Lack of SPNS1 results in accumulation of lysolipids and lysosomal storage disease in mouse models.

Accumulation of sphingolipids, especially sphingosines, in the lysosomes is a key driver of several lysosomal storage diseases. The transport mechanism for sphingolipids from the lysosome remains unclear. Here, we identified SPNS1, which shares the highest homology to SPNS2, a sphingosine-1-phosphate (S1P) transporter, functions as a transporter for lysolipids from the lysosome. We generated Spns1-KO cells and mice and employed lipidomic and metabolomic approaches to reveal SPNS1 ligand identity. Global KO of Spns1 caused embryonic lethality between E12.5 and E13.5 and an accumulation of sphingosine, lysophosphatidylcholines (LPC), and lysophosphatidylethanolamines (LPE) in the fetal livers. Similarly, metabolomic analysis of livers from postnatal Spns1-KO mice presented an accumulation of sphingosines and lysoglycerophospholipids including LPC and LPE. Subsequently, biochemical assays showed that SPNS1 is required for LPC and sphingosine release from lysosomes. The accumulation of these lysolipids in the lysosomes of Spns1-KO mice affected liver functions and altered the PI3K/AKT signaling pathway. Furthermore, we identified 3 human siblings with a homozygous variant in the SPNS1 gene. These patients suffer from developmental delay, neurological impairment, intellectual disability, and cerebellar hypoplasia. These results reveal a critical role of SPNS1 as a promiscuous lysolipid transporter in the lysosomes and link its physiological functions with lysosomal storage diseases.

Ecological changes in subtidal macrobenthic communities of the Taean coast following the Hebei Spirit oil spill: A 10-year longitudinal study.

We examined long-term response (2008-2017) of the macrobenthos to the Hebei Spirit oil spill that occurred around the Taean coast, Korea, in December 2007. Oil concentrations were below the Korea/US environmental standards as of January 2008. Organic matter, chlorophyll-a, and zooplankton abundance dominated by Noctiluca scintillans were higher after the spill. Macrobenthic diversity recovered to pre-incident (2007) level in 2011. Biomass exceeded that level in 2011 and the increase prolonged for 5 years. Cross-correlation and regression analyses showed that chlorophyll-a at year t and zooplankton abundance at t-2 had a significant relationship with macrobenthic biomass at t (p < 0.05 for both), suggesting the transfer of increased organic matter (transformed from crude oil within the pelagic ecosystem) into the benthic ecosystem. Coastal wetlands around the incident area, vulnerable to oil pollution and slowly remobilizing accumulated oil, seemed to affect pelagic ecosystem processes and the unexpectedly increased and sustained biomass.

δ-Catenin engages the autophagy pathway to sculpt the developing dendritic arbor.

The development of the dendritic arbor in pyramidal neurons is critical for neural circuit function. Here, we uncovered a pathway in which δ-catenin, a component of the cadherin-catenin cell adhesion complex, promotes coordination of growth among individual dendrites and engages the autophagy mechanism to sculpt the developing dendritic arbor. Using a rat primary neuron model, time-lapse imaging, immunohistochemistry, and confocal microscopy, we found that apical and basolateral dendrites are coordinately sculpted during development. Loss or knockdown of δ-catenin uncoupled this coordination, leading to retraction of the apical dendrite without altering basolateral dendrite dynamics. Autophagy is a key cellular pathway that allows degradation of cellular components. We observed that the impairment of the dendritic arbor resulting from δ-catenin knockdown could be reversed by knockdown of autophagy-related 7 (ATG7), a component of the autophagy machinery. We propose that δ-catenin regulates the dendritic arbor by coordinating the dynamics of individual dendrites and that the autophagy mechanism may be leveraged by δ-catenin and other effectors to sculpt the developing dendritic arbor. Our findings have implications for the management of neurological disorders, such as autism and intellectual disability, that are characterized by dendritic aberrations.

Neuron-Type Specific Loss of CDKL5 Leads to Alterations in mTOR Signaling and Synaptic Markers.

