Beta-mannosidosis in a domestic cat associated with a missense variant in MANBA.

A 6-month-old cat of unknown ancestry presented for a neurologic evaluation due to progressive motor impairment. Complete physical and neurologic examinations suggested the disorder was likely to be hereditary, although the signs were not consistent with any previously described inherited disorders in cats. Due to the progression of disease signs including severely impaired motor function and cognitive decline, the cat was euthanized at approximately 10.5 months of age. Whole genome sequence analysis identified a homozygous missense variant c.2506G > A in MANBA that predicts a p.Gly836Arg alteration in the encoded lysosomal enzyme ß -mannosidase. This variant was not present in the whole genome or whole exome sequences of any of the 424 cats represented in the 99 Lives Cat Genome dataset. ß -Mannosidase enzyme activity was undetectable in brain tissue homogenates from the affected cat, whereas α-mannosidase enzyme activities were elevated compared to an unaffected cat. Postmortem examination of brain and retinal tissues revealed massive accumulations of vacuolar inclusions in most cells, similar to those reported in animals of other species with hereditary ß -mannosidosis. Based on these findings, the cat likely suffered from ß -mannosidosis due to the abolition of ß -mannosidase activity associated with the p.Gly836Arg amino acid substitution. p.Gly836 is located in the C-terminal region of the protein and was not previously known to be involved in modulating enzyme activity. In addition to the vacuolar inclusions, some cells in the brain of the affected cat contained inclusions that exhibited lipofuscin-like autofluorescence. Electron microscopic examinations suggested these inclusions formed via an autophagy-like process.

Impact of Multiple Sclerosis Risk Polymorphism rs7665090 on MANBA Activity, Lysosomal Endocytosis, and Lymphocyte Activation.

Deficiencies in Mannosidase ß (MANBA) are associated with neurological abnormalities and recurrent infections. The single nucleotide polymorphism located in the 3'UTR of MANBA, rs7665090, was found to be associated with multiple sclerosis (MS) susceptibility. We aimed to study the functional impact of this polymorphism in lymphocytes isolated from MS patients and healthy controls. A total of 152 MS patients and 112 controls were genotyped for rs7665090. MANBA mRNA expression was quantified through qPCR and MANBA enzymatic activity was analyzed. Upon phytohemagglutinin stimulation, immune activation was evaluated by flow cytometry detection of CD69, endocytic function, and metabolic rates with Seahorse XFp Analyzer, and results were stratified by variation in rs7665090. A significantly reduced gene expression (p < 0.0001) and enzymatic activity (p = 0.018) of MANBA were found in lymphocytes of MS patients compared to those of controls. The rs7665090*GG genotype led to a significant ß-mannosidase enzymatic deficiency correlated with lysosomal dysfunction, as well as decreased metabolic activation in lymphocytes of MS patients compared to those of rs7665090*GG controls. In contrast, lymphocytes of MS patients and controls carrying the homozygous AA genotype behaved similarly. Our work provides new evidence highlighting the impact of the MS-risk variant, rs7665090, and the role of MANBA in the immunopathology of MS.

Variant c.2158-2A>G in MANBA is an important and frequent cause of hereditary hearing loss and beta-mannosidosis among the Czech and Slovak Roma population- evidence for a new ethnic-specific variant.

BACKGROUND: The Roma are a European ethnic minority threatened by several recessive diseases. Variants in MANBA cause a rare lysosomal storage disorder named beta-mannosidosis whose clinical manifestation includes deafness and mental retardation. Since 1986, only 23 patients with beta-mannosidosis and biallelic MANBA variants have been described worldwide. RESULTS: We now report on further 10 beta-mannosidosis patients of Roma origin from eight families in the Czech and Slovak Republics with hearing loss, mental retardation and homozygous pathogenic variants in MANBA. MANBA variant c.2158-2A>G screening among 345 anonymized normal hearing controls from Roma populations revealed a carrier/heterozygote frequency of 3.77%. This is about 925 times higher than the frequency of this variant in the gnomAD public database and classifies the c.2158-2A>G variant as a prevalent, ethnic-specific variant causing hearing loss and mental retardation in a homozygous state. The frequency of heterozygotes/carriers is similar to another pathogenic variant c.71G>A (p.W24*) in GJB2, regarded as the most frequent variant causing deafness in Roma populations. CONLCUSION: Beta-mannosidosis, due to a homozygous c.2158-2A>G MANBA variant, is an important and previously unknown cause of hearing loss and mental retardation among Central European Roma.

