Nitrogen-Sparing Mechanisms in Chlamydomonas: Reduce, Reuse, Recycle, and Reallocate

It is a universal axiom that when times are hard, the practices of reducing, reusing, and recycling waste and resources—arguably good habits even at the best of times—can make a critical difference to one's well-being and ability to thrive, or even survive. It turns out that this is as true for the single-celled green alga Chlamydomonas reinhardtii as for any of us. Nitrogen is an essential macronutrient, and a costly one for plants, many of which live in N-limited conditions. N availability is one of the most limiting factors to crop yields and one of the most expensive to supply. Excessive runoff of N from agricultural lands is not only costly, but also contributes to N pollution, leading to reduced water quality and dead zones in many coastal waters worldwide. There is also increased interest in the use of algae in biotechnology applications, as an important source for biofuels, bioplastics, fertilizer, pharmaceuticals, and a variety of other products. Therefore, it is of considerable importance to gain a deep understanding of N metabolism and N use efficiency, in algae as well as land plants.

An immediate response of algae and other microorganisms to nutrient limitation is to put more effort into acquisition by upregulating specific transporters and other components of nutrient uptake and to begin mobilizing stored resources. The next step is to employ “austerity measures” involving nutrient sparing and recycling (reviewed in Merchant and Helmann, 2012). New work by Schmollinger et al. (pages 1410–1435) investigates these mechanisms operating in N-limited C. reinhardtii cells via deep sequencing of mRNA and parallel quantitative proteomics.

C. reinhardtii is a facultative autotroph that can grow on organic carbon sources, and the authors used acetate-grown cells to minimize potentially confounding results due to changes in photosynthetic rate. As expected, the transcriptome responded immediately to reduced N by upregulating transporters for N acquisition, followed by increased release of N from abundant molecules (ribosomes, Rubisco, and chlorophyll) from upregulation of associated degradation pathways. There was also a decrease in many proteins involved in the Calvin-Benson cycle, photosynthetic electron transport, and photophosphorylation. By contrast, N-depleted cells showed an increase in proteins involved in mitochondrial respiration, suggesting that N-starved cells grown on acetate maintain a bioenergetic preference for respiration.

N deficiency also leads to the accumulation of lipid bodies in C. reinhardtii (Boyle et al., 2012), which is further enhanced in acetate-grown cells (Goodson et al., 2011). Proteins associated with early steps of fatty acid metabolism were found to be significantly reduced in N-starved cells, emphasizing the role of recycling of thylakoid membranes in TAG synthesis, rather than de novo fatty acid synthesis, under these conditions.

An increase in N-use efficiency was seen in that the abundant proteins that decreased under N deficiency, such as Rubisco and ribosomal subunits, are rich in amino acids with N-containing side chains, whereas proteins that increased in N-starved medium have less N in side chains. Overall, the proteins induced upon N starvation had ∼6% less nitrogen compared with proteins whose abundance decreased, suggesting a proteome-wide N-sparing mechanism of changes in acid composition (see figure). It may not be merely coincidental that transporters and other proteins involved in N acquisition (upregulated under N deprivation) also have a lower than average N content. C. reinhardtii adds reallocation of scarce resources to the three Rs of environmental planning!

A proteome-wide N-sparing mechanism under N deprivation. Molar carbon-to-nitrogen ratio in C. reinhardtii cells after transfer to N-containing (+N) or N-free (−N) media (top panel). Distributions of nitrogen atoms in amino acid side chains of upregulated (green) or downregulated (red) proteins, relative to the whole proteome (gray) (bottom panel). (Reprinted from Schmollinger et al. [2014], Figures 6C and 6D.)