3-Hydroxyisobutyryl-CoA hydrolase involved in isoleucine catabolism regulates triacylglycerol accumulation in Phaeodactylum tricornutum

Since methylmalonyl-CoA epimerase appears to be absent in the majority of photosynthetic organisms, including diatoms, (S)-methylmalonyl-CoA, the intermediate of isoleucine (Ile) catabolism, cannot be metabolized to (R)methylmalonyl-CoA then to succinyl-CoA. In this study, propionyl-CoA carboxylase (PCC) RNAi silenced strains and 3-hydroxyisobutyryl-CoA hydrolase (HIBCH) overexpression strains were constructed to elucidate the Ile degradation pathway and its influence on lipid accumulation in Phaeodactylum tricornutum based on growth, neutral lipid content and metabolite profile analysis. Knockdown of PCC disturbed the metabolism of Ile through propionyl-CoA to methylmalonyl-CoA, as illustrated by much higher Ile content at day 6. However, Ile decreased to comparable levels to the wild-type at day 10. PCC silencing redirected propionyl-CoA to acetyl-CoA via a modified beta-oxidation pathway, and transcript levels for some branched-chain amino acid (BCAA) degradation-related genes, especially HIBCH, significantly upregulated in the PCC mutant, which enhanced the BCAA degradations and thus resulted in higher triacylglycerol (TAG) content. Overexpression of HIBCH accelerates Ile degradation and results in a lowered Ile content in the overexpression strains, thus enhancing carbon skeletons to the tricarboxylic acid cycle and giving rise to increasing TAG accumulation. Our study provides a good strategy to obtain high-lipid-yield transgenic diatoms by modifying the propionyl-CoA metabolism. This article is part of the themed issue &#39;The peculiar carbon metabolism in diatoms&#39;.</p

Concomitant percutaneous coronary intervention and transcatheter aortic valve replacement for aortic stenosis complicated with acute STEMI: a case report and literature review

Aortic stenosis (AS) complicated with acute ST-segment elevation myocardial infarction (STEMI) is a life-threatening emergency with high mortality. A 75-year-old male patient attended the emergency department of Wuhan Asia Heart Hospital in December 2021 with chest pain for 2 days and exacerbation for 1 h. The electrocardiogram (ECG) indicated atrial fibrillation with rapid ventricular response and ST-segment depression. Echocardiography showed severe AS and mild/moderate aortic insufficiency. The patient refused coronary angiography and further invasive procedures and then requested discharge, but he had recurrent chest pain on the third day. The ECG showed an extensive anterior wall STEMI. During preoperative preparation, he suffered from cardiogenic shock (CS). Concomitant percutaneous coronary intervention (PCI) and transcatheter aortic valve replacement (TAVR) was performed, but he died of CS and multiple organ failure 4 days after surgery. Patients with AS and STEMI might be susceptible to CS during perioperative period of concomitant PCI and TAVR, which requires proactive prevention

A high sensitivity iron-dependent bioreporter used to measure iron bioavailability in freshwaters

A Nostoc sp. PCC 7120 iron bioreporter containing iron-regulated schizokinen transporter gene alr0397 promoter fused to the luxAB genes was examined to optimize its response to bioavailable iron. Doseresponse relationships between luciferase activity and free ferric ion (Fe3+) concentrations pFe (-lg [Fe3+]) were generated by measuring luciferase activities of the bioreporter in trace metalbuffered Fraquil medium with various incubation times. The results were best demonstrated by sigmoidal curves (pFe 18.821.7, Fe3+ = 10-18.810-21.7 M) with the linear range extending from pFe 19.621.5 (Fe3+ = 10-19.610-21.5 M) after a 12-h incubation time. Optimal conditions for the use of this bioreporter to sense the iron bioavailability were determined to be: a 12-h exposure time, initial cell density of OD730 nm = 0.06, high nitrate (100 mu M), high phosphate (10 mu M), moderate Co2+ (0.122.5 nM), Zn2+ (0.1612 nM), Cu2+ (0.0450 nM), and wide range of Mn2+ concentration (0.922300 nM). The applicability of using this iron bioreporter to assess iron availability in the natural environment has been tested using water samples from eutrophic Taihu, Donghu, and Chaohu lakes. It is indicated that the bioreporter is a useful tool to assess bioavailable iron in various water quality samples, especially in eutrophic lakes with high bioavailable iron.A Nostoc sp. PCC 7120 iron bioreporter containing iron-regulated schizokinen transporter gene alr0397 promoter fused to the luxAB genes was examined to optimize its response to bioavailable iron. Doseresponse relationships between luciferase activity and free ferric ion (Fe3+) concentrations pFe (-lg [Fe3+]) were generated by measuring luciferase activities of the bioreporter in trace metalbuffered Fraquil medium with various incubation times. The results were best demonstrated by sigmoidal curves (pFe 18.821.7, Fe3+ = 10-18.810-21.7 M) with the linear range extending from pFe 19.621.5 (Fe3+ = 10-19.610-21.5 M) after a 12-h incubation time. Optimal conditions for the use of this bioreporter to sense the iron bioavailability were determined to be: a 12-h exposure time, initial cell density of OD730 nm = 0.06, high nitrate (100 mu M), high phosphate (10 mu M), moderate Co2+ (0.122.5 nM), Zn2+ (0.1612 nM), Cu2+ (0.0450 nM), and wide range of Mn2+ concentration (0.922300 nM). The applicability of using this iron bioreporter to assess iron availability in the natural environment has been tested using water samples from eutrophic Taihu, Donghu, and Chaohu lakes. It is indicated that the bioreporter is a useful tool to assess bioavailable iron in various water quality samples, especially in eutrophic lakes with high bioavailable iron

