Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcriptional response of the marine pennate diatom Phaeodactylum tricornutum to Fe limitation

ABSTRACT: Diatoms, which are responsible for up to 40% of the 45 to 50 billion metric tons of organic carbon production each year in the sea, are particularly sensitive to Fe stress. Here we describe the transcriptional response of the pennate diatom Phaeodactylum tricornutum to Fe limitation using a partial genome microarray based on EST and genome sequence data. Processes carried out by components rich in Fe, such as photosynthesis, mitochondrial electron transport and nitrate assimilation are down-regulated to cope with the reduced cellular iron quota. This retrenchment is compensated by nitrogen (N) and carbon (C) reallocation from protein and storage carbohydrate degradation, adaptations to chlorophyll biosynthesis and pigment metabolism, removal of excess electron s by mitochondrial alternative oxidase (AOX), augmented Fe-independent oxidative stress responses, and sensitized iron capture mechanisms. Keywords: Marine phytoplankton, pinnate diatom Wild-type Phaeodactylum tricornutum was grown under Fe replete (10,000 nM) and Fe limiting (5nM) conditions. Partial genome gene expression analysis of iron-inducible genes was conducted using a two-color competitive hybridization microarray.

ORGANISM(S): Phaeodactylum tricornutum

SUBMITTER: Chris Bowler

PROVIDER: E-GEOD-8675 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Whole-cell response of the pennate diatom Phaeodactylum tricornutum to iron starvation.

Allen Andrew E AE Laroche Julie J Maheswari Uma U Lommer Markus M Schauer Nicolas N Lopez Pascal J PJ Finazzi Giovanni G Fernie Alisdair R AR Bowler Chris C

Proceedings of the National Academy of Sciences of the United States of America 20080724 30

Marine primary productivity is iron (Fe)-limited in vast regions of the contemporary oceans, most notably the high nutrient low chlorophyll (HNLC) regions. Diatoms often form large blooms upon the relief of Fe limitation in HNLC regions despite their prebloom low cell density. Although Fe plays an important role in controlling diatom distribution, the mechanisms of Fe uptake and adaptation to low iron availability are largely unknown. Through a combination of nontargeted transcriptomic and metab ...[more]

PMID: 18653757

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Project description:Genes that are constitutively expressed across multiple environmental stimuli are crucial to quantifying differentially expressed genes, particularly when employing quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) assays. However, the identification of these potential reference genes in non-model organisms is challenging and is often guided by expression patterns in distantly related organisms. Here, transcriptome datasets from the diatom Thalassiosira pseudonana grown under replete, phosphorus-limited, iron-limited, and phosphorus and iron co-limited nutrient regimes were analyzed through literature-based searches for homologous reference genes, k-means clustering, and Analysis of Sequence Counts (ASC) to identify putative reference genes. A total of 9759 genes were identified and screened for stable expression. Literature-based searches surveyed 18 generally accepted reference genes, revealing 101 homologs in T. pseudonana with variable expression and a wide range of mean tags per million. K-means analysis parsed the whole transcriptome into 15 clusters. The two most stable clusters contained 709 genes but still had distinct patterns in expression. ASC analyses identified 179 genes that were stably expressed (posterior probability < 0.1 for 1.25 fold change). Genes known to have a stable expression pattern across the test treatments, like actin, were identified in this pool of 179 candidate genes. ASC can be employed on data without biological replicates and was more robust than the k-means approach in isolating genes with stable expression. The intersection of the genes identified through ASC with commonly used reference genes from the literature suggests that actin and ubiquitin ligase may be useful reference genes for T. pseudonana and potentially other diatoms. With the wealth of transcriptome sequence data becoming available, ASC can be easily applied to transcriptome datasets from other phytoplankton to identify reference genes. Axenic T. pseudonana CCMP 1335 was grown at 14Â°C under 24 hour light (120 Âµmol photons m-2 s-1) after Dyhrman et al. (2012) in f/2 plus silica chelated media made from surface Sargasso Sea water. Nitrate, silica, vitamins, and trace metals were at f/2 concentrations (Guillard and Ryther 1962), while iron and phosphate were modified across treatments. In brief, triplicate cultures of replete (36 ÂµM PO4, 400 nM Fe), P-limited (0.4 ÂµM PO4, 400 nM Fe), Fe-limited (36 ÂµM PO4, 40 nM Fe), and Co-limited (0.4 ÂµM PO4, 40 nM Fe) treatments were harvested when growth deviated from the replete control. Growth was monitored by cell counts. Biomass was harvested onto 0.2 Âµm filters and flash frozen in liquid nitrogen and total RNA was extracted as described in Dyhrman et al. (2012). Tag-seq sequencing of the transcriptome was performed by Illumina with a polyA selection and NlaIII digestion, resulting in 21 bp sequence reads or tags (Dyhrman et al., 2012). Libraries were of varied sizes as follows: replete (~12 million), P-limited (~13 million), Fe-limited (~23 million), and Co-limited (~26 million). Tags were mapped to gene models (predicted protein coding regions) with a pipeline designed by Genesifter Inc., requiring 100% identity and covering 9759 genes. Tag counts within a gene were pooled and normalized to the size of the library, with resulting data expressed in tags per million (tpm). Genes with normalized tag counts less than 2.5 tpm for each of the four treatments were excluded (Figure S1) , leaving 7380 genes in the analysis.

Dataset Information

Transcriptional response of the marine pennate diatom Phaeodactylum tricornutum to Fe limitation

Publications

Whole-cell response of the pennate diatom Phaeodactylum tricornutum to iron starvation.

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