CDKL5 disorder is a devastating neurodevelopmental disorder associated with epilepsy, developmental retardation, autism, and related phenotypes. Mutations in the CDKL5 gene, encoding CDKL5, have been identified in this disorder. CDKL5 is a protein with homology to the serine-threonine kinases and incompletely characterized function. We generated and validated a murine model bearing a floxed allele of CDKL5 and polyclonal antibodies to CDKL5. CDKL5 is well expressed in the cortex, hippocampus, and striatum, localized to synaptosomes and nuclei and developmentally regulated in the hippocampus. Using Cre-mediated mechanisms, we deleted CDKL5 from excitatory CaMKIIα-positive neurons or inhibitory GABAergic neurons. Our data indicate that loss of CDKL5 in excitatory neurons of the cortex or inhibitory neurons of the striatum differentially alters expression of some components of the mechanistic target of rapamycin (mTOR) signaling pathway. Further loss of CDKL5 in excitatory neurons of the cortex or inhibitory neurons of the striatum leads to alterations in levels of synaptic markers in a neuron-type specific manner. Taken together, these data support a model in which loss of CDKL5 alters mTOR signaling and synaptic compositions in a neuron-type specific manner and suggest that CDKL5 may have distinct functional roles related to cellular signaling in excitatory and inhibitory neurons. Thus, these studies provide new insights into the biology of CDKL5 and suggest that the molecular pathology in CDKL5 disorder may have distinct neuron-type specific origins and effects.

Mutations in VPS13D lead to a new recessive ataxia with spasticity and mitochondrial defects.

OBJECTIVE: To identify novel causes of recessive ataxias, including spinocerebellar ataxia with saccadic intrusions, spastic ataxias, and spastic paraplegia. METHODS: In an international collaboration, we independently performed exome sequencing in 7 families with recessive ataxia and/or spastic paraplegia. To evaluate the role of VPS13D mutations, we evaluated a Drosophila knockout model and investigated mitochondrial function in patient-derived fibroblast cultures. RESULTS: Exome sequencing identified compound heterozygous mutations in VPS13D on chromosome 1p36 in all 7 families. This included a large family with 5 affected siblings with spinocerebellar ataxia with saccadic intrusions (SCASI), or spinocerebellar ataxia, recessive, type 4 (SCAR4). Linkage to chromosome 1p36 was found in this family with a logarithm of odds score of 3.1. The phenotypic spectrum in our 12 patients was broad. Although most presented with ataxia, additional or predominant spasticity was present in 5 patients. Disease onset ranged from infancy to 39 years, and symptoms were slowly progressive and included loss of independent ambulation in 5. All but 2 patients carried a loss-of-function (nonsense or splice site) mutation on one and a missense mutation on the other allele. Knockdown or removal of Vps13D in Drosophila neurons led to changes in mitochondrial morphology and impairment in mitochondrial distribution along axons. Patient fibroblasts showed altered morphology and functionality including reduced energy production. INTERPRETATION: Our study demonstrates that compound heterozygous mutations in VPS13D cause movement disorders along the ataxia-spasticity spectrum, making VPS13D the fourth VPS13 paralog involved in neurological disorders. Ann Neurol 2018.

Differential regulation of apical-basolateral dendrite outgrowth by activity in hippocampal neurons.

Hippocampal pyramidal neurons have characteristic dendrite asymmetry, characterized by structurally and functionally distinct apical and basolateral dendrites. The ability of the neuron to generate and maintain dendrite asymmetry is vital, since synaptic inputs received are critically dependent on dendrite architecture. Little is known about the role of neuronal activity in guiding maintenance of dendrite asymmetry. Our data indicate that dendrite asymmetry is established and maintained early during development. Further, our results indicate that cell intrinsic and global alterations of neuronal activity have differential effects on net extension of apical and basolateral dendrites. Thus, apical and basolateral dendrite extension may be independently regulated by cell intrinsic and network neuronal activity during development, suggesting that individual dendrites may have autonomous control over net extension. We propose that regulated individual dendrite extension in response to cell intrinsic and neuronal network activity may allow temporal control of synapse specificity in the developing hippocampus.

Cadherins and catenins in dendrite and synapse morphogenesis.

