New insights into direct cell surface-to-chloroplast trafficking of organic and inorganic iron substrates in marine diatoms

Andrew Allen1, Jernej Turnsek2, Tyler Coale3, Katherine Barbeau4, John K. Brunson5, Vince A Bielinksi2 and Robert H Lampe6, (1)J. Craig Venter Institute La Jolla, La Jolla, CA, United States, (2)J. Craig Venter Institute La Jolla, La Jolla, United States, (3)University of California San Diego, Scripps Institution of Oceanography, La Jolla, CA, United States, (4)Scripps Institution of Oceanography, Geosciences Research Division, La Jolla, CA, United States, (5)Scripps Institution of Oceanography, La Jolla, CA, United States, (6)Scripps Institution of Oceanography, La Jolla, United States
We recently showed that diatoms utilize phytotransferrin to access inorganic iron with remarkable efficiency, but this process is contingent on carbonate ion concentration. As ocean acidification reduces carbonate concentration, inorganic iron uptake may be discouraged in favor of carbonate-independent uptake. We describe components of a reduction-dependent siderophore acquisition pathway that is not sensitive to carbonate availability but relies on a bacterial derived receptor protein and provides a viable alternative to inorganic iron uptake under certain conditions. This form of iron uptake entails close association between diatoms and siderophore producing organisms during low iron conditions. Understanding the caveats associated with iron source preference in diatoms will help predict the impacts of climate change on microbial community structure in high nitrate low chlorophyll ecosystems. While recent studies have confirmed endocytosis to be required for acquisition of both inorganic and organic iron, proteins downstream of these cell surface associated uptake systems are unknown. Iron uptake related vesicles have been observed, but the proteins present in these compartments are largely unknown. We applied APEX2-based subcellular proteomics to catalog proximal proteins of phytotransferrin in the model diatom Phaeodactylum tricornutum. Proteins encoded by poorly characterized iron-sensitive genes were identified including three that are expressed from a chromosomal gene cluster. In subsequent experiments, two of these showed unambiguous colocalization with phytotransferrin in a region adjacent to the chloroplast. Further phylogenetic and domain analyses suggest they are likely involved in intracellular iron processing. Data related to the putative role and function of other proteins found to associate with iron trafficking vesicles will be discussed. Taken together, our ­­­data enable proposition of a model connecting cell surface iron binding, internalization and intracellular trafficking, followed by offloading to the chloroplast in Phaeodactylum tricornutum.