The galactocerebrosidase enzyme contributes to the maintenance of a functional hematopoietic stem cell niche

Visigalli, Ilaria; Ungari, Silvia; Martino, Sabata; Park, Hye-Jung; Cesani, Martina; Gentner, Bernhard; Sergi, Lucia Sergi; Orlacchio, Aldo; Naldini, Luigi; Biffi, Alessandra

doi:10.1182/blood-2009-12-256461

Cited by 53 publications

(50 citation statements)

References 34 publications

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“…Overexpression of galactocerebrosidase in HSCs is known to be toxic and results in functional impairment and apoptosis 45 . However, IDS-overexpression in HSCs did not cause these problems and we observed reconstitution of each cell lineage.…”

Section: Discussionmentioning

confidence: 99%

Hematopoietic Stem Cell Gene Therapy Corrects Neuropathic Phenotype in Murine Model of Mucopolysaccharidosis Type II

et al. 2015

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Mucopolysaccharidosis type II (MPS II) is a neuropathic lysosomal storage disorder caused by a deficiency of iduronate-2-sulfatase (IDS), which leads to the accumulation of glycosaminoglycans (GAGs). We demonstrated that biochemical alterations in the brains of MPS II mice are not corrected by bone marrow transplantation (BMT) or enzyme replacement therapy, although BMT has been shown to be effective for other neurodegenerative MPSs, such as Hurler syndrome. In this study, we demonstrated that lentiviral isogeneic hematopoietic stem cell (HSC) gene therapy corrected neuronal manifestations by ameliorating lysosomal storage and autophagic dysfunction in the brains of MPS II mice. IDS-transduced HSCs increased enzyme activity both in various visceral organs and the CNS. Decreased levels of GAGs were observed in many organs, including cerebra, after transplantation of IDS-transduced HSCs. In addition, lentiviral HSC gene therapy normalized the secondary accumulation of autophagic substrates, such as p62 and ubiquitin-protein conjugates, in cerebra. Furthermore, in contrast to naive MPS II mice, there was no deterioration of neuronal function observed in transplant recipients. These results indicated that lentiviral HSC gene therapy is a promising approach for the treatment of CNS lesions in MPS II.

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Section: Discussionmentioning

confidence: 99%

Hematopoietic Stem Cell Gene Therapy Corrects Neuropathic Phenotype in Murine Model of Mucopolysaccharidosis Type II

et al. 2015

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“…1A) (1). GALC function has also been implicated in cancer cell metabolism, primary open-angle glaucoma and the maintenance of a hematopoietic stem cell niche (2)(3)(4).…”

mentioning

confidence: 99%

Structural snapshots illustrate the catalytic cycle of β-galactocerebrosidase, the defective enzyme in Krabbe disease

Hill

Graham

Read

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Glycosphingolipids are ubiquitous components of mammalian cell membranes, and defects in their catabolism by lysosomal enzymes cause a diverse array of diseases. Deficiencies in the enzyme β-galactocerebrosidase (GALC) cause Krabbe disease, a devastating genetic disorder characterized by widespread demyelination and rapid, fatal neurodegeneration. Here, we present a series of highresolution crystal structures that illustrate key steps in the catalytic cycle of GALC. We have captured a snapshot of the short-lived enzyme-substrate complex illustrating how wild-type GALC binds a bona fide substrate. We have extensively characterized the enzyme kinetics of GALC with this substrate and shown that the enzyme is active in crystallo by determining the structure of the enzyme-product complex following extended soaking of the crystals with this same substrate. We have also determined the structure of a covalent intermediate that, together with the enzymesubstrate and enzyme-product complexes, reveals conformational changes accompanying the catalytic steps and provides key mechanistic insights, laying the foundation for future design of pharmacological chaperones.β-galactosylceramidase | lysosomal storage disease | glycosyl hydrolase | pharmacological chaperone therapy T he recycling and degradation of eukaryotic membrane components occurs in the lysosome and is essential for cellular maintenance. The molecular mechanisms of lysosomal lipid degradation are primarily informed by the study of a class of human diseases, sphingolipidoses, which are caused by inherited defects in glycosphingolipid catabolism. Krabbe disease is a devastating neurodegenerative disorder that is caused by deficiencies in the lysosomal enzyme β-galactocerebrosidase (GALC) (enzyme commission 3.2.1.46). It is essential for the catabolism of galactosphingolipids, including the principal lipid component of myelin, β-D-galactocerebroside ( Fig. 1A) (1). GALC function has also been implicated in cancer cell metabolism, primary open-angle glaucoma and the maintenance of a hematopoietic stem cell niche (2-4).GALC catalyzes the hydrolysis of β-D-galactocerebroside to β-D-galactose and ceramide, as well as the breakdown of psychosine to β-D-galactose and sphingosine. In both cases, removal of the galactosyl moiety is thought to occur via a retaining twostep glycosidic bond hydrolysis reaction (5, 6). Our recent structure of murine GALC identified two active site glutamate residues geometrically consistent with this mechanism (7). In the first step, the carboxylate group of E258 is hypothesized to perform a nucleophilic attack at ring position C 1 , forming an enzymesubstrate intermediate, releasing the first product (ceramide or sphingosine) as the leaving group. In the second step, E182 is thought to act as a general acid/base to deprotonate a water molecule, which then attacks the ring, releasing the enzyme and the second product (galactose).Defects in GALC lead to the accumulation of cytotoxic metabolites that elicit complex, and still only partially under...

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“…For instance, increased concentrations of lactosylceramide [22], a lipid raft component implicated in cell-cell and cell-matrix interactions and in signaling events linked to cell differentiation, apoptosis, development and oncogenesis [23], may contribute to tissue injury. In addition, GALC silencing may decrease intra cellular ceramide and sphingosine, thus affecting cell survival and function [18].…”

Section: Resultsmentioning

confidence: 99%

“…Alternatively, they may provide evidence for the systemic presence of diffusible toxic molecule(s) -including psychosine itself -with powerful antiangiogenic effects. These vascular defects are likely to aggravate the evolution of the diseased CNS and may impair therapeutic interventions, including repopulation of bone marrow after hematopoietic stem-cell transplantation [18].…”

Section: Galc Deficiency Leads To Systemic Vascular Defectsmentioning

confidence: 99%

The role of the endothelium in globoid-cell leukodystrophy: unexpected revelations

Presta¹,

Belleri²,

Cox³

2014

Future Neurology

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The galactocerebrosidase enzyme contributes to the maintenance of a functional hematopoietic stem cell niche

Cited by 53 publications

References 34 publications

Hematopoietic Stem Cell Gene Therapy Corrects Neuropathic Phenotype in Murine Model of Mucopolysaccharidosis Type II

Hematopoietic Stem Cell Gene Therapy Corrects Neuropathic Phenotype in Murine Model of Mucopolysaccharidosis Type II

Structural snapshots illustrate the catalytic cycle of β-galactocerebrosidase, the defective enzyme in Krabbe disease

The role of the endothelium in globoid-cell leukodystrophy: unexpected revelations

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