Potential impact of stress activated retrotransposons on genome evolution in a marine diatom

<p>Abstract</p> <p>Background</p> <p>Transposable elements (TEs) are mobile DNA sequences present in the genomes of most organisms. They have been extensively studied in animals, fungi, and plants, and have been shown to have important functions in genome dynamics and species evolution. Recent genomic data can now enlarge the identification and study of TEs to other branches of the eukaryotic tree of life. Diatoms, which belong to the heterokont group, are unicellular eukaryotic algae responsible for around 40% of marine primary productivity. The genomes of a centric diatom, <it>Thalassiosira pseudonana</it>, and a pennate diatom, <it>Phaeodactylum tricornutum</it>, that likely diverged around 90 Mya, have recently become available.</p> <p>Results</p> <p>In the present work, we establish that LTR retrotransposons (LTR-RTs) are the most abundant TEs inhabiting these genomes, with a much higher presence in the <it>P. tricornutum </it>genome. We show that the LTR-RTs found in diatoms form two new phylogenetic lineages that appear to be diatom specific and are also found in environmental samples taken from different oceans. Comparative expression analysis in <it>P. tricornutum </it>cells cultured under 16 different conditions demonstrate high levels of transcriptional activity of LTR retrotransposons in response to nitrate limitation and upon exposure to diatom-derived reactive aldehydes, which are known to induce stress responses and cell death. Regulatory aspects of <it>P. tricornutum </it>retrotransposon transcription also include the occurrence of nitrate limitation sensitive <it>cis</it>-regulatory components within LTR elements and cytosine methylation dynamics. Differential insertion patterns in different <it>P. tricornutum </it>accessions isolated from around the world infer the role of LTR-RTs in generating intraspecific genetic variability.</p> <p>Conclusion</p> <p>Based on these findings we propose that LTR-RTs may have been important for promoting genome rearrangements in diatoms.</p

High efficiency transformation by electroporation of the freshwater alga Nannochloropsis limnetica

The microalgal genus of Nannochloropsis is considered one of the most promising organisms for the production of biofuels due to their high lipid content. Transformation systems for marine Nannochloropsis species have been established in the recent decade, however, genetic manipulation of Nannochloropsis limnetica, the only known freshwater species in this genus, is not yet available. Based on established marine Nannochloropsis species electrotransformation protocol, nuclear genetic transformation was established in N. limnetica, meanwhile the appropriate antibiotic selection concentration and electric field strength of electroporation were determined. For the selection of transformants in N. limnetica on plates, 0.07 mu gmL(-1) of zeocin or 5 mu gmL(-1) of hygromycin B was proved sufficient, and the transformation efficiency was &lt;2x10(-8) with a single pulse ranging from 2200 to 2600V using 2-mm electroporation cuvettes. Pretreatment of N. limnetica with 10mM lithium acetate and 3mM dithiothreitol before electroporation increased transformation efficiency hundreds of times, and the highest transformation efficiency of 10-11x10(-6) was obtained with an electric field strength of 12,000V/cm. Our results help to expand the biotechnological applications of this freshwater species and provide means for successful electrotransformation of other microalgae as well.Graphic abstractHigh-efficiency transformation of freshwater Nannochloropsis pretreatment of N. limnetica with 10 mM lithium acetate and 3 mM dithiothreitol before electroporation increased transformation efficiency hundredsoftimes. [GRAPHICS] .</p