Hereditary ß-mannosidosis in a dog: Clinicopathological and molecular genetic characterization.

Hereditary ß-mannosidosis causing progressive lysosomal neuropathy and other clinical signs, has been previously described in humans, Nubian goats, and Salers cattle. Here we report the clinicopathological, metabolic, and molecular genetic features of canine beta-mannosidase (MANBA, EC 3.2.1.25) deficiency. A 1-year-old male mix-breed dog from St. Kitts was presented with progressive stumbling, weakness, and regurgitation. Vacuolated lymphocytes were observed on the blood film. Postmortem findings included marked enlargement of nerves, megaesophagus, and internal hydrocephalus. Vacuolated macrophages, neurons, and secretory epithelial cells suggested an oligosaccharide storage disease. Plasma concentration of the ß-mannosidosis specific oligosaccharide was approximately 75 fold that of controls. The plasma beta-mannosidase activity was severely reduced to ~5% of controls; five other lysosomal acid hydrolase activities were increased or within their normal reference interval. Genomic sequencing of this dog's MANBA gene identified a homozygous exonic five bp tandem duplication in the penultimate exon of the MANBA gene (c.2377_2381dupTATCA) which results in a reading frame shift, altering the subsequent amino acid sequence and creating a premature stop codon. The truncated beta-mannosidase enzyme is expected to be dysfunctional. This enzyme deficiency causes the accumulation of un-degraded oligosaccharides in cells, which affect the myelination of the peripheral and central nervous systems. This insertion was not encountered in 121 and 80-screened samples from dogs on St. Kitts (all were homozygous for wild-type) and Philadelphia region (wild-type), respectively. In conclusion, canine ß-mannosidosis has similar clinicopathological features with some human patients, but milder signs than in ruminants and more severe than in knockout mice. Hence, dogs with ß-mannosidosis could become a valuable disease model for the human disease.

Biochemical and clinical response after umbilical cord blood transplant in a boy with early childhood-onset beta-mannosidosis.

BACKGROUND: Deficiency in the enzyme ß-mannosidase was described over three decades ago. Although rare in occurrence, the presentation of childhood-onset ß-mannosidase deficiency consists of hypotonia in the newborn period followed by global development delay, behavior problems, and intellectual disability. No effective pharmacologic treatments have been available. METHODS: We report 2-year outcomes following the first umbilical cord blood transplant in a 4-year-old boy with early childhood-onset disease. RESULTS: We show restoration of leukocyte ß-mannosidase activity which remained normal at 2 years posttransplant, and a simultaneous increase in plasma ß-mannosidase activity and dramatic decrease in urine-free oligosaccharides were also observed. MRI of the brain remained stable. Neurocognitive evaluation revealed test point gains, although the magnitude of improvement was less than expected for age, causing lower IQ scores that represent a wider developmental gap between the patient and unaffected peers. CONCLUSION: Our findings suggest that hematopoietic cell transplant can correct the biochemical defect in ß-mannosidosis, although preservation of the neurocognitive trajectory may be a challenge.

ß-Mannosidosis in German Shepherd Dogs.

A neurological disease was investigated in 3 German Shepherd pups from the same litter that failed to grow normally, appeared stiff, were reluctant to move, and were deaf. They developed intermittent seizures and ataxia and had proprioceptive defects. Histopathology showed severe vacuolation of neurons, astrocytes in nervous tissue, renal tubular epithelial cells, and macrophages in nervous tissue, spleen, and liver. Vacuoles appeared empty with no storage material stained by periodic acid-Schiff (PAS) or Sudan black stains, leading to a diagnosis of a lysosomal storage disease and in particular an oligosaccharidosis. Biochemical and genomic studies showed that this was ß-mannosidosis, not previously diagnosed in dogs. A c.560T>A transition in exon 4 of the MANBA gene was found, which segregated in these and other family members in a manner consistent with it being the causative mutation of an autosomal recessive disease. This mutation led to substitution of isoleucine to asparagine at position 187 of the 885 amino acid enzyme, a change expected to have functional significance.

