Search

The liver has a high demand for phosphatidylcholine (PC), particularly in overnutrition, where reduced phospholipid levels have been implicated in the development of nonalcoholic fatty liver disease (NAFLD). Whether other pathways exist in addition to de novo PC synthesis that contribute to hepatic PC pools remains unknown. Here, we identified the lysophosphatidylcholine (LPC) transporter major facilitator superfamily domain containing 2A (Mfsd2a) as critical for maintaining hepatic phospholipid pools. Hepatic Mfsd2a expression was induced in patients having NAFLD and in mice in response to dietary fat via glucocorticoid receptor action. Mfsd2a liver- specific deficiency in mice (L2aKO) led to a robust nonalcoholic steatohepatitis-like (NASH-like) phenotype within just 2 weeks of dietary fat challenge associated with reduced hepatic phospholipids containing linoleic acid. Reducing dietary choline intake in L2aKO mice exacerbated liver pathology and deficiency of liver phospholipids containing polyunsaturated fatty acids (PUFAs). Treating hepatocytes with LPCs containing oleate and linoleate, two abundant blood-derived LPCs, specifically induced lipid droplet biogenesis and contributed to phospholipid pools, while LPC containing the omega-3 fatty acid docosahexaenoic acid (DHA) promoted lipid droplet formation and suppressed lipogenesis. This study revealed that PUFA-containing LPCs drive hepatic lipid droplet formation, suppress lipogenesis, and sustain hepatic phospholipid pools- -processes that are critical for protecting the liver from excess dietary fat., Introduction Hepatic accumulation of dietary lipids is caused by excessive uptake and impairment in lipid disposal. There are 4 major routes whereby liver maintains the balance of hepatic lipids, namely, [...]

Excess lipid accumulation is an early signature of nonalcoholic fatty liver disease (NAFLD). Although liver receptor homolog 1 (LRH-1) (encoded by NR5A2) is suppressed in human NAFLD, evidence linking this phospholipid-bound nuclear receptor to hepatic lipid metabolism is lacking. Here, we report an essential role for LRH-1 in hepatic lipid storage and phospholipid composition based on an acute hepatic KO of LRH-1 in adult mice (LRH-1AAV8-Cre mice). Indeed, LRH-1-deficient hepatocytes exhibited large cytosolic lipid droplets and increased triglycerides (TGs). LRH-1-deficient mice fed high-fat diet displayed macrovesicular steatosis, liver injury, and glucose intolerance, all of which were reversed or improved by expressing wild-type human LRH-1. While hepatic lipid synthesis decreased and lipid export remained unchanged in mutants, elevated circulating free fatty acid helped explain the lipid imbalance in LRH-1AAV8-Cre mice. Lipidomic and genomic analyses revealed that loss of LRH-1 disrupts hepatic phospholipid composition, leading to lowered arachidonoyl (AA) phospholipids due to repression of Elovl5 and Fads2, two critical genes in AA biosynthesis. Our findings reveal a role for the phospholipid sensor LRH-1 in maintaining adequate pools of hepatic AA phospholipids, further supporting the idea that phospholipid diversity is an important contributor to healthy hepatic lipid storage.

Patients with autosomal recessive microcephaly 15 caused by deficiency in the sodium-dependent lysophosphatidylcholine (LPC) transporter major facilitator superfamily domain-containing 2a (Mfsd2a) present with both microcephaly and hypomyelination, suggesting an important role for LPC uptake by oligodendrocytes in the process of myelination. Here we demonstrate that Mfsd2a is specifically expressed in oligodendrocyte precursor cells (OPCs) and is critical for oligodendrocyte development. Single-cell sequencing of the oligodendrocyte lineage revealed that OPCs from OPC-specific Mfsd2a-KO mice (2aOKO mice) underwent precocious differentiation into immature oligodendrocytes and impaired maturation into myelinating oligodendrocytes, correlating with postnatal brain hypomyelination. 2aOKO mice did not exhibit microcephaly, a finding consistent with the notion that microcephaly is the consequence of an absence of LPC uptake at the blood-brain barrier rather than a deficiency in OPCs. Lipidomic analysis showed that OPCs and iOLs from 2aOKO mice had significantly decreased levels of phospholipids containing omega-3 fatty acids, with a corresponding increase in unsaturated fatty acids, the latter being products of de novo synthesis governed by Srebp-1. RNA-Seq indicated activation of the Srebp-1 pathway and defective expression of regulators of oligodendrocyte development. Taken together, these findings indicate that the transport of LPCs by Mfsd2a in OPCs is important for maintaining OPC state to regulate postnatal brain myelination., Introduction Myelin is a conserved membrane structural component of axons derived from the plasma membrane of oligodendrocytes that is essential for saltatory axon conduction and axon energetics (1-3). Myelination in [...]