High-efficiency nuclear transformation of the diatom Phaeodactylum tricornutum by electroporation

We established a high-efficiency nuclear transformation method for the diatom Phaeodactylum tricornutum using an electroporation system. Based on a universal electroporation protocol, the conditions for the introduction of exogenous DNA including electric field strength and plasmid form were optimized. Following optimization, the diatom cells could be transformed with exogenous gene easily, the maximum transformation frequency obtained was 2.8 x 10(-5) cells. The cotransformation of P. tricornutum with a non-selective GUS gene together with the selectable resistance gene has also been achieved using our new method and found to be very efficient (up to 60%). The electroporation procedure described in this article offers a number of advantages, including simplicity, general utility, low-cost and high efficiency. The described method also provides some clue for developing electroporation transformation system in other eukaryotic microalgae. (C) 2013 Elsevier B.V. All rights reserved

Screening High CO2-Tolerant Oleaginous Microalgae from Genera Desmodesmus and Scenedesmus

Microalgae from genus Scenedesmus sensu lato (including Desmodesmus and Scenedesmus) were reported to be particularly suitable candidates for CO2 biomitigation. In this study, 16 strains from Scenedesmus sensu lato were obtained from different climate zones of China and their phylogenetic positions were determined. Seven strains out of the 16 showed high CO2 tolerance and grew much faster under 20% CO2 than air condition. Two representatives from genera Desmodesmus (NMD46) and Scenedesmus (HBX310) respectively were selected due to their higher lipid productivity, and the maximum value of 146 mg L-1 day(-1) was achieved in NMD46. Triacylglycerols increased with the rising of CO2 levels from 0.04 to 15% in NMD46, while they changed little in HBX310. High CO2 level decreased the polyunsaturated fatty acid content in NMD46 but increased it in HBX310. NMD46 is more suitable for standardized biodiesel production in view of its lipid and fatty acid composition responses to high CO2

J. Appl. Phycol.

The growth and toxin content of the dinoflagellate Alexandrium tamarense ATHK was markedly affected by culture methods. In early growth phase at lower cell density static or mild agitation methods were beneficial to growth, but continuous agitation or aeration, to some extent, had an adverse effect on cell growth. Static culture in 2 L Erlenmeyer flasks had the highest growth rate (0.38 d(-1)) but smaller cell size compared with other culture conditions. Cells grown under aerated conditions possessed low nitrogen and phosphorus cell yields, namely high N and P cell-quota. At day 18, cells grown in continuous agitated and 1 h aerated culture entered the late stationary phase and their cellular toxin contents were higher (0.67 and 0.54 pg cell(-1)) compared with cells grown by other culture methods (0.27-0.49 pg cell(-1)). The highest cell density and cellular toxin content were 17190 cells mL(-1) and 1.26 pg cell(-1) respectively in an airlift photobioreactor with two-step culture. The results indicate that A. tamarense could be grown successfully in airlift photobioreactor by a two-step culture method, which involved cultivating the cells statically for 4 days and then aerating the medium. This provides an efficient way to enhance cell and toxin yield of A. tamarense.The growth and toxin content of the dinoflagellate Alexandrium tamarense ATHK was markedly affected by culture methods. In early growth phase at lower cell density static or mild agitation methods were beneficial to growth, but continuous agitation or aeration, to some extent, had an adverse effect on cell growth. Static culture in 2 L Erlenmeyer flasks had the highest growth rate (0.38 d(-1)) but smaller cell size compared with other culture conditions. Cells grown under aerated conditions possessed low nitrogen and phosphorus cell yields, namely high N and P cell-quota. At day 18, cells grown in continuous agitated and 1 h aerated culture entered the late stationary phase and their cellular toxin contents were higher (0.67 and 0.54 pg cell(-1)) compared with cells grown by other culture methods (0.27-0.49 pg cell(-1)). The highest cell density and cellular toxin content were 17190 cells mL(-1) and 1.26 pg cell(-1) respectively in an airlift photobioreactor with two-step culture. The results indicate that A. tamarense could be grown successfully in airlift photobioreactor by a two-step culture method, which involved cultivating the cells statically for 4 days and then aerating the medium. This provides an efficient way to enhance cell and toxin yield of A. tamarense