ß-Mannosidosis caused by a novel homozygous intragenic inverted duplication in MANBA.

ß-Mannosidosis is a lysosomal storage disorder characterized by accumulation of disaccharides due to deficiency of the lysosomal enzyme ß-mannosidase. The disease is caused by mutations in MANBA and is extremely rare in humans. Although the clinical presentation is heterogeneous, common symptoms include various degrees of developmental delay, behavioral disturbances, hearing loss, and frequent infections. We report a 15-yr-old girl presenting with mild intellectual disability, sensorineural hearing loss, severe behavioral disturbances, dysmorphic traits, and evolving angiokeratomas. Copy-number variation analysis of next-generation sequencing (NGS) data indicated increased coverage in exons 8-11 of MANBA Low ß-mannosidase activity (1 µkatal/kg protein, refv 25-40) established the diagnosis of ß-mannosidosis. Whole-genome sequencing (WGS) and cDNA analysis revealed a novel homozygous intragenic inverted duplication in MANBA, where a 13.1-kb region between introns 7 and 11 was duplicated and inserted in an inverted orientation, creating a 67-base nonduplicated gap at the insertion point. Both junctions showed microhomology regions. The inverted duplication resulted in exon skipping of exons 8-9 or 8-10. Our report highlights the importance of copy-number variation analysis of data from NGS and in particular the power of WGS in the identification and characterization of copy-number variants.

The structure of mammalian ß-mannosidase provides insight into ß-mannosidosis and nystagmus.

ß-Mannosidase is a lysosomal enzyme from the glycosyl hydrolase family 2 that cleaves the single ß(1-4)-linked mannose at the nonreducing end of N-glycosylated proteins, and plays an important role in the polysaccharide degradation pathway. Mutations in the MANBA gene, which encodes the ß-mannosidase, can lead to the lysosomal storage disease ß-mannosidosis, as well as nystagmus, an eye condition characterized by involuntary eye movements. Here, we present the first structures of a mammalian ß-mannosidase in both the apo- and mannose-bound forms. The structure is similar to previously determined ß-mannosidase structures with regard to domain organization and fold, however, there are important differences that underlie substrate specificity between species. Additionally, in contrast to most other ligand-bound ß-mannosidases from bacterial and fungal sources where bound sugars were in a boat-like conformation, we find the mannose in the chair conformation. Evaluation of known disease mutations in the MANBA gene provides insight into their impact on disease phenotypes. Together, these results will be important for the design of therapeutics for treating diseases caused by ß-mannosidase deficiency. DATABASE: Structural data are available in the Protein Data Bank under the accession numbers 6DDT and 6DDU.

Angioqueratomas (parte 2): [revisión] / Angiokeratomas (Part 2): [review]

Los angioqueratomas son lesiones vasculares relativamente infrecuentes que consisten en pápulas hiperqueratósicas rojo-violáceas. Éstas pueden ser únicas o múltiples, estar localizadas en un único segmento corporal o ser generalizadas y estar o no asociadas a otras enfermedades subyacentes. El presente trabajo abordará en profundidad los angioqueratomas generalizados complementando la primera parte en la que se trataron los angioqueratomas localizados.

Angiokeratomas are a relatively non-frequent group of vascular lesions that consist on hyperkeratotic red-violaceous papules. Lesions can be solitary or multiple, localized or generalized and may be associated or not with a systemic disease. The present work is a throughout review on generalized angiokeratomas and it is a complement of the first part in which localized angiokeratomas have been discussed.

A MANBA mutation resulting in residual beta-mannosidase activity associated with severe leukoencephalopathy: a possible pseudodeficiency variant.

