Vacuoles of diatoms
Final Report Abstract
In the vacuolar project we have identified 11 vacuolar proteins of the diatom Phaeodactylum tricornutum in silico, which show in in vivo experiments a localization in the tonoplast of the marbled vacuole of the diatom. These vacuolar proteins include factors having homologs in the model plant Arabidopsis thaliana, but also unknown tonoplast proteins. For the latter we applied two search criteria by which we identified vacuolar proteins: a signal peptide-dependent approach or via detection of a putative vacuolar targeting motif. Targeting of proteins in the endomembrane system is specified by targeting Signals and di-leucine as well as a tyrosine-based motifs are important examples. In a screen for targeting motifs and by manipulating them, we have shown that both mentioned motifs are, depending an the proteins, involved in vacuolar protein targeting. Moreover, both targeting motifs can be functionally present in one protein in parallel and mutation can lead to different mis-localization effects. Plant-type vacuolar sorting receptors (ptVSRs) are type I membrane proteins and involved in receptor-mediated trafficking of soluble vacuolar proteins in plant cells. We have identified three different ptVSR genes in the genome of P. tricornutum, which showed, after expression as a eGFP-fusion protein, a slightly different subcellular localization. Furthermore, we tested the expression of ptVSRs, in which the membrane spanning C-terminal region was deleted. This led to the secretion of one ptVSR member in P. tricornutum. Thus, manipulated ptVSRs of the diatom can be used in future to identify soluble vacuolar proteins bound to the mutated receptor in the medium. In addition, diatom strains, in which CRISPR/CAS9 induced mutations in one vacuolar protein were identified, haue been initially characterized. Finally, we discussed, according to recent models, if the presence of ptVSRs in diatoms can contribute to the reconstruction of the evolution of the host cell of diatoms. Although the presence of ptVSRs in diatoms is hard to explain, we discuss several models for the origin of these receptors in diatoms and came to the conclusion that a green history of diatoms might explain their presence. Taken together, the outcome of this project provides a solid basis for further investigations in respect to the vacuolar proteome of diatoms, the targeting routes of vacuolar proteins, mutational analyses and the implications an the evolution the host of diatoms.
Publications
- (2016) Addressing various compartments of the diatom model organism Phaeodactylum tricornutum via sub-cellular marker Proteins. Algal Research 20:249-257
Liu X, Hempel F, Stork S, Bolte K, Moog D, Heimerl T, Maier UG,Zauner S
(See online at https://doi.org/10.1016/j.algal.2016.10.018) - (2017) The central vacuole of the diatom Phaeodactylum tricornutum: identification of new vacuolar membrane Proteins and of a functional di-leucine-based targeting motif. Protist 168:271-282
Schreiber V, Dersch J, Puzik K, Bäcker O, Liu X, Stork S, Schulz J, Heimerl T, Klingl A, Zauner S, Maier UG
(See online at https://doi.org/10.1016/j.protis.2017.03.001)