Docosahexaenoic acid (DHA) is an essential omega‐3 polyunsaturated fatty acid implicated in cognitive functions by promoting synaptic protein expression. While alterations of specific DHA‐containing phospholipids have been described in the neocortex of patients with Alzheimer's disease (AD), the status of these lipids in dementia with Lewy bodies (DLB), known to manifest aggregated α‐synuclein‐containing Lewy bodies together with variable amyloid pathology, is unclear. In this study, post‐mortem samples from the parietal cortex of 25 DLB patients and 17 age‐matched controls were processed for phospholipidomics analyses using a liquid chromatography–tandem mass spectrometry (LC–MS/MS) platform. After controlling for false discovery rate, six out of the 46 identified putative DHA‐phospholipid species were significantly decreased in DLB, with only one showing increase. Altered putative DHA‐phospholipid species were subsequently validated with further LC–MS/MS measurements. Of the DHA‐containing phospholipid (DCP) species showing decreases, five negatively correlated with soluble beta‐amyloid (Aβ42) levels, whilst three also correlated with phosphorylated α‐synuclein (all p < 0.05). Furthermore, five of these phospholipid species correlated with deficits of presynaptic Rab3A, postsynaptic neurogranin, or both (all p < 0.05). Finally, we found altered immunoreactivities of brain lysolipid DHA transporter, MFSD2A, and the fatty acid binding protein FABP5 in DLB parietal cortex. In summary, we report alterations of specific DCP species in DLB, as well as their associations with markers of neuropathological burden and synaptopathology. These results support the potential role of DHA perturbations in DLB as well as therapeutic targets. [ABSTRACT FROM AUTHOR]

Mfsd2a is the major transporter of the omega-3 fatty acid docosahexaenoic acid (DHA) into brain, with Mfsd2a-knockout mice showing reduced DHA in brain, neuronal cell loss in hippocampus and cerebellum, behavioural disorders and reduced brain size; DHA is transported in a sodium-dependent manner, in the form of lysophosphatidylcholines (LPCs) carrying long-chain fatty acids. Building the blood-brain barrier The blood-brain barrier serves a vital function in maintaining the necessary environment for brain function but is an inconvenient obstacle to brain-directed therapeutics. Two papers published in this issue of Nature report the involvement of Mfsd2a, a member of the major facilitator superfamily regarded previously as an orphan transporter, in two aspects of blood-brain barrier function. David Silver and colleagues identify Mfsd2a as the major transporter for uptake of the omega fatty acid docosahexaenoic acid (DHA) into the brain. Mfsd2a is exclusively expressed in the endothelium of the blood-brain barrier, and Mfsd2a-knockout mice have reduced levels brain DHA, neuronal loss and reduced brain size and function. Chenghua Gu and colleagues find a role for Mfsd2 as a regulator of blood-brain barrier development and function: the barrier becomes 'leaky' in Mfsd2a-deficient mice, possibly a result of increased transcellular vesicular transport. Docosahexaenoic acid (DHA) is an omega-3 fatty acid that is essential for normal brain growth and cognitive function.sup.1,2,3,4. Consistent with its importance in the brain, DHA is highly enriched in brain phospholipids.sup.5,6,7. Despite being an abundant fatty acid in brain phospholipids, DHA cannot be de novo synthesized in brain and must be imported across the blood-brain barrier, but mechanisms for DHA uptake in brain have remained enigmatic. Here we identify a member of the major facilitator superfamily--Mfsd2a (previously an orphan transporter)--as the major transporter for DHA uptake into brain. Mfsd2a is found to be expressed exclusively in endothelium of the blood-brain barrier of micro-vessels. Lipidomic analysis indicates that Mfsd2a-deficient (Mfsd2a-knockout) mice show markedly reduced levels of DHA in brain accompanied by neuronal cell loss in hippocampus and cerebellum, as well as cognitive deficits and severe anxiety, and microcephaly. Unexpectedly, cell-based studies indicate that Mfsd2a transports DHA in the form of lysophosphatidylcholine (LPC), but not unesterified fatty acid, in a sodium-dependent manner. Notably, Mfsd2a transports common plasma LPCs carrying long-chain fatty acids such LPC oleate and LPC palmitate, but not LPCs with less than a 14-carbon acyl chain. Moreover, we determine that the phosphor-zwitterionic headgroup of LPC is critical for transport. Importantly, Mfsd2a-knockout mice have markedly reduced uptake of labelled LPC DHA, and other LPCs, from plasma into brain, demonstrating that Mfsd2a is required for brain uptake of DHA. Our findings reveal an unexpected essential physiological role of plasma-derived LPCs in brain growth and function., Author(s): Long N. Nguyen [sup.1] , Dongliang Ma [sup.2] , Guanghou Shui [sup.3] , Peiyan Wong [sup.2] , Amaury Cazenave-Gassiot [sup.3] , Xiaodong Zhang [sup.2] , Markus R. Wenk [sup.3] [...]

