Phosphorus and nitrogen starvation reveal life-cycle specific responses in the metabolome of Emiliania huxleyi (Haptophyta)

The coccolithophore Emiliania huxleyi is a microalga with biogeochemical and biotechnological relevance, due to its high abundance in the ocean and its ability to form intricate calcium carbonate structures. Depletion of macronutrients in oceanic waters is very common and will likely enhance with advancing climate change. We present the first comprehensive metabolome study analyzing the effect of phosphorus (P) and nitrogen (N) starvation on the diploid and haploid life-cycle stage, applying various metabolome analysis methods to gain new insights in intracellular mechanisms to cope with nutrient starvation. P-starvation led to an accumulation of many generic and especially N-rich metabolites, including lipids, osmolytes, and pigments. This suggests that P-starvation primarily arrests cell-cycling due to lacking P for nucleic acid synthesis, but that enzymatic functionality is widely preserved. Also, the de-epoxidation ratio of the xanthophyll cycle was upregulated in the diploid stage under P-starvation, indicating increased nonphotochemical quenching, a response typically observed under high light stress. In contrast, N-starvation resulted in a decrease of most central metabolites, also P-containing ones, especially in the diploid stage, indicating that most enzymatic functionality ceased. The two investigated nutrient starvation conditions caused significantly different responses, contrary to previous assumptions derived from transcriptomic studies. Data highlight that instantaneous biochemical flux is a more dominant driver of the metabolome than the transcriptomically rearranged pathway patterns. Due to the fundamental nature of the observed responses it may be speculated that microalgae with similar nutrient requirements can cope better with P-starvation than with N-starvation

An international effort towards developing standards for best practices in analysis, interpretation and reporting of clinical genome sequencing results in the CLARITY Challenge

There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data were donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease-causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance. RESULTS: A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization. CONCLUSIONS: The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups

Single-Shot Sub-microsecond Mid-infrared Spectroscopy on Protein Reactions with Quantum Cascade Laser Frequency Combs

The kinetic analysis of irreversible protein reactions requires an analytical technique that provides access to time-dependent infrared spectra in a single shot. Here, we present a spectrometer based on dual-frequency-comb spectroscopy using mid-infrared frequency combs generated by quantum cascade lasers. Attenuation of the intensity of the combs by molecular vibrational resonances results in absorption spectra covering 55 cm–1 in the fingerprint region. The setup has a native resolution of 0.3 cm–1, noise levels in the μOD range, and achieves sub-microsecond time resolution. We demonstrate the simultaneous recording of both spectra and transients of the photoactivated proton pump bacteriorhodopsin. More importantly, a single shot, i.e., a single visible light excitation, is sufficient to extract spectral and kinetic characteristics of several intermediates in the bacteriorhodopsin photocycle. This development paves the way for the noninvasive analysis of enzymatic conversions with high time resolution, broad spectral coverage, and minimal sample consumption.ISSN:1520-6882ISSN:0003-270

Phosphorus and nitrogen starvation reveal life-cycle specific responses in the metabolome of Emiliania huxleyi (Haptophyta), Supplementary Table 3

The coccolithophore Emiliania huxleyi is a microalga with biogeochemical and biotechnological relevance, due to its high abundance in the ocean and its ability to form intricate calcium carbonate structures. Depletion of macronutrients in oceanic waters is very common and will likely enhance with advancing climate change. We present the first comprehensive metabolome study analyzing the effect of phosphorus (P) and nitrogen (N) starvation on the diploid and haploid life-cycle stage, applying various metabolome analysis methods to gain new insights in intracellular mechanisms to cope with nutrient starvation. P-starvation led to an accumulation of many generic and especially N-rich metabolites, including lipids, osmolytes and pigments. This suggests that P-starvation primarily arrests cell-cycling due to lacking P for nucleic acid synthesis, but that enzymatic functionality is widely preserved. Also, the de-epoxidation ratio of the xanthophyll cycle was upregulated in the diploid stage under P-starvation, indicating increased nonphotochemical quenching, a response typically observed under high light stress. In contrast, N-starvation resulted in a decrease of most central metabolites, also P-containing ones, especially in the diploid stage, indicating that most enzymatic functionality ceased. The two investigated nutrient starvation conditions caused significantly different responses, contrary to previous assumptions derived from transcriptomic studies. Data highlight that instantaneous biochemical flux is a more dominant driver of the metabolome than the transcriptomically rearranged pathway patterns. Due to the fundamental nature of the observed responses it may be speculated that microalgae with similar nutrient requirements can cope better with P-starvation than with N-starvation