RNA-seq of cultured parasites supplemented with sodium L-lactate.
Ontology highlight
ABSTRACT:
PROVIDER: PRJEB25413 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR2367120_1.fastq.gz | Fastqsanger.gz | |||
| ERR2367120_2.fastq.gz | Fastqsanger.gz | |||
| ERR2367121_1.fastq.gz | Fastqsanger.gz | |||
| ERR2367121_2.fastq.gz | Fastqsanger.gz | |||
| ERR2367122_1.fastq.gz | Fastqsanger.gz |
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Project description:This study examined the difference in gene expression pattern between early ring stage parasites grown in medium with or without 15 mM sodium L-lactate supplementation for 5 hours using RNA-seq approach.
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Project description:L. plantarum is known to possess an L-lactate inducible lactate racemase activity (Goffin et al. 2005. J. Bacteriol. 187:6750). In the present study, microarrays were used in order to identify all genes that are up-regulated by L-lactate, but not by a racemic mixture of D- and L-lactate. A mutant of L. plantarum NCIMB8826 deficient for NAD-dependent L-lactate activity (TF101; Ferain et al. 1994. 176:596), and thus producing no L-lactate, was grown in MRS medium at 28°C until mid-exponential phase (OD600nm 0.75). The culture was then divided into 3 sub-cultures. Optically pure sodium L-lactate (200 mM) was added to the first sub-culture (TF101 + L-lac 200 mM). An equimolar mixture of sodium D- and L-lactate (100 mM each) was added to the second sub-culture (TF101 + L/D-lac 200 mM). The third sub-culture was not treated (TF101; reference sample). The three sub-cultures were further incubated at 28°C for 1h30 (a time known to be sufficient for induction of lactate racemase activity by L-lactate). Cells were harvested by centrifugation. Microarray data were used ot identify genes that are specifically induced by L-lactate (comparison of TF101 with TF101 + L-lac 200 mM), but not by DL-lactate (comparison of TF101 with T101 + L/D-lac 200 mM). There are no biological replicates.
2013-07-01 | E-GEOD-43518 | biostudies-arrayexpress
Project description:Growth to mid exponential phase in M9 minimal media supplemented with 0.5% lactate versus growth in rich LB media Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Media: M9 minimal media with 0.5% lactate Keywords: Logical Set
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Project description:Background: Previously, we could show that L-lactate affects cultured bovine granulosa cells (GC) in a specific manner driving the cells into an early pre-ovulatory phenotype. Here we studied genome wide effects in L-lactatetreated GC to further elucidate the underlying mechanisms that are responsible for the L-lactate induced transformation. Cultured estrogen producing GC treated either with L-lactate or vehicle control were subjected to mRNA microarray analysis. Results: The analysis revealed 487 differentially expressed clusters, representing 461 annotated genes. Of these, 333 (= 318 genes) were identified as up- and 154 (= 143 genes) as down-regulated. As the top up-regulated genes we detected TXNIP, H19 and AHSG as well as our previously established marker transcripts RGS2 and PTX3. The top down-regulated genes included VNN1, SLC27A2 and GFRA1, but also MYC and the GC marker transcript CYP19A1. Pathway analysis with differentially expressed genes indicated “cAMP-mediated signaling” and “Axon guidance signaling” among the most affected pathways. Furthermore, estradiol, progesterone and Vegf were identified as potential upstream regulators. An effector network analysis by IPA provided first hints that processes of “angiogenesis” and “vascularization”, but also “cell movement” appeared to be activated, whereas “organismal death” was predicted to be inhibited. Conclusions: Our data clearly show that L-lactate alters gene expression in cultured bovine GC in a broad, but obviously specific manner. Pathway analysis revealed that the mode of L-lactate action in GC initiates angiogenic processes, but also migratory events like cell movement and axonal guidance signaling, thus supporting the transformation of GC into an early luteal phenotype.
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Project description:CD8+ T cells infiltrate virtually every tissue to find and destroy infected or mutated cells. They often traverse varying oxygen levels and nutrient-deprived microenvironments. High glycolytic activity in tissues can result in extended exposure of cytotoxic T cells to the metabolite lactate. Lactate can be immunosuppressive, at least in part due to its association with tissue acidosis. We show here that the lactate anion is well tolerated by CD8+ T cells in pH neutral conditions. We describe how lactate is taken up by activated CD8+ T cells and is capable of displacing glucose as a carbon source. Activation in the presence of a pH neutral form of lactate significantly alters the CD8+ T cell transcriptome, including the expression of key effector differentiation markers such as granzyme B and interferon-gamma. Our studies reveal the novel metabolic features of lactate utilization by activated CD8+ T cells, and highlight the importance of lactate in shaping the differentiation and activity of cytotoxic T cells. Method: CD8+ T cells were purified from mouse splenocytes and activated for 72h with anti-CD3/CD28 dynabeads (Thermo Fisher, 11456D) and 10 U/ml recombinant human IL-2, in either plain media (RPMI 1640 supplemented with 10% Fetal Bovine Serum, 50 µM 2-mercaptoethanol, and 100 U/ml penicillin-streptomycin), or in the presence of 40 mM sodium lactate. After activation, cells were washed twice with PBS and cell pellets were snap-frozen in RLT Plus lysis buffer (Qiagen, #1053393). Total RNA was extracted with the Qiagen’s RNeasy kit according to the manufacturer's instructions. All samples were quality checked with Agilent Tapestation RNA screen tape. To construct libraries suitable for Illumina sequencing, the Illumina TruSeq Stranded mRNA Sample preparation protocol which includes cDNA synthesis, ligation of adapters, and amplification of indexed libraries was used. The yield and quality of the amplified libraries were analysed using Qubit by Thermo Fisher and the Agilent Tapestation. The indexed cDNA libraries were normalised and combined, and the pools were sequenced on the Nextseq 550 for a 50-cycle v2.5 sequencing run, generating 71 bp single-end reads (and 2*10 bp index reads). Fastq files were generated by demultiplexing with bcl2fastq (v2.20.0.422). STAR 2.7.5b was used to map the fastq files to the mouse reference genome (mm10/GRCm38) and to remove PCR duplicates. Uniquely mapped reads were counted in annotated exons using featureCounts v1.5.1. The gene annotations (Mus_musculus.GRCm38.99.gtf) and reference genome were obtained from Ensembl. The count table from featureCounts was imported into R/Bioconductor and differential gene expression was performed using the EdgeR package and its general linear models pipeline.
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