Amino Acid Changes during Energy Storage Compounds Accumulation of Microalgae under the Nitrogen Depletion

The nitrogen depletion stress is widely used to promote energy storage compound (ESC) production of microalgae, such as starch and lipids. Our cultivation results and most reports show that during the nitrogen depletion, the fast ESC’s accumulation happens around the overall nitrogen content lowered to the half of normal cells. It indicates that the cells may take an active nitrogen reassembly to rebalance the requirement of nitrogen, in which the amino acid conversion should play an important role. So here, using a marine strain, Isochrysis zhanjiangensis, as the model to give a detail view on metabolic, transcriptomic and proteomic levels within the “golden period” of ESC’s accumulation. To monitor the metabolic transition in response to nitrogen starvation, the intracellular metabolite fluctuation within 32 h was profiled by GC-MS and LC-MS scanned in selected ion monitoring mode for the first time. These techniques identified and quantified the levels of 14 SMAs, 2 carbohydrates involved in the TCA cycle and glycolysis, and 28 free amino acids (AAs). The pulsed increase of pyruvate, which is the precursor of acetyl-CoA and fatty acids (FAs), indicated a potential to produce more FAs. Although overall AAs showed a decreasing trend under the experimental conditions, Ala and Phe showed increased levels initially. Meanwhile, the transcriptomic and proteomic studies were utilized, and the nitrogen metabolic pathways were studied in this ESC’s fast accumulation period. It is found that gamma-aminobutyric acid (GABA) and other non-protein AAs also play important roles in the regulation of energy metabolism

EvoMoE: An Evolutional Mixture-of-Experts Training Framework via Dense-To-Sparse Gate

Mixture-of-experts (MoE) is becoming popular due to its success in improving the model quality, especially in Transformers. By routing tokens with a sparse gate to a few experts (i.e., a small pieces of the full model), MoE can easily increase the model parameters to a very large scale while keeping the computation cost in a constant level. Most existing works just initialize some random experts, set a fixed gating strategy (e.g., Top-k), and train the model from scratch in an ad-hoc way. We identify that these MoE models are suffering from the immature experts and unstable sparse gate, which are harmful to the convergence performance. In this paper, we propose an efficient end-to-end MoE training framework called EvoMoE. EvoMoE starts from training one single expert and gradually evolves into a large and sparse MoE structure. EvoMoE mainly contains two phases: the expert-diversify phase to train the base expert for a while and spawn multiple diverse experts from it, and the gate-sparsify phase to learn an adaptive sparse gate and activate a dynamic number of experts. EvoMoE naturally decouples the joint learning of both the experts and the sparse gate and focuses on learning the basic knowledge with a single expert at the early training stage. Then it diversifies the experts and continues to train the MoE with a novel Dense-to-Sparse gate (DTS-Gate). Specifically, instead of using a permanent sparse gate, DTS-Gate begins as a dense gate that routes tokens to all experts, then gradually and adaptively becomes sparser while routes to fewer experts. Evaluations are conducted on three popular models and tasks, including RoBERTa for masked language modeling task, GPT for language modeling task and Transformer for machine translation task. The results show that EvoMoE outperforms existing baselines, including Switch, BASE Layer, Hash Layer and StableMoE

Regulatory mechanisms of oxidative species and phytohormones in marine microalgae Isochrysis zhangjiangensis under nitrogen deficiency

Oxidative species and phytohormones are important factors in plants that can regulate stress responses. In this study, the content variations of four oxidative species, O-2(-), H2O2, ClO-, and NO, and the phytohormones, salicylic acid (SA), jasmonic acid (JA), and folic acid (FA), were investigated in the microalgae Isochrysis zhangjiangensis under nitrogen deficiency. The O-2(-) and H2O2 contents were found to be positively correlated with the photosynthetic activity of the microalgae, while the opposite relationship was observed for NO and ClO-. In higher plants, there is a balance model for NO and ROS interactions, which revealed that the cooperation of NO and H2O2 regulates the pathogen-induced hypersensitive response-associated cell death. Because the NO and H2O2 levels in both nitrogen-replete and nitrogen-depleted I. zhangjiangensis were not simultaneously high, we proposed that as a unicellular lower plant, the microalgae I. zhangjiangensis follows this model and regulates the different types of intracellular oxidative species, especially when they were cultured from nitrogen sufficient condition to corresponding nitrogen deficient condition, to avoid cell death. Together with the corresponding changes in the SA, JA and FA contents in the nitrogen-stressed microalgae, we proposed that the balance model of oxidative species interactions, as well as the correlations among nitrogen assimilation, photosynthesis and the biosyntheses of SA, JA, and FA, may originally exist in algae, and plants have followed this model and maintained these correlations all along to protect themselves from stressful conditions during their evolutions from alga to higher plants. (C) 2016 Published by Elsevier B.V

Molecular characterization of CO2 sequestration and assimilation in microalgae and its biotechnological applications

Microalgae are renewable feedstock for sustainable biofuel production, cell factory for valuable chemicals and promising in alleviation of greenhouse gas CO2. However, the carbon assimilation capacity is still the bottleneck for higher productivity. Molecular characterization of CO2 sequestration and assimilation in microalgae has advanced in the past few years and are reviewed here. In some cyanobacteria, genes for 2-oxoglytarate dehydrogenase was replaced by four alternative mechanisms to fulfill TCA cycle. In green algae Coccomyxa subellipsoidea C-169, alternative carbon assimilation pathway was upregulated under high CO2 conditions. These advances thus provide new insights and new targets for accelerating CO2 sequestration rate and enhancing bioproduct synthesis in microalgae. When integrated with conventional parameter optimization, molecular approach for microalgae modification targeting at different levels is promising in generating value-added chemicals from green algae and cyanobacteria efficiently in the near future. (c) 2017 Elsevier Ltd. All rights reserved

Determination of Internal Controls for Quantitative Gene Expression of Isochrysis zhangjiangensis at Nitrogen Stress Condition

Isochrysis zhangjiangensis is a potential marine microalga for biodiesel production, which accumulates lipid under nitrogen limitation conditions, but the mechanism on molecular level is veiled. Quantitative real-time polymerase chain reaction (qPCR) provides the possibility to investigate the gene expression levels, and a valid reference for data normalization is an essential prerequisite for firing up the analysis. In this study, five housekeeping genes, actin (ACT), alpha-tubulin (TUA), beta-tubulin (TUB), ubiquitin (UBI), 18S rRNA (18S) and one target gene, diacylglycerol acyltransferase (DGAT), were used for determining the reference. By analyzing the stabilities based on calculation of the stability index and on operating the two types of software, geNorm and bestkeeper, it showed that the reference genes widely used in higher plant and microalgae, such as UBI, TUA and 18S, were not the most stable ones in nitrogen-stressed I. zhangjiangensis, and thus are not suitable for exploring the mRNA expression levels under these experimental conditions. Our results show that ACT together with TUB is the most feasible internal control for investigating gene expression under nitrogen-stressed conditions. Our findings will contribute not only to future qPCR studies of I. zhangjiangensis, but also to verification of comparative transcriptomics studies of the microalgae under similar conditions