This project focused on the evolution and selection of a particular metalloenzyme family, the superoxide dismutases (SOD), which utilize the redox sensitive and thus biogeochemically significant metals Fe, Mn, Cu/Zn, or Ni.  I successfully showed that in the secondary red algae, diatoms, historically high Mn measurements could be accounted for by the protective role and constitutive expression of MnSODs in the plastid.  This was previously unknown to the field (Wolfe-Simon et al., 2006).  In addition, the importance of Mn to secondary red algae also suggests a possible role for Mn in their ascent to dominance of the marine system during the Mesozoic.  Furthermore, the Mn and FeSOD genes are related due to an ancient gene duplication event.  I reviewed (Wolfe-Simon et al., 2005) and extended our understanding of the molecular evolution and phylogenetic radiation of the Mn and FeSOD isoforms in photoautotrophs by running detailed maximum likelihood and Bayesian phylogenetic analyses (Wolfe-Simon et al., in review).  The eukaryotic subcellular residence of the proteins today suggests that they were inherited originally from the symbiont ancestor which led to the rise of the modern eukaryotic organelles (Wolfe-Simon et al. 2006).

[Collaborators: ...]

Did trace metal availability support the dominance of cyanobacteria during a substantial amount of Earth's early history?  Currently, my data suggest that cyanobacteria have biological mechanisms to cope with reduced Fe and reduced Cu conditions as compared to algae and other photosynthetic eukaryotes (Wolfe-Simon et al., in prep).  Cyanobacteria dominated the oceans until approximately 0.5-0.8 billion years ago (Ga) despite drastic environmental changes.  Eukaryotic photoautotrophs evolved during this time (2.7-1.5 Ga) but their rise to ecological prominence and multicellularity was delayed. Biochemical strategies to cope with changing metal availability may have been critical. A major change in metal availability was the decrease in ocean Fe after 1.8 Ga. This decrease would have been accompanied by a decrease in Cu if the oceans became euxinic (reduced and sulfur rich) at this time.  Hence, I am investigating differences in the ways that cyanobacteria and algae cope with Fe and Cu stress. I found that the Cu quota of the cyanobacterium Synechocystis PCC6803 does not increase under Fe-deficiency. This contrasts with previous studies with eukaryotic photoautotrophs showing increased use of Cu under Fe-deficiency.  For cyanobacteria, production of compensatory Cu proteins presumably involves reallocation of intracellular Cu rather than increased Cu uptake.  The implication is that cyanobacteria were better equipped to cope with Fe ocean redox changes at this time when Cu may have been scarce.

How did the redox conditions of the environment, and thus micronutrient availability, help drive the rise of algae?  As trace metal concentrations have varied over Earth history, a major geochemical trend is the shift from abundantly soluble Fe to Cu due to the affect of increased oxygen in the atmosphere over geologic time (see above).  There is evidence in the literature that algae and other photosynthetic eukaryotes biochemically utilize Cu in a stoichiometrically larger proportion than cyanobacteria.  I have tested the evolutionary validity of this hypothesis and shown the increased cellular quota of Cu as well as the more economical and efficient use of Cu in green algae when compared to cyanobacteria.  For example, green algae increase their expression of the PSI Cu protein plastocyanin when under Fe-deficiency to replace the Fe-containing protein cytochrome c₆.  Furthermore, they modulate their expression of other Fe-rich PSI proteins including PsaA and psaC.   Another example relates back to my doctoral work in that higher plants utilize CuZnSOD while more primitive plants (including some early eukaryotic photoautotrophs) rely on the constitutive expression of FeSOD.  By combining controlled laboratory experiments, biochemical analyses, and molecular genetic modeling I will be able to forge a deeper understanding of historically relevant events for the Proterozoic evolution of photosynthetic life.

[Collaborators: Ariel Anbar, Petra Fromme, Vim Vermaas]

Nitrogen is vital for all known oxygenic photoautotrophs. The three main pathways leading to nitrogen assimilation all require Mo. Nitrogenase (Nif), responsible for the biological fixation of N₂ gas to ammonia, is known to posses one atom of Mo per functional subgroup. Nitrate reductase (NarB) also has one atom of Mo and reduced nitrate to nitrite for further processing by the cell. MORE TO COME. This project forms the basis for the PhD thesis of Jennifer Glass. Jennifer is a PhD candidate working under the guidance of Ariel Anbar in the School of Earth and Space Exploration at Arizona State University. Jennifer worked with me during my postdoctoral fellowship at ASU and continues to be a close collaborator. For further information on this project, please follow this link.

[Collaborators: Jennifer Glass, Ariel Anbar]

Arsenic as a prebiotic chemical analog of phosphate (Wolfe-Simon et al 2008). Essentially, Arsenic (in the oxidized 5+ state as arsenate) is biologically, so similar to phosphate that many enzymes cannot recognize the difference. This constitutes the basis for much of the toxicity of arsenate and so most detoxification pathways in biology aim to reduce arsenate to more volatile forms for easier removal from biological systems. However, due to the increased mobility of reduced arsenic species, often the toxicity of arsenic increases as the redox state decreases.

Inorganically, As and P have very similar atomic radii and electronegativity (especially, compared to nitrogen which is directly above both on the periodic table). In fact, in contrast to phosphorus, arsenic readily forms sulfides and thus may have been available to early life at hydrothermal vents and similar environments. Given the distinct similarity between these two elements my coauthors and I assemble plausible TESTABLE hypotheses regarding the liklihood of life arising to either originally incorporate arsenate in a functional sense, and/or more speculatively, alternative forms of life utilizing a genetic system entirely based on arsenic. Check out our manuscript and decide for yourself!

To futher this hypothesis, we have embarked on two different approaches to test assimilatory arsenic utilization. Firstly, we are collaborating with Dr. Steve Benner and Dr. Nicole Leal at The Foundation for Applied Molecular Evolution to measure the spontanous incorporation of arsenate into a DNA backbone. In concert to this purely in vitro approach, we are also examining arsenate-rich/phosphate-poor environments to hunt and enrich cultures for organisms utilizing arsenate in novel and unique modes.

[Collaborators: Ariel Anbar, Steve Benner, Nicole Leal, Paul Davies, Dirk Schulze-Milch, Ann Pearson.]