The actin cytoskeleton contributes to essential processes across eukaryotes. Our current understanding of the actin cytoskeleton is based on experiments that were performed using just a handful of species. While most cytoskeletal research has aimed at understanding human health, researchers have discovered and characterized many components using single-celled organisms. For example, capping protein and the branched actin-nucleating Arp2/3 complex were first identified and purified from the single-celled amoeba Acanthamoeba castellanii  and actin-binding coronin was discovered in the slime mold Dictyostelium discoideum . The multifunctional actin-binding protein twinfilin was discovered in the budding yeast Saccharomyces cerevisiae .
Recent work has illuminated unique characteristics of the actin cytoskeleton in unicellular, green algae Chlamydomonas reinhardtii, which expresses two separate compensatory actins and lacks any known activators of the Arp2/3 complex , which is required for nucleating actin branches in other organisms. Microalgal species have evolved from diverse ancestral sources, and many of these species have uniquely divergent cytoskeletal characteristics. This and their easy maintenance in a lab setting make microalgae an ideal system to study the evolution of the actin cytoskeleton.
The following questions guide this project:
Can we identify novel actin regulators in diverse species lacking canonical regulators?
The spatial and temporal regulation of actin filament assembly, disassembly, and modification is essential across eukaryotes. Many of the proteins responsible for regulating these processes are conserved across species; however, recent work has uncovered that several organisms lack clear homologs to these important regulators . Among these regulators is the Arp2/3 complex, which nucleates branched actin filaments necessary to carry out endocytosis, phagocytosis, cell motility, and many more functions . This complex is inactive without a class of proteins called nucleation promoting factors (NPFs) . Surprisingly, several algal species lack clear homologs to any known NPFs, raising the question of how this complex is regulated in a large group of organisms.
Can we uncover novel actin-dependent functions using diverse organisms that carry out unique cellular processes?
Actin filament polymerization is essential for many essential cellular functions, including division, adhesion, and motility . Although actin was first described over a century ago , new contributions of the actin cytoskeleton continue to be discovered, including involvement in chloroplast movement  and DNA damage repair . The actin cytoskeleton’s contributions to microalgal cell functions remain poorly understood and further research could uncover contributions of actin assembly and disassembly to unique functions.
Actin polymerization is important for cell motility, usually through the formation of branched actin networks at the leading edge of crawling cells. Many microalgal species have cell walls and move instead by microtubule-based flagellar motility. However, some members of the chlorarachniophyte family perform both flagellar motility and amoeboid motility ! Working with multiple species from this family, including members of the genera Bigelowiella, Amorphochlora, Lotharella, and Gymnochlora, we have discovered that members of this family form light-dependent extensions through putative branched actin nucleation to perform cell motility and predation.
Learn more about these findings:
The actin cytoskeleton contributes to the structure of many cells. Many algal species have unique and intricate morphologies, especially diatoms. Phaeodactylum tricornutum is one of the first diatoms with a sequenced genome and is found in three main morphotypes, making it an ideal species to study. Interestingly, while this species has homologs to actin and various actin-binding proteins, it lacks homologs to the Arp2/3 complex subunits. Surprisingly, we observed several P. tricornutum cells forming protoplasts and separating from their cell wall when treated with the myosin II inhibitor (-)-blebbistatin and the Arp2/3 complex inhibitor CK-666. However, recent attempts to replicate these findings have been unsuccessful, likely due to the sensitivity of cell wall-less protoplasts under harsh conditions.
Additionally, members of the order Chlamydomonadales express a conventional actin and a divergent, non-conventional actin. These cells also contain homologs to most of the Arp2/3 complex subunits and recent work has shown Arp2/3 complex function is important for Chlamydomonas reinhardtii physiology . Interestingly, while regulation of this complex is vitally important, Chlamydomonadales lack any clear homologs to the Arp2/3 complex-activating nucleation-promoting factors (NPFs) . We are using three Chlamydomonadales species (Chlamydomonas reinhardtii, Gonium pectorale, and Volvox carteri) to understand how the Arp2/3 complex is regulated in species lacking known NPFs.
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