BACKGROUND: beta-Mannosidosis (OMIM 248510) is a rare inborn lysosomal storage disorder caused by the deficient activity of beta-mannosidase, an enzyme encoded by a single gene (MANBA) located on chromosome 4q22-25. To date, only 20 cases of this autosomal recessive disorder have been described and 14 different MANBA mutations were incriminated in the disease. These are all null mutations or missense mutations that abolish beta-mannosidase activity. In this study, we characterized the molecular defect of a new case of beta-mannosidosis, presenting with a severe neurological disorder. METHODS: Genomic DNA was isolated from peripheral blood leukocytes of the patient to allow MANBA sequencing. The identified mutation was engineered by site-directed mutagenesis and the mutant protein was expressed through transient transfection in HEK293T cells. The beta-mannosidase expression and activity were respectively assessed by Western blot and fluorometric assay in both leukocytes and HEK293T cells. RESULTS: A missense disease-associated mutation, c.1922G>A (p.Arg641His), was identified for which the patient was homozygous. In contrast to previously described missense mutations, this substitution does not totally abrogate the enzyme activity but led to a residual activity of about 7% in the patient's leukocytes, 11% in lymphoblasts and 14% in plasma. Expression studies in transfected cells also resulted in 7% residual activity. CONCLUSION: Correlations between MANBA mutations, residual activity of beta-mannosidase and the severity of the ensuing neurological disorder are discussed. Whether the c.1922G>A mutation is responsible for a yet undescribed pseudodeficiency of beta-mannosidase is also discussed.

Beta-mannosidosis: a new cause of spinocerebellar ataxia.

Beta-mannosidosis (OMIM 248510) is an inborn lysosomal storage disorder caused by deficiency of beta-mannosidase activity. This enzyme is encoded by a single gene (MANBA), located on chromosome 4q22-25. This autosomal recessive disorder is characterized by a wide range of symptoms including mental retardation, behavioural problems, hearing loss, recurrent respiratory infections, angiokeratoma, facial dysmorphism, skeletal deformation, seizures, hypotonia, demyelinating polyneuropathy, and hepatosplenomegaly. The age of symptom onset is variable. We describe a 14-year clinical follow-up of a patient with beta-mannosidase deficiency with symptoms of mental retardation, progressive spasticity and cerebellar ataxia, a clinical spectrum that so far has never been reported in beta-mannosidosis. A novel mutation in the MANBA gene was found in our patient. Evoked potentials were in favour of a demyelinating pathology of the central nervous system. Serial MRI showed generalized cortical and subcortical atrophy in the absence of white matter changes suggesting an additional axonal pathophysiological component.

Identification of two novel beta-mannosidosis-associated sequence variants: biochemical analysis of beta-mannosidase (MANBA) missense mutations.

Beta-mannosidosis (OMIM # 248510) is an autosomal-recessive lysosomal storage disorder caused by deficiency of the lysosomal enzyme beta-mannosidase (MANBA, E.C. 3.2.1.25). The disorder has been reported in goat, cattle and man. The human disorder is rare and only 20 cases in 16 families have been reported. We have sequenced the exons and exon-intron borders in a European patient with infantile onset of beta-mannosidosis. The patient was compound heterozygous for a silent mutation (c.375A>G) in exon 3 causing alternative splicing, and a missense mutation (c.1513T>C, p.Ser505Pro) in exon 12. The alternative splicing event deleted four nucleotides from the transcript and was predicted to result in premature termination of translation. In order to evaluate the consequence of the missense mutation, we inserted the human beta-mannosidase gene into an expression vector, performed site-directed mutagenesis and expressed the normal and mutant enzyme in COS-7 cells. We also included the previously reported beta-mannosidosis-associated missense mutations c.544C>T (p.Arg182Trp) and c.1175G>A (p.Gly392Glu), which were found in patients presenting a milder phenotype. Cells transfected with the wild-type construct showed a 33-fold increase in beta-mannosidase activity compared to mock-transfected cells, whereas cells transfected with the mutant constructs showed no detectable increase in activity. We propose that the milder phenotype described in some beta-mannosidosis patients with missense mutations in the MANBA gene is not due to residual beta-mannosidase activity, but rather caused by epigenetic and/or environmental factors.

Angiokeratoma corporis diffusum in human beta-mannosidosis: Report of a new case and a novel mutation.

