Intraspecific variability in membrane proteome, cell growth, and morphometry of the invasive marine neurotoxic dinoflagellate Alexandrium pacificum grown in metal-contaminated conditions

Abstract

Over the past decades, the occurrence, distribution and intensity of harmful algal blooms involving the dinoflagellate Alexandrium pacificum have increased in marine coastal areas disturbed by anthropogenic inputs. This invasive species produces saxitoxin, which causes the paralytic shellfish poisoning syndrome in humans upon consumption of contaminated seafood. Blooms of A. pacificum have been reported in metal-contaminated coastal ecosystems, suggesting some ability of these microorganisms to adapt to and/or resist in metal stress conditions. This study seeks to characterize the modifications in membrane proteomes (by 2-D electrophoresis coupled to LC-MS/MS), cell growth and morphometry (measured with an inverted microscope), in response to metal stress (addition of Zn2+, Pb2+, Cu2+ and Cd2+), in two Mediterranean A. pacificum strains: SG C10-3 and TAR C5-4F, respectively isolated from the Santa Giusta Lagoon (Sardinia, Italy) and from the Tarragona seaport (Spain), both metal-contaminated ecosystems. In the SG C10-3 cultures grown in a metal cocktail, cell growth was significantly delayed, and cell size increased (22% of 37.5 μm cells after 25 days of growth). Conversely, no substantial change was observed for cell growth or cell size in the TAR C5-4F cultures grown in a metal cocktail (P > 0.10), thus indicating intraspecific variability in the responses of A. pacificum strains to metal contamination. Regardless of the conditions tested, the total number of proteins constituting the membrane proteome was significantly higher for TAR C5-4F than for SG C10-3, which may help TAR C5-4F to thrive better in contaminated conditions. For both strains, the total number of proteins constituting the membrane proteomes was significantly lower in response to metal stress (29% decrease in the SG C10-3 proteome: 82 ± 12 proteins for controls, and 58 ± 12 in metal-contaminated cultures; 17% decrease in the TAR C5-4F proteome: 101 ± 8 proteins for controls, and 84 ± 5 in metal-contaminated cultures). Moreover, regardless of the strain, proteins with significantly modified expression in response to stress were mainly Please note that this is an author-produced PDF of an article accepted for publication following peer review. The definitive publisher-authenticated version is available on the publisher Web site. down-regulated (representing 45% of the proteome for SG C10-3 and 38% for TAR C5-4F), clearly showing the harmful effects of the metals. Protein down-regulation may affect cell transport (actin and phospholipid scramblase in SG C10-3), photosynthesis (RUBISCO in SG C10-3, light-harvesting protein in TAR C5-4F, and high-CO2-inducing periplasmic protein in both strains), and finally energy metabolism (ATP synthase in both strains). However, other modifications in protein expression may confer to these A. pacificum strains a capacity for adaptation and/or resistance to metal stress conditions, for example by (i) limiting the metal entry through the plasma membrane of the SG C10-3 cells (via the down-regulation of scramblase...

Metal stresses modify soluble proteomes and toxin profiles in two Mediterranean strains of the distributed dinoflagellate Alexandrium pacificum

“…A proteomic study on the Mediterranean A. pacificum strain ACT03 (isolated from Thau Lagoon, France) exposed to monometallic stresses revealed upregulated proteins (ATP synthase) that contribute to an adaptive proteomic response of this dinoflagellate in metal-contaminated ecosystems (Jean et al, 2017). Chetouhi et al (2020) have combined ecotoxicoproteomic (membrane proteomes) and physiological (morphometry) approaches to compare the responses to polymetallic stress (zinc, lead, copper and cadmium used in cocktail) of the Mediterranean A. pacificum strains SG C10-3 and TAR C5-4F, respectively coming from the metal-contaminated Mediterranean areas of the Santa Giusta Lagoon (Luglié et al, 2002) and the Tarragona seaport (Bravo et al, 2008). In these conditions, the strains showed differences in cell growth and morphometry, in addition to different modifications of the membrane proteomes, potentially conferring to the ability for adaptation and/or tolerance under metal stresses.…”

OMICS Approaches to Assess Dinoflagellate Responses to Chemical Stressors

“…The emergence of high-throughput proteomics using mass spectrometry enables the identification of thousands of proteins and the development of protein databases for dinoflagellate proteins ( Table 1 ). This facilitates the investigation of proteome modifications under stress conditions [ 19 ]. Recent studies notably focused on dinoflagellate responses to nutrient starvations.…”

“…In response to metal stress, both soluble and membrane proteomes are modified in two Alexandrium species and suggest a decrease in photosynthesis efficiency [ 19 , 44 ] ( Figure 1 ). In A. catenella , the ATP synthase upregulation is proposed as a compensatory effect of the overall proteome downregulation and photosynthesis reduction as a strategy to improve energy production and metal removal from the cell [ 44 ] ( Figure 1 ).…”

“…Conversely, in A. pacificum , the ATP synthase subunit is downregulated and the subsequent ATP synthesis reduction may constitute a strategy to limit oxidative stress. Additionally, the downregulation of the scramblase protein may be an adaptive response to decrease metal uptake [ 19 ] ( Figure 1 ). Regarding defensive strategies under zinc and lead stress, oxidative stress proteins are downregulated (e.g., proteasome subunit) [ 44 ].…”

“…The downregulation of metal-binding transporter is a strategy to limit metal entry in the cell The ATP synthase downregulation is a strategy to reduce oxidative stress [19] Proteomic responses to phosphorus deficiency to understand HABs' formation Lipid content, docosahexaenoic acid (DHA) productivity, fatty acid composition and metabolomic analysis under different nitrogen-feeding strategies…”