Spectrum of LYST mutations in Chediak-Higashi syndrome: a report of novel variants and a comprehensive review of the literature.

INTRODUCTION: Chediak-Higashi syndrome (CHS) is a rare autosomal recessive disorder characterised by partial oculocutaneous albinism, a bleeding diathesis, immunological dysfunction and neurological impairment. Bi-allelic loss-of-function variants in LYST cause CHS. LYST encodes the lysosomal trafficking regulator, a highly conserved 429 kDa cytoplasmic protein with an unknown function. METHODS: To further our understanding of the pathogenesis of CHS, we conducted clinical evaluations on individuals with CHS enrolled in our natural history study. Using genomic DNA Sanger sequencing, we identified novel pathogenic LYST variants. Additionally, we performed an extensive literature review to curate reported LYST variants and classified these novel and reported variants according to the American College of Medical Genetics/Association for Molecular Pathology variant interpretation guidelines. RESULTS: Our investigation unveiled 11 novel pathogenic LYST variants in eight patients with a clinical diagnosis of CHS, substantiated by the presence of pathognomonic giant intracellular granules. From these novel variants, together with a comprehensive review of the literature, we compiled a total of 147 variants in LYST, including 61 frameshift variants (41%), 44 nonsense variants (30%), 23 missense variants (16%), 13 splice site variants or small genomic deletions for which the coding effect is unknown (9%), 5 in-frame variants (3%) and 1 start-loss variant (1%). Notably, a genotype-phenotype correlation emerged, whereby individuals harbouring at least one missense or in-frame variant generally resulted in milder disease, while those with two nonsense or frameshift variants generally had more severe disease. CONCLUSION: The identification of novel pathogenic LYST variants and improvements in variant classification will provide earlier diagnoses and improved care to individuals with CHS.

Diagnosis of Chediak Higashi disease in a 67-year old woman.

Chediak-Higashi disease is a rare disease caused by bi-allelic mutations in the lysosomal trafficking regulator gene, LYST. Individuals typically present in early childhood with partial oculocutaneous albinism, a bleeding diathesis, recurrent infections secondary to immune dysfunction, and risk of developing hemophagocytic lymphohistiocytosis (HLH). Without intervention, mortality is high in the first decade of life. However, some individuals with milder phenotypes have attenuated hematologic and immunologic presentations, and lower risk of HLH. Both classic and milder phenotypes develop progressive neurodegeneration in early adulthood. Here we present a remarkable patient diagnosed with Chediak-Higashi disease at age 67, many decades after the diagnosis is usually established. Diagnosis was suspected by observing the pathognomonic granules within leukocytes, and confirmed by identification of bi-allelic mutations in LYST, reduced LYST mRNA expression, enlarged lysosomes within fibroblasts, and decreased NK cell lytic activity. This case further expands the phenotype of Chediak-Higashi disease and highlights the need for increased awareness. Individuals with milder phenotypes may escape early diagnosis, but identification is important for close monitoring of potential complications, and to further our understanding of the function of LYST.

CELSR2, encoding a planar cell polarity protein, is a putative gene in Joubert syndrome with cortical heterotopia, microophthalmia, and growth hormone deficiency.

Joubert syndrome is a ciliopathy characterized by a specific constellation of central nervous system malformations that result in the pathognomonic "molar tooth sign" on imaging. More than 27 genes are associated with Joubert syndrome, but some patients do not have mutations in any of these genes. Celsr1, Celsr2, and Celsr3 are the mammalian orthologues of the drosophila planar cell polarity protein, flamingo; they play important roles in neural development, including axon guidance, neuronal migration, and cilium polarity. Here, we report bi-allelic mutations in CELSR2 in a Joubert patient with cortical heterotopia, microophthalmia, and growth hormone deficiency. © 2017 Wiley Periodicals, Inc.

Neuronal endolysosomal transport and lysosomal functionality in maintaining axonostasis.

Lysosomes serve as degradation hubs for the turnover of endocytic and autophagic cargos, which is essential for neuron function and survival. Deficits in lysosome function result in progressive neurodegeneration in most lysosomal storage disorders and contribute to the pathogenesis of aging-related neurodegenerative diseases. Given their size and highly polarized morphology, neurons face exceptional challenges in maintaining cellular homeostasis in regions far removed from the cell body where mature lysosomes are enriched. Neurons therefore require coordinated bidirectional intracellular transport to sustain efficient clearance capacity in distal axonal regions. Emerging lines of evidence have started to uncover mechanisms and signaling pathways regulating endolysosome transport and maturation to maintain axonal homeostasis, or "axonostasis," that is relevant to a range of neurologic disorders. In this review, we discuss recent advances in how axonal endolysosomal trafficking, distribution, and lysosomal functionality support neuronal health and become disrupted in several neurodegenerative diseases.

Lipid-mediated motor-adaptor sequestration impairs axonal lysosome delivery leading to autophagic stress and dystrophy in Niemann-Pick type C.

