Manganese co-localizes with calcium and phosphorus in Chlamydomonas acidocalcisomes and is mobilized in manganese-deficient conditions

Abstract

Exposing cells to excess metal concentrations well beyond the cellular quota is a powerful tool for understanding the molecular mechanisms of metal homeostasis. Such improved understanding may enable bioengineering of organisms with improved nutrition and bioremediation capacity. We report here that Chlamydomonas reinhardtii can accumulate manganese (Mn) in proportion to extracellular supply, up to 30-fold greater than its typical quota and with remarkable tolerance. As visualized by X-ray fluorescence microscopy and nanoscale secondary ion MS (nanoSIMS), Mn largely co-localizes with phosphorus (P) and calcium (Ca), consistent with the Mn-accumulating site being an acidic vacuole, known as the acidocalcisome. Vacuolar Mn stores are accessible reserves that can be mobilized in Mn-deficient conditions to support algal growth. We noted that Mn accumulation depends on cellular polyphosphate (polyP) content, indicated by 1) a consistent failure of C. reinhardtii vtc1 mutant strains, which are deficient in polyphosphate synthesis, to accumulate Mn and 2) a drastic reduction of the Mn storage capacity in P-deficient cells. Rather surprisingly, X-ray absorption spectroscopy, EPR, and electron nuclear double resonance revealed that only little Mn2+ is stably complexed with polyP, indicating that polyP is not the final Mn ligand. We propose that polyPs are a critical component of Mn accumulation in Chlamydomonas by driving Mn relocation from the cytosol to acidocalcisomes. Within these structures, polyP may, in turn, escort vacuolar Mn to a number of storage ligands, including phosphate and phytate, and other, yet unidentified, compounds.

Polyphosphate: A Multifunctional Metabolite in Cyanobacteria and Algae

“…However, very little is known about the physiological relevance of these observed variations. The number and size of polyP bodies in cyanobacterial and algal cells can vary greatly, but they generally increase under stress conditions (Stevens et al, 1985;Jensen, 1993;Docampo et al, 2010;Aksoy et al, 2014;Tsednee et al, 2019). As in other single-cell eukaryotes, the acidocalcisome membranes in C. reinhardtii harbor the VTC complex, which is responsible for the polyP polymerase and H + -PPase activities (acidocalcisome membranes also harbor a V-type ATPase driven proton pump); the H + -PPases, with homologues in three eukaryotic clades (not present in opisthokonts and Amoebozoa), are activated by pyrophosphate (Rea et al, 1992;Kim et al, 1994;Rodrigues et al, 1999;McIntosh and Vaidya, 2002;Drozdowicz et al, 2003;Au et al, 2006;Serrano et al, 2007;Hsu et al, 2009;Tsai et al, 2014;Schilling et al, 2017;Shah et al, 2017;Segami et al, 2018a;Segami et al, 2018b) and also function in acidification of the lumen of vacuoles in land plants (Venter et al, 2006;Kajander et al, 2013;Asaoka et al, 2014).…”

Metabolic control of acclimation to nutrient deprivation dependent on polyphosphate synthesis

“…Intracellular polyP aggregates were found more than a hundred years ago, and over the past three decades, many studies have shown that different organisms require polyP to cope with various stress conditions including high temperature, oxidative stress, osmotic shock, survival during stationary phase, heavy metal accumulation, and nutrient deprivation, among others (1,5,(13)(14)(15)(16)(17)(18). Moreover, polyP synthesis affects critical cellular processes, including DNA replica-tion, transcription, translation, quorum sensing, and protein folding, and is essential for normal infectivity of many parasites (1,5,(18)(19)(20)(21)(22)(23).…”

“…For example, the Lon protease interacts with polyP, activating degradation of ribosomal polypeptides during amino acid starvation of Escherichia coli (21). Last, polyP can serve as a chelator of divalent cations (Ca 2+ , Fe 2+ , Cd 2+ , Mn 2+ , and Cu 2+ , among others) and control intracellular cation availability (15,27). Despite this growing number of discoveries concerning the impact of polyP on cellular processes, many with seemingly unrelated functions, we still know little about how this polymer and its dynamic synthesis and degradation help cells acclimate to stress.…”

Sabeeha Merchant

“…The existence of a reference genome fueled a large number of deep transcriptome-sequencing studies using many different growth conditions, allowing the identification of numerous candidate genes based on expression profiles. More recently, work in the laboratory has turned to a precise accounting of all metal atoms within the cell, as well as their subcellular localization, helped by innovations in microscopy techniques (Tsednee et al, 2019;Hong-Hermesdorf et al, 2014).…”