.

Highlights d Disrupting the TFAP2A regulatory domain causes branchiooculofacial syndrome (BOFS) d Enhancer disconnection leads to monoallelic and reduced TFAP2A expression d Placing enhancers and genes within a TAD is not sufficient for enhancer adoption d Patient-specific hiPSC-derived neural crest to uncover longrange pathomechanisms

In guanosine diphosphate (GDP)-mannose pyrophosphorylase A (GMPPA), we identified a homozygous nonsense mutation that segregated with achalasia and alacrima, delayed developmental milestones, and gait abnormalities in a consanguineous Pakistani pedigree. Mutations in GMPPA were subsequently found in ten additional individuals from eight independent families affected by the combination of achalasia, alacrima, and neurological deficits. This autosomal-recessive disorder shows many similarities with triple A syndrome, which is characterized by achalasia, alacrima, and variable neurological deficits in combination with adrenal insufficiency. GMPPA is a largely uncharacterized homolog of GMPPB. GMPPB catalyzes the formation of GDP-mannose, which is an essential precursor of glycan moieties of glycoproteins and glycolipids and is associated with congenital and limb-girdle muscular dystrophies with hypoglycosylation of α-dystroglycan. Surprisingly, GDP-mannose pyrophosphorylase activity was unchanged and GDP-mannose levels were strongly increased in lymphoblasts of individuals with GMPPA mutations. This suggests that GMPPA might serve as a GMPPB regulatory subunit mediating feedback inhibition of GMPPB instead of displaying catalytic enzyme activity itself. Thus, a triple-A-like syndrome can be added to the growing list of congenital disorders of glycosylation, in which dysregulation rather than mere enzyme deficiency is the basal pathophysiological mechanism.

Acquisition of a somatic nonsense mutation in the cells of the haematopoietic system might revert the cellular growth repression caused by the germline mutations (ie, second-site reversion mutations). Unexpected lack of haematological features in the two patients would be explained by the reversion mutations.

Triple A syndrome is a rare autosomal recessive disorder characterized by ACTH-resistant adrenal failure, alacrima, achalasia, and progressive neurological manifestations. The majority of cases are associated with mutations in the AAAS gene, which encodes a novel, 60-kDa WD-repeat nuclear pore protein, alacrima-achalasia-adrenal insufficiency neurological disorder (ALADIN) of unknown function. Our aim was to elucidate the functional role of ALADIN by determining its interacting protein partners using the bacterial two-hybrid (B2-H) technique. Nonidentical cDNA fragments were identified from both a HeLa S-3 cell and human cerebellar cDNA library that encoded the full-length ferritin heavy chain protein (FTH1). This interaction was confirmed by both co-immunoprecipitation and fluorescence lifetime imaging microscopy-fluorescence resonance energy transfer studies. Immunoblotting showed that fibroblasts from triple A patients (with known AAAS mutations) lack nuclear FTH1, suggesting that the nuclear translocation of FTH1 is defective. Cells transfected with FTH1 and visualized by confocal microscopy had very little nuclear FTH1, but when cotransfected with AAAS, FTH1 is readily visible in the nuclei. Therefore, FTH1 nuclear translocation is enhanced when ALADIN is coexpressed in these cells. In addition to its well known iron storage role, FTH1 has been shown to protect the nucleus from oxidative damage. Apoptosis of neuronal cells induced by hydrogen peroxide was significantly reduced by transfection of AAAS or by FTH1 or maximally by both genes together. Taken together, this work offers a plausible mechanism for the progressive clinical features of triple A syndrome.

The triple A syndrome (MIM#231550) is a rare autosomal recessive disorder characterized by adrenocorticotropic hormone (ACTH) resistant adrenal failure, achalasia, alacrima, and a variety of neurological and dermatological features. The triple A syndrome is caused by mutations in the AAAS gene, which encodes a protein known as ALADIN (ALacrima Achalasia aDrenal Insufficiency Neurologic disorder). ALADIN is a new WD-repeat protein that has no significant homology to any previously identified WD-repeat protein. It has been shown that it colocalizes with nuclear pore complexes (NPCs), a finding that strongly suggests an involvement of ALADIN in nucleocytoplasmic transport. An investigation of 110 families with triple A syndrome disclosed mutation hot spots including Q15K (exon 1), and S293P (exon 8), which occur in 17 and 21 families from different geographical regions, respectively. The variable phenotype of all patients cannot be correlated with the localization and the nature of the ALADIN mutations. Thus, modifying genes/factors may be involved in the progression of this neurodegenerative disease. The lack of AAAS mutations in eight patients and negative linkage to chromosome 12q13 in three families are suggestive of genetic heterogeneity. To examine the cellular localization of ALADIN mutants causing triple A syndrome, we investigated nine different ALADIN-mutants: 2 nonsense (W84X, Q456X), 2 frameshift (F157fsX171, G397fsX414) and 5 point mutations (Q15K, L25P, H160R, S263P, L381R) by transfection experiments with green fluorescence protein. Mutants were predominantly localized in the cytoplasm, but also found in the nucleus indicating that ALADIN is essential for NPC targeting. To investigate physiological functions of ALADIN in vivo, we generated and analysed Aaas-/- knockout mice by homologous recombination in embryonic stem cells. Surprisingly, required animals lack any gross abnormality in adrenal and nervous system function. Further studies have to investigate the role of ALADIN at NPCs and to identify interacting proteins. Functional analyses of ALADIN may permit further understanding of its role for adrenocortical function and neurodevelopment.