BACKGROUND: Human beta-mannosidosis, a rare disorder of oligosaccharide catabolism, results from a deficiency of beta-mannosidase activity. So far, mutational analysis has been performed in only seven families and revealed 11 mutations in the MANBA gene which encodes the enzyme beta-mannosidase. OBJECTIVES: We report here a 36-year-old Arab female with beta-mannosidosis who presented with mental retardation and multiple angiokeratomas. We describe in this patient a novel null mutation and review the previously reported MANBA gene mutations and their clinical correlations. METHODS: Histopathology, ultrastructural analysis, and enzyme assays were performed. Sequencing of cDNA and genomic DNA analysis was conducted in a search for a mutation in the MANBA gene. RESULTS: Histopathology of a skin biopsy specimen from the patient showed the characteristic findings of angiokeratoma. Electron microscopy showed cytoplasmic vacuolation. Enzymatic activity of beta-mannosidase in the patient's serum, leukocytes, and fibroblasts was less than 1% of control values. Sequencing of the MANBA cDNA revealed a G-->A transition in exon 6 at nucleotide position c.693, resulting in the formation of a stop codon (W231X). LIMITATIONS: Only one family was studied. CONCLUSIONS: A new case of human beta-mannosidosis is presented and the first MANBA gene mutation from Arab ancestry is reported. Reviewing the reported MANBA gene mutations does not reveal a clear genotype-phenotype correlation. The importance of angiokeratoma corporis diffusum as the clue to the diagnosis of beta-mannosidosis and other lysosomal storage diseases is emphasized.

Molecular analysis in two beta-mannosidosis patients: description of a new adult case.

beta-Mannosidosis is a lysosomal storage disorder caused by deficiency of beta-mannosidase. Thirteen families with cases of beta-mannosidosis have been described including one case previously reported by our group. We present clinical and biochemical data in a new adult case, and the molecular analyses in both this new case and the one previously reported. We detected four novel mutations: p.R182W, p.G392E, p.W466X and c.1848delA. Discrepancies between genomic DNA and cDNA results when detecting this last deletion suggested a nonsense-mediated decay cell process (NMD).

Beta-mannosidosis mice: a model for the human lysosomal storage disease.

Beta-mannosidase, a lysosomal enzyme which acts exclusively at the last step of oligosaccharide catabolism in glycoprotein degradation, functions to cleave the unique beta-linked mannose sugar found in all N-linked oligosaccharides of glycoproteins. Deficiency of this enzyme results in beta-mannosidosis, a lysosomal storage disease characterized by the cellular accumulation of small oligosaccharides. In human beta-mannosidosis, the clinical presentation is variable and can be mild, even when caused by functionally null mutations. In contrast, two existing ruminant animal models have disease that is consistent and severe. To further explore the molecular pathology of this disease and to investigate potential treatment strategies, we produced a beta-mannosidase knockout mouse. Homozygous mutant mice have undetectable beta-mannosidase activity. General appearance and growth of the knockout mice are similar to the wild-type littermates. At >1 year of age, these mice exhibit no dysmorphology or overt neurological problems. The mutant animals have consistent cytoplasmic vacuolation in the central nervous system and minimal vacuolation in most visceral organs. Thin-layer chromatography demonstrated an accumulation of disaccharide in epididymis and brain. This mouse model closely resembles human beta-mannosidosis and provides a useful tool for studying the phenotypic variation in different species and will facilitate the study of potential therapies for lysosomal storage diseases.

Beta-mannosidosis with angiokeratoma corporis diffusum.

Beta-mannosidosis is a lysosomal disorder which is caused by a deficiency of beta-mannosidase. This disorder was first described in goats. Twelve human cases have already been reported. We present the first case in Japan in whom the diagnosis was reached from angiokeratoma corporis diffusum. Futhermore, mental retardation, hearing loss, and renal failure were also detected. Pseudoxanthoma elasticum was also present, but whether it is a complication of beta-mannosidosis or not remains unknown. The activity level of beta-mannosidase in the patient's plasma was only 2% of the normal range, while that in the patient's mother was 40%. We suggest that beta-mannosidosis should be one of the differential diagnoses when lysosomal enzyme disorders are suspected in cases of angiokeratoma corporis diffusum.