Molecular dissection of cytochalasan activities on actin and actin-binding proteins in situ and in vitro

• Applicants: Professor Dr. Klemens Rottner; Professorin Dr. Theresia Stradal
• Subject Area: Cell Biology ; Biochemistry ; Pharmacology
• Term: since 2026
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 582196148

Eukaryotic cell biology is fundamentally influenced by the cytoskeleton, particularly actin filaments and microtubules, which are essential for processes such as proliferation, cell division, metabolism, adhesion, endocytosis, and intracellular trafficking. While actin filament remodeling remains an area of active investigation, the precise roles of inhibitors, especially from the cytochalasan family of secondary fungal metabolites, are critical to understand. These insights not only have pharmacological importance but also contribute to developing molecular tools for studying actin dynamics. Actin filament polymerization initiates with a kinetically unfavorable step called nucleation, in which 3-4 actin monomers associate to form a filament stub, resulting in a polarized filament with distinct barbed (rapidly-growing) and pointed (slowly-growing) ends. The assembly of actin filaments is tightly regulated and largely restricted to certain subcellular locations by nucleators like the Arp2/3 complex and formins. Small molecular inhibitors, such as latrunculins and cytochalasans, disrupt these dynamics by sequestering monomers or inhibiting filament elongation, respectively. Despite being utilized for decades, there is a lack of comprehensive data explaining how the structural variations among different cytochalasans affect their activities and how actin and actin binding proteins (ABPs) respond to their binding. Cells express diverse actin structures linked to various remodelling processes, including contractile rings, stress fibers, and cell edge protrusions during migration. All actin structures undergo continuous turnover, balancing polymerization and depolymerization, making them susceptible to compounds that interfere with these processes. Cytochalasans, produced by a complex of polyketide synthase and non-ribosomal peptide synthetase, are bioactive hybrid compounds found in many Ascomycota species. With over 400 natural structures identified, cytochalasans can impact every actin structure, through predominantly interfering with the barbed end's growth. Cytochalasins A, B, C, and D were first described in 1967 and shown to inhibit cytokinesis (causing multinucleation) and cellular motility. Their effects are mirrored by tryptophan-bearing cytochalasans, such as chaetoglobosins A-F, which also induce multinucleation in cells. Finally, some cytochalasans such as Aspochalasin D in vitro do not bind to actin at all. Understanding these mechanisms will illuminate the complex interactions of cytochalasans with actin structures and their regulatory proteins, providing insight into cellular functions and potential therapeutic applications. The current application deals with the elaborate, detailed analysis of both actin binding and non-actin binding cytochalasans and extends to identify the protein targets of these compound representatives. We will also dissect the targets mediating cytotoxicity and those that account for altering actin dynamics.

• DFG Programme: Research Grants