Major Facilitator Superfamily Domain containing 2a (Mfsd2a) is a sodium-dependent lysophosphatidylcholine (LPC) transporter expressed at the blood–brain barrier that constitutes the main pathway by which the brain obtains omega-3 fatty acids, such as docosahexanoic acid. Mfsd2a deficiency in humans results in severe microcephaly, underscoring the importance of LPC transport by Mfsd2a for brain development. Biochemical studies and recent cryo-electron microscopy (cryo-EM) structures of Mfsd2a bound to LPC suggest that Mfsd2a transports LPC via an alternating access mechanism between outward-facing and inward-facing conformational states in which the LPC inverts during transport between the outer and inner leaflet of a membrane. However, direct biochemical evidence of flippase activity by Mfsd2a has not been demonstrated and it is not understood how Mfsd2a could invert LPC between the outer and inner leaflet of the membrane in a sodium-dependent manner. Here, we established a unique in vitro assay using recombinant Mfsd2a reconstituted in liposomes that exploits the ability of Mfsd2a to transport lysophosphatidylserine (LPS) coupled with a small molecule LPS binding fluorophore that allowed for monitoring of directional flipping of the LPS headgroup from the outer to the inner liposome membrane. Using this assay, we demonstrate that Mfsd2a flips LPS from the outer to the inner leaflet of a membrane bilayer in a sodium- dependent manner. Furthermore, using cryo-EM structures as guides together with mutagenesis and a cell-based transport assay, we identify amino acid residues important for Mfsd2a activity that likely constitute substrate interaction domains. These studies provide direct biochemical evidence that Mfsd2a functions as a lysolipid flippase. [ABSTRACT FROM AUTHOR]

The lysosome is central to the degradation of proteins, carbohydrates, and lipids and their salvage back to the cytosol for reutilization. Lysosomal transporters for amino acids, sugars, and cholesterol have been identified, and the metabolic fates of these molecules in the cytoplasm have been elucidated. Remarkably, it is not known whether lysosomal salvage exists for glycerophospholipids, the major constituents of cellular membranes. By using a transport assay screen against orphan lysosomal transporters, we identified the major facilitator superfamily protein Spns1 that is ubiquitously expressed in all tissues as a proton-dependent lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE) transporter, with LPC and LPE being the lysosomal breakdown products of the most abundant eukaryotic phospholipids, phosphatidylcholine and phosphatidylethanolamine, respectively. Spns1 deficiency in cells, zebrafish embryos, and mouse liver resulted in lysosomal accumulation of LPC and LPE species with pathological consequences on lysosomal function. Flux analysis using stable isotope-labeled phospholipid apolipoprotein E nanodiscs targeted to lysosomes showed that LPC was transported out of lysosomes in an Spns1-dependent manner and re-esterified back into the cytoplasmic pools of phosphatidylcholine. Our findings identify a phospholipid salvage pathway from lysosomes to the cytosol that is dependent on Spns1 and critical for maintaining normal lysosomal function. [ABSTRACT FROM AUTHOR]

Choline is an essential nutrient, but how cells acquire it was not known. Two studies by Kenny et al. and Tsuchiya et al. identified the plasma membrane proteins FLVCR1 and FLVCR2 to be the bona fide choline transporters mediating choline uptake for de novo synthesis of phospholipids in all cells. [ABSTRACT FROM AUTHOR]

Western-type diets are linked to obesity and diabetes partly because of their high-saturated fatty acid (SFA) content. We found that SFAs, but not unsaturated fatty acids (USFAs), reduced lipid droplets (LDs) within pancreatic ß cells. Mechanistically, SFAs, but not USFAs, reduced LD formation by inducing S-acylation and proteasomal, mediated degradation of fat storage-inducing transmembrane protein 2 (FIT2), an endoplasmic reticulum (ER) resident protein important for LD formation. Targeted ablation of FIT2 reduced ß cell LD numbers, lowered ß cell ATP levels, reduced Ca2+ signaling, dampened vesicle exocytosis, down-regulated ß cell transcription factors, up-regulated unfolded protein response genes, and finally, exacerbated diet-induced diabetes in mice. Subsequent mass spectrometry studies revealed increased C16:0 ceramide accumulation in islets of diet-induced diabetes mice lacking ß cell FIT2. Inhibition of ceramide synthases ameliorated the enhanced ER stress and improved insulin secretion. FIT2 was reduced in mouse diabetic islets, and separately, overexpression of FIT2 increased the number of intracellular LDs and rescued SFA-induced ER stress and apoptosis, thereby highlighting the protective role of FIT2 and LDs against ß cell lipotoxicity. [ABSTRACT FROM AUTHOR]