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. 2022 Apr 14;13(1):1995.
doi: 10.1038/s41467-022-29603-y.

Adaptive responses of marine diatoms to zinc scarcity and ecological implications

Affiliations

Adaptive responses of marine diatoms to zinc scarcity and ecological implications

Riss M Kellogg et al. Nat Commun. .

Abstract

Scarce dissolved surface ocean concentrations of the essential algal micronutrient zinc suggest that Zn may influence the growth of phytoplankton such as diatoms, which are major contributors to marine primary productivity. However, the specific mechanisms by which diatoms acclimate to Zn deficiency are poorly understood. Using global proteomic analysis, we identified two proteins (ZCRP-A/B, Zn/Co Responsive Protein A/B) among four diatom species that became abundant under Zn/Co limitation. Characterization using reverse genetic techniques and homology data suggests putative Zn/Co chaperone and membrane-bound transport complex component roles for ZCRP-A (a COG0523 domain protein) and ZCRP-B, respectively. Metaproteomic detection of ZCRPs along a Pacific Ocean transect revealed increased abundances at the surface (<200 m) where dZn and dCo were scarcest, implying Zn nutritional stress in marine algae is more prevalent than previously recognized. These results demonstrate multiple adaptive responses to Zn scarcity in marine diatoms that are deployed in low Zn regions of the Pacific Ocean.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1. Growth responses of diatoms to varying [Zn2+] and [Co2+] and initial detection of ZCRPs in T. pseudonana.
Growth rates of four diatoms over a range of a [Zn2+] and b [Co2+]. Data are presented as mean values of biological duplicate cultures. Data is available in Supplementary Table 1. Global proteomic analyses comparing the proteomes of pooled biological duplicate cultures (n = 2) of T. pseudonana in c high vs. low added Zn and d high vs. low added Co. Each point is an identified protein with the mean of technical triplicate abundance scores in one treatment plotted against the mean of abundance scores in another treatment. The solid line denotes 1:1 abundance. Error bars in c are the standard deviation of technical triplicate measurements.
Fig. 2
Fig. 2. Expression trends of ZCRPs in four marine diatom species.
Mean spectral counting abundance scores of ZCRP-A and ZCRP-B in a 6 treatments of T. pseudonana, b 12 treatments of wild-type P. tricornutum, c 6 treatments of P. delicatissima, and d 3 Zn treatments of Chaetoceros RS19 as measured by global proteomic analysis. For all diatoms, spectral counts were derived from pooled biological duplicate cultures (n = 2). Where available, data are presented as mean values ± the standard deviation of technical triplicate measurements with individual data points overlaid (white circles). Data presented without error bars were measured in technical singlicate. ND, not detected. No proteomic data is available for Chaetoceros RS19 grown in Co treatments as ample biomass for proteomic analysis was not available. e Spectral counts of ZCRP-A and f ZCRP-B compared to growth rates of P. tricornutum in Co treatments (μCo) and Zn treatments (μZn). Data available from Supplementary Table 1.
Fig. 3
Fig. 3. Algal ZCRP-A sequence alignments and predicted structures.
a Sequence alignment of the E. coli YjiA protein compared to ZCRP-A proteins in C. reinhardtii, P. tricornutum, T. pseudonana, P. delicatissima, and Chaetoceros RS19 generated using MUSCLE. Four conserved GTPase domains including the putative CXCC metal binding motif are labeled. Asterisks denote the glutamate (E) and cysteine (C) residues predicted to bind Zn2+. Notably, one of the metal binding E resides in the G2/Switch I region has been replaced by valine (V; red box) in Chaetoceros RS19. b Predicted structures of ZCRP-A homologs modeled using the well-characterized Zn2+ and Co2+ binding COG0523 GTPase YjiA protein in E. coli as a template (PDB entry 4IXM.2.b). The predicted ligand Zn2+ is indicated by a red arrow. Close-up views show the predicted Zn2+ binding site composed of two glutamate (E) and one cysteine (C) residue.
Fig. 4
Fig. 4. Confocal and epifluorescent micrographs of the ZCRP-A and ZCRP-B proteins fused to YFP and overexpressed in P. tricornutum.
a Localization of ZCRP-A to the chloroplasts and b localization of ZCRP-B to the cell membrane. YFP fluorescence is false-colored green and chlorophyll autofluorescence is false-colored red. Composite images are stacks of the individual channels c, d differential interference contrast (DIC), e, f yellow fluorescent protein (YFP), and g, h chlorophyll autofluorescence (Chl auto). i Micrographs of Zn-limited wild-type and ZCRP-A knockout (KO) P. tricornutum cells showing morphological differences. For ah and i, results were validated > 10 times. j Topology predictions from five sub-methods (OCTOPUS, Philius, PolyPhobius, SCAMPI, and SPOCTOPUS), consensus prediction (TOPCONS), and predicted ΔG values for P. tricornutum ZCRP-B generated using the TOPCONS webserver (https://topcons.cbr.su.se/),. k Extent of Co uptake after 24 h for wild-type (WT), ZCRPA-knockout (KO), and ZCRPA-overexpression (OE) lines of P. tricornutum normalized to fluorescence units (fsu). Data are presented as mean values ± the standard deviation of biological triplicate cultures (n = 3). Individual data points are overlaid as white circles. The extent of Co uptake was found to be significantly larger in the ZCRPA-OE line compared to the wild-type via one-way ANOVA (f(3) = 23.16, p = 0.000268) and post hoc Dunnett test (p = 0.00048).
Fig. 5
Fig. 5. Comparison of α-CA, ι-CA, and ZCRP abundances.
Spectral counting abundance scores of a alpha CA, iota CA, and b ZCRP-A and ZCRP-B detected in Zn and Co treatments of P. tricornutum measured by global proteomic analysis. Data are plotted as means ± the standard deviation of technical triplicate measurements of pooled biological duplicate cultures (n = 2). Protein names are shown with their corresponding JGI protein ID.
Fig. 6
Fig. 6. Metaproteomic detection of ZCRP homologs in the Southern Pacific.
a Transect sampled for dissolved Zn and metaproteomics during the KM1128 METZYME research expedition on the R/V Kilo Moana October 1–25, 2011 from Oʻahu, Hawaiʻi to Apia, Samoa, in the tropical South Pacific. b The concentration of total dissolved Zn and c the concentration of total dissolved Co measured in the upper 1000 m in color scale along the METZYME transect. d Total spectral counts of the ZCRP-A homolog in the dinoflagellate Azadinium spinosum and e total spectral counts of the ZCRP-B homolog in the diatom Pseudo-nitzschia fraudulenta and measured in the upper 1000 m in color scale along the METZYME transect. f Total spectral counts of ZCRP-A homologs detected in the dinoflagellate A. spinosum, the diatom H. tamensis, and the haptophytes E. huxleyi, and Phaeocystis sp. compared to total dissolved Zn concentration. g Total spectral counts of ZCRP-B homologs detected in the dinoflagellates G. spinifera, Symbiodinium sp., A. spinosum, and in the diatoms P. fraudulenta and L. danicus compared to dZn. Detected species correspond to nearest taxonomic matches.
Fig. 7
Fig. 7. Phytoplankton pigment concentrations measured over the METZYME (KM1128) transect.
The concentration of a chlorophyll a, b fucoxanthin, and c 19′-hexanoyloxyfucoxanthin (19′-Hex) measured in the upper 500 m in color scale along the transect. This data was originally described in Saito et al. 2014 and Cohen et al. 2021 and is available on BCO-DMO (https://www.bco-dmo.org/project/2236).

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