Neurons are highly polarized specialized cells. Neuronal integrity and functional roles are critically dependent on dendritic architecture and synaptic structure, function and plasticity. The cadherins are glycosylated transmembrane proteins that form cell adhesion complexes in various tissues. They are associated with a group of cytosolic proteins, the catenins. While the functional roles of the complex have been extensively investigates in non-neuronal cells, it is becoming increasingly clear that components of the complex have critical roles in regulating dendritic and synaptic architecture, function and plasticity in neurons. Consistent with these functional roles, aberrations in components of the complex have been implicated in a variety of neurodevelopmental disorders. In this review, we discuss the roles of the classical cadherins and catenins in various aspects of dendrite and synapse architecture and function and their relevance to human neurological disorders. Cadherins are glycosylated transmembrane proteins that were initially identified as Ca(2+)-dependent cell adhesion molecules. They are present on plasma membrane of a variety of cell types from primitive metazoans to humans. In the past several years, it has become clear that in addition to providing mechanical adhesion between cells, cadherins play integral roles in tissue morphogenesis and homeostasis. The cadherin family is composed of more than 100 members and classified into several subfamilies, including classical cadherins and protocadherins. Several of these cadherin family members have been implicated in various aspects of neuronal development and function. (1-3) The classical cadherins are associated with a group of cytosolic proteins, collectively called the catenins. While the functional roles of the cadherin-catenin cell adhesion complex have been extensively investigated in epithelial cells, it is now clear that components of the complex are well expressed in central neurons at different stages during development. (4,5) Recent exciting studies have shed some light on the functional roles of cadherins and catenins in central neurons. In this review, we will provide a brief overview of the cadherin superfamily, describe cadherin family members expressed in central neurons, cadherin-catenin complexes in central neurons and then focus on role of the cadherin-catenin complex in dendrite morphogenesis and synapse morphogenesis, function and plasticity. The final section is dedicated to discussion of the emerging list of neural disorders linked to cadherins and catenins. While the roles of cadherins and catenins have been examined in several different types of neurons, the focus of this review is their role in mammalian central neurons, particularly those of the cortex and hippocampus. Accompanying this review is a series of excellent reviews targeting the roles of cadherins and protocadherins in other aspects of neural development.

δ-Catenin Regulates Spine Architecture via Cadherin and PDZ-dependent Interactions.

The ability of neurons to maintain spine architecture and modulate it in response to synaptic activity is a crucial component of the cellular machinery that underlies information storage in pyramidal neurons of the hippocampus. Here we show a critical role for δ-catenin, a component of the cadherin-catenin cell adhesion complex, in regulating spine head width and length in pyramidal neurons of the hippocampus. The loss of Ctnnd2, the gene encoding δ-catenin, has been associated with the intellectual disability observed in the cri du chat syndrome, suggesting that the functional roles of δ-catenin are vital for neuronal integrity and higher order functions. We demonstrate that loss of δ-catenin in a mouse model or knockdown of δ-catenin in pyramidal neurons compromises spine head width and length, without altering spine dynamics. This is accompanied by a reduction in the levels of synaptic N-cadherin. The ability of δ-catenin to modulate spine architecture is critically dependent on its ability to interact with cadherin and PDZ domain-containing proteins. We propose that loss of δ-catenin during development perturbs synaptic architecture leading to developmental aberrations in neural circuit formation that contribute to the learning disabilities in a mouse model and humans with cri du chat syndrome.

Neuronal and non-neuronal functions of the AP-3 sorting machinery.

Vesicles selectively exchange lipids, membrane proteins and luminal contents between organelles along the exocytic and endocytic routes. The repertoire of membrane proteins present in these vesicles is crucial for their targeting and function. Vesicle composition is determined at the time of their biogenesis by cytosolic coats. The heterotetrameric protein adaptor protein complex 3 (AP-3), a coat component, participates in the generation of a diverse group of secretory organelles and lysosome-related organelles. Recent work has shed light on the mechanisms that regulate AP-3 and the trafficking pathways controlled by this adaptor. Phenotypic analysis of organisms carrying genetic deficiencies in the AP-3 pathway highlight its role regulating the targeting of lysosomal, melanosomal and synaptic vesicle-specific membrane proteins. Synaptic vesicles from AP-3-deficient mice possess altered levels of neurotransmitter and ion transporters, molecules that ultimately define the type and amount of neurotransmitter stored in these vesicles. These findings reveal a complex picture of how AP-3 functions in multiple tissues, including neuronal tissue, and expose potential links between endocytic sorting mechanisms and the pathogenesis of psychiatric disorders such as schizophrenia.

To knockout in 129 or in C57BL/6: that is the question.