Niemann-Pick disease type C (NPC) is a neurodegenerative lysosomal storage disorder characterized by lipid accumulation in endolysosomes. An early pathologic hallmark is axonal dystrophy occurring at presymptomatic stages in NPC mice. However, the mechanisms underlying this pathologic change remain obscure. Here, we demonstrate that endocytic-autophagic organelles accumulate in NPC dystrophic axons. Using super-resolution and live-neuron imaging, we reveal that elevated cholesterol on NPC lysosome membranes sequesters kinesin-1 and Arl8 independent of SKIP and Arl8-GTPase activity, resulting in impaired lysosome transport into axons, contributing to axonal autophagosome accumulation. Pharmacologic reduction of lysosomal membrane cholesterol with 2-hydroxypropyl-ß-cyclodextrin (HPCD) or elevated Arl8b expression rescues lysosome transport, thereby reducing axonal autophagic stress and neuron death in NPC. These findings demonstrate a pathological mechanism by which altered membrane lipid composition impairs lysosome delivery into axons and provide biological insights into the translational application of HPCD in restoring axonal homeostasis at early stages of NPC disease.

Lipid-mediated impairment of axonal lysosome transport contributing to autophagic stress.

Efficient degradation of autophagic vacuoles (AVs) generated at axon terminals by mature lysosomes enriched in the cell body represents an exceptional challenge that neurons face in maintaining cellular homeostasis. Here, we discuss our recent findings revealing a lipid-mediated impairment of lysosome transport to distal axons contributing to axonal AV accumulation in the neurodegenerative lysosomal storage disorder Niemann-Pick disease type C (NPC). Using transmission electron microscopy, we observed a striking buildup of endocytic and autophagic organelles in NPC dystrophic axons, indicating defects in the clearance of organelles destined for lysosomal degradation. We further revealed that elevated cholesterol on NPC lysosome membranes abnormally sequesters motor-adaptors of axonal lysosome delivery, resulting in impaired anterograde lysosome transport into distal axons that disrupts maturation of axonal AVs during their retrograde transport route. Together, our study demonstrates a mechanism by which altered membrane lipid composition compromises axonal lysosome trafficking and positioning and shows that lowering lysosomal lipid levels rescues lysosome transport into NPC axons, thus reducing axonal autophagic stress at early stages of NPC disease.

The secret life of degradative lysosomes in axons: delivery from the soma, enzymatic activity, and local autophagic clearance.

Lysosomal degradation of protein aggregates and damaged organelles is essential for maintaining cellular homeostasis. This process in neurons is challenging due to their highly polarized architecture. While enzymatically active degradative lysosomes are enriched in the cell body, their trafficking and degradation capacity in axons remain elusive. We recently characterized the axonal delivery of degradative lysosomes by applying a set of fluorescent probes that selectively label active forms of lysosomal hydrolases on cortical neurons in microfluidic devices. We revealed that soma-derived degradative lysosomes rapidly influx into distal axons and target to autophagosomes and Parkinson disease-related SNCA/α-synuclein cargos for local degradation. Disrupting axon-targeted delivery of degradative lysosomes induces axonal autophagic stress. We demonstrate that the axon is an active compartment for local degradation, establishing a foundation for future investigations into axonal lysosome trafficking and functionality in neurodegenerative diseases and lysosomal storage disorders associated with axonal pathology and macroautophagy/autophagy stress.

Neuronal Soma-Derived Degradative Lysosomes Are Continuously Delivered to Distal Axons to Maintain Local Degradation Capacity.

Neurons face the challenge of maintaining cellular homeostasis through lysosomal degradation. While enzymatically active degradative lysosomes are enriched in the soma, their axonal trafficking and positioning and impact on axonal physiology remain elusive. Here, we characterized axon-targeted delivery of degradative lysosomes by applying fluorescent probes that selectively label active forms of lysosomal cathepsins D, B, L, and GCase. By time-lapse imaging of cortical neurons in microfluidic devices and standard dishes, we reveal that soma-derived degradative lysosomes rapidly influx into distal axons and target to autophagosomes and Parkinson disease-related α-synuclein cargos for local degradation. Impairing lysosome axonal delivery induces an aberrant accumulation of autophagosomes and α-synuclein cargos in distal axons. Our study demonstrates that the axon is an active compartment for local degradation and reveals fundamental aspects of axonal lysosomal delivery and maintenance. Our work establishes a foundation for investigations into axonal lysosome trafficking and functionality in neurodegenerative diseases.

A clinical report of Chediak-Higashi syndrome in infancy with a novel genotype from the Indian subcontinent.

Chediak-Higashi syndrome (CHS; OMIM no. 214500) is an inherited multisystem disorder presenting with hypopigmentation and a propensity to infections due to immunological dysfunction. CHS generally presents in infancy with a fatal outcome, but less severe cases can present in adulthood. Treatment with bone marrow transplantation can be life-saving, so establishing a correct diagnosis is critical. The presence of large granules on examination of peripheral blood smears is suggestive of the diagnosis of CHS in most centers. However, sequencing of the lysosomal trafficking, LYST, gene confirms the diagnosis and can provide a prognosis regarding disease severity. In the case presented here, we performed molecular testing to identify the causative mutation and tabulated published mutation data from 2009 to 2014. We found a novel frameshift mutation in our case and concluded that frameshift and nonsense are the most common types of mutation in CHS, but this may be biased due to underdiagnosis of the milder and atypical forms of the disease.