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Elucidation and Engineering of Sphingolipid Biosynthesis Pathway inYarrowia lipolyticaforProduction of Sphingoid Bases and Glycosylceramides

ABSTRACT: Elucidation and Engineering of Sphingolipid Biosynthesis Pathway inYarrowia lipolyticaforProduction of Sphingoid Bases and Glycosylceramides

PROVIDER: PRJNA1055003 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR27297102_1.fastq.gz	Fastqsanger.gz
	SRR27297102_2.fastq.gz	Fastqsanger.gz
	SRR27297103_1.fastq.gz	Fastqsanger.gz
	SRR27297103_2.fastq.gz	Fastqsanger.gz
	SRR27297104_1.fastq.gz	Fastqsanger.gz

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Elucidation and Engineering of Sphingolipid Biosynthesis Pathway inYarrowia lipolyticaforProduction of Sphingoid Bases and Glycosylceramides

Project description:This study focused on understanding and modifying the sphingolipid biosynthesis pathway in the yeast Yarrowia lipolytica. By altering specific biological processes within the yeast, the creation of new types of lipid molecules became possible. The research findings highlight the high potential of this yeast in producing non-acetylated sphingoid bases and human-type glucosylceramides, which could serve as valuable components in cosmetics or nutritional supplements.

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Project description:Treating Saccharomyces cerevisiae cells with myriocin reduces the intracellular free pool of most amino acids. We find that transcriptional changes within the first 6 hours of drug treatment seem to have little impact on free pool changes. Rather the changes are driven early by post-translational events including impaired amino acid transporter activity, due to a reduced rate of sphingolipid biosynthesis, and an increased rate of endocytosis of at least some amino acid transporters, particularly the major methionine transporter Mup1.

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Complete pathway elucidation and heterologous reconstitution of Rhodiola salidroside biosynthesis

Project description:Salidroside is a bioactive tyrosine-derived phenolic natural product found in medicinal plants under the Rhodiola genus. In addition to their roles in traditional medicine as anti-fatigue and anti-anoxia adaptogens, Rhodiola total extract and salidroside have also displayed medicinal properties as anti-cardiovascular diseases and anti-cancer agents. The resulting surge in global demand of Rhodiola plants and salidroside has driven some species close to extinction. Here, we report the full elucidation of Rhodiola salidroside biosynthetic pathway utilizing the first comprehensive transcriptomics and metabolomics datasets for Rhodiola rosea. Unlike the previously proposed pathway involving separate decarboxylation and deamination enzymatic steps from tyrosine to the key intermediate 4-hydroxyphenylacetaldehyde, Rhodiola contains a pyridoxal phosphate (PLP)-dependent 4-hydroxyphenylacetaldehyde synthase (4HPAAS) that directly converts tyrosine to 4-hydroxyphenylacetaldehyde. We further identified genes encoding the subsequent 4-hydroxyphenylacetaldehyde reductase (4HPAR) and tyrosol:UDP-glucose 8-O- glucosyltransferase (T8GT), respectively, to complete salidroside biosynthesis in Rhodiola. We show that heterologous production of salidroside can be achieved in yeast Saccharomyces cerevisiae as well as in plant Nicotiana benthamiana through transgenic expression of Rhodiola salidroside biosynthetic genes. This study provides new tools for engineering sustainable production of salidroside in heterologous hosts.

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Project description:Defects of the endosomal and lysosomal systems have been increasingly linked to adult neurodegenerative diseases1,2. Deficiencies in the retrograde trafficking pathway between endosomes and the Golgi apparatus lead to forms of Alzheimer’s and Parkinson’s diseases in the case of retromer mutations, or to the rare progressive cerebello-cerebral atrophy type 2 (PCCA2) for mutations of the Golgi-associated retrograde protein (GARP) complex3. Mutations in GARP subunits lead to abnormal cellular lipid metabolism, with accumulation of sterol esters and sphingoid bases, and concomitant lysosomal dysfunction in yeast, murine, or human cells4,5. However, whether these lipid changes cause neurodegeneration and if so, which lipids are toxic, is unknown. Here, we show that GARP defects lead to global changes of cellular protein distribution, including missorting of numerous disease-linked enzymes of sphingolipid metabolism, and to the accumulation of sphingolipid intermediates in wobbler fibroblasts. Inhibiting sphingolipid synthesis in wobbler mice, a murine model of GARP deficiency with features of amyotrophic lateral sclerosis (ALS), improved wellness scores and grip strength, and increased lifespan. Our findings indicate that regulation of sphingolipids by the GARP complex is essential for neuronal health and suggest that inhibition of sphingolipid synthesis might provide a therapeutic strategy for treatment of neurodegenerative diseases due to defects in retrograde endosome-to-Golgi membrane trafficking.

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Project description:Growing evidence correlated changes in bioactive sphingolipids, particularly sphingosine-1-phosphate (S1P) and ceramides, with coronary artery diseases. Furthermore, specific plasma ceramide species can predict major cardiovascular events. Dysfunction of the endothelium lining lesion-prone areas plays a pivotal role in the initiation and progression of atherosclerosis. Yet, how sphingolipid metabolism and signaling change and contribute to endothelial dysfunction and atherosclerosis remain poorly understood. By using a mouse model of coronary atherosclerosis, we demonstrated that hemodynamic stress induces an early metabolic rewiring of endothelial sphingolipid de novo biosynthesis favoring S1P signaling over ceramide as protective response. Furthermore, our data are paradigm shift from the current believe that ceramide accrual contributes to endothelial dysfunction. The de novo biosynthesis of sphingolipids is commenced by serine palmitoyltransferase (SPT), and is downregulated by NOGO-B, an ER membrane protein. We showed that Nogo-B is upregulated by hemodynamic stress in myocardial endothelial cells (EC) of ApoE-/- mice and is expressed in the endothelium lining coronary lesions in mice and human. We demonstrated that mice lacking Nogo-B specifically in EC (Nogo-A/BECKOApoE-/-) were resistant to coronary atherosclerosis development and progression, and mortality. Fibrous cap thickness was significantly increased in Nogo-A/BECKOApoE-/- mice and correlated with reduced necrotic core and macrophage infiltration. Mechanistically, the deletion of Nogo-B in EC sustained the rewiring of sphingolipid metabolism towards S1P, imparting an atheroprotective transcriptional signature that refrain coronary atherogenesis and its progression. These findings also set forth the foundation for sphingolipid-based therapeutics to reduce the treat this condition.

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Project description:rs07-05_sphingolipids-cold - sphingo-1 - The cold choc response seems to be partly triggered by Sphingolipid species. To date no gene response as been associated to sphingolipid signaling pathway in plant. Our aim is to identify among the cold induced genes the ones regulated by sphingolipids and to try to define a sphingolipid pathway specific group of genes. - 7ml of 5 days-old cells suspensions were incubated in presence of different sphingolipid pathway inhibitors, 30 min to 2 hours depending in the coumpound (all were resuspended in DMSO and control were done with DMSO). Then a 30 min cold choc was applied before cells were harvested and frozen in cold nitrogen. RNA were then extracted. FB1 and DMS were from Alexis , Myr from Cayman, TSP from matreya. 6 dye-swap - treated vs untreated comparison

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