Traditionally, knockout experiments are performed in ES cells derived from the 129 mouse strain, followed by backcrossing with the more robust C57BL/6 strain. C57BL/6-derived ES cells have only occasionally been used in this process. We compared C57BL/6- with 129-derived ES cells directly and reviewed the literature. We found that, although some steps are less efficient, the advantages of C57BL/6 mice more than compensate for these drawbacks.

Mutations in a novel gene encoding a CRAL-TRIO domain cause human Cayman ataxia and ataxia/dystonia in the jittery mouse.

Cayman ataxia is a recessive congenital ataxia restricted to one area of Grand Cayman Island. Comparative mapping suggested that the locus on 19p13.3 associated with Cayman ataxia might be homologous to the locus on mouse chromosome 10 associated with the recessive ataxic mouse mutant jittery. Screening genes in the region of overlap identified mutations in a novel predicted gene in three mouse jittery alleles, including the first mouse mutation caused by an Alu-related (B1 element) insertion. We found two mutations exclusively in all individuals with Cayman ataxia. The gene ATCAY or Atcay encodes a neuron-restricted protein called caytaxin. Caytaxin contains a CRAL-TRIO motif common to proteins that bind small lipophilic molecules. Mutations in another protein containing a CRAL-TRIO domain, alpha-tocopherol transfer protein (TTPA), cause a vitamin E-responsive ataxia. Three-dimensional protein structural modeling predicts that the caytaxin ligand is more polar than vitamin E. Identification of the caytaxin ligand may help develop a therapy for Cayman ataxia.

Photoreceptor degeneration and rd1 mutation in the grizzled/mocha mouse strain.

The mocha mouse is a spontaneous mutant carrying a defective adaptor-like protein complex AP-3delta subunit. We examined retinal function and histology of the mocha mutant. We found that not only mocha homozygotes but also other littermates in the inbred strain are blind due to severe defects in both rod and cone photoreceptors on electroretinogram recordings. The functional deficit was caused by rapid, early postnatal photoreceptor degeneration. Genotyping confirmed the presence of a viral insertion of rd1 gene in the mocha strain. We conclude that rd1 allele contamination is primarily responsible for photoreceptor degeneration, and caution against behavioral tests with visual cues in the present stocks.

Genetic and phenotypic analysis of the mouse mutant mh2J, an Ap3d allele caused by IAP element insertion.

Mocha (mh), a mouse model for Hermansky-Pudlak syndrome (HPS), is characterized by platelet storage pool deficiency, pigment dilution, and deafness as well as neurological abnormalities. The trans-Golgi/endosome adaptor-related complex AP-3 is missing in mh mice owing to a deletion in the gene encoding the delta subunit. Mice mutant for a second allele, mh(2J), are as hyperactive as mh, and display both spike wave absence and generalized tonic clonic seizures, but have less coat color dilution, no hearing loss, and no hypersynchronized EEG. Here we show that the mh(2J) mutation is due to an IAP element insertion in the Ap3d gene leading to a C-terminally truncated protein. Despite correct assembly of the AP-3 complex and localization to the trans-Golgi network and endosomes, AP-3 function in neurons remains impaired. While mh mice show a severe reduction of vesicular zinc (TIMM staining) owing to mislocalization and degradation of the Zinc transporter ZnT-3, the TIMM and ZnT-3 staining patterns in mh(2J) varies, with normal expression in hippocampal mossy fibers, but abnormal patterns in neocortex. These results indicate that the N-terminal portion of the delta subunit is sufficient for AP-3 complex assembly and subcellular localization to the TGN/endosomes, while subsequent function is regulated in part by cell-specific interactions with the C-terminal portion.

Mouse models for psychiatric disorders.

Genes involved in psychiatric disorders are difficult to identify, and those that have been proposed so far remain ambiguous. As it is unrealistic to expect the development of, say, a 'schizophrenic' or 'autistic' mouse, mice are unlikely to have the same role in gene identification in psychiatry as circling mice did in the discovery of human deafness genes. However, many psychiatric disorders are associated with intermediate phenotypes that can be modeled and studied in mice, including physiological or anatomical brain changes and behavioral traits. Mouse models help to evaluate the effect of a human candidate gene mutation on an intermediate trait, and to identify new candidate genes. Once a gene or pathway has been identified, mice are also used to study the interplay of different genes in that system.