Journal: Nature Plants

Article Title: Environmental gradients reveal stress hubs pre-dating plant terrestrialization

doi: 10.1038/s41477-023-01491-0

Figure Lengend Snippet: a , DIC and confocal micrographs of Mesotaenium endlicherianum SAG 12.97 cells accumulating LDs (arrows) upon exposure to different temperature/light conditions (abbreviations) of the gradient table for 89 h or 216 h. For confocal microscopy, algae were cultured independent of table conditions at 75 µmol photons m −2 s −1 and 22 °C for 22 days. LDs are visible as distinct globular structures and were stained with BODIPY (false-coloured green; 493 nm excitation, 503 nm emission); chlorophyll autofluorescence in false-coloured purple; for each condition, at least ten micrographs were taken, all showing similar phenotypes of the cells. b , Violin plots of LD quantification after 9 days of exposure to different environmental conditions; significance grouping (Mann–Whitney U ) is based on P < 0.05; see also Supplementary Fig. . c , Heat map of row-scaled z scores of the expression of homologues for LD biogenesis and function (see also Supplementary Fig. ). Conditions are displayed at the bottom as symbols in different colours; best Arabidopsis hits (via BLASTp) are shown on the right. d , e , Proteomic investigation into lipid-enriched phases extracted from Mesotaenium ; note the enrichment in hallmark proteins of LDs. Volcano plot showing significantly (false discovery rate (FDR) <0.05) enriched Mesotaenium proteins in the lipid-enriched (LD) versus the TE ( d ). Bar plots show the relative, normalized iBAQ values for ten LD signature proteins detected in Mesotaenium ( e ). Bottom bar plot shows the log 2 enrichment of proteins characteristic for subcellular compartments. LL, ML and HL, low, moderate and high light; LT, MT and HT, low, moderate and high temperature, respectively; ER, endoplasmic reticulum. f , LD proteins of M. endlicherianum localize to LDs in tobacco pollen tubes: cLSM images of transiently expressed proteins appended to mCherry in transiently transformed N. tabacum pollen tubes. LDs were stained with BODIPY 493/503; for each construct, the images are representative of at least nine micrographs of transformed pollen tubes per fusion construct. Scale bars, 10 µm. g , h , Lipid composition in M. endlicherianum LDs of 12- to 25-week-old cultures and standards for sterol esters (SE), FFA, free sterols (free S), TAG and DAG via analytical TLC ( g ) and preparative TLC followed by GC for profiling ( h ). i , Full lipid profiles assessed via GC.

Article Snippet: Confocal laser scanning microscope was done on a Zeiss LSM780 (Carl Zeiss) set as in Müller et al. . For the staining of the LD structures, we used the neutral lipid specific stain BODIPY 493/503 (EM/EX) (Merck).

Techniques: Confocal Microscopy, Algae, Cell Culture, Staining, MANN-WHITNEY, Expressing, Transformation Assay, Construct