Self-Organization of Cellular Structures

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Self-Organization of Secretory Compartments.

Y. Liu1, N. Bharucha1, E. Papanikou1, B. S. Glick1; 1Molecular Genetics and Cell Biology, The University of Chicago, Chicago, IL

During the past decade, the concept of self-organization has been increasingly used to describe the biogenesis of secretory compartments. We have applied this concept to analyze transitional ER (tER) sites and the Golgi apparatus. These studies have focused on two budding yeasts: Pichia pastoris, which has a few large tER sites that are linked to Golgi stacks, and Saccharomyces cerevisiae, which has numerous small tER sites and a nonstacked Golgi. We found that P. pastoris tER sites are long-lived structures that form de novo, fuse upon collision, and grow or shrink to attain a steady-state size. This behavior can be explained by a self- organization model. Specifically, we postulate that capture of new tER components is balanced by shrinkage driven by the budding of COPII coated transport vesicles. To test this model, we used a genetic screen to identify Sec16 as a key player in tER organization. Sec16 is a large peripheral ER membrane protein that interacts with multiple COPII components. Our results suggest that Sec16 partitions between the ER membrane and the cytosol, and that the membrane-associated form acts as a negative regulator of ER export. According to this view, Sec16 functions primarily to control tER dynamics rather than to nucleate tER site formation. Interestingly, the redistribution of Sec16 to the cytosol is influenced by association of the central conserved domain of Sec16 with the COPII component Sec13. The combined data imply that Sec16 defines a new level of regulation for the ER export system. To study Golgi organization, we took advantage of the nonstacked Golgi cisternae in S. cerevisiae. Our 4D confocal microscopy studies revealed that Golgi cisternae mature by progressively acquiring and then losing early resident Golgi proteins followed by late resident Golgi proteins. Recently, we have found that early Golgi compartments are closely associated with tER sites even in S. cerevisiae, consistent with the idea that Golgi cisternae assemble by the coalescence of ER-derived vesicles. However, the mechanisms of subsequent Golgi maturation are more obscure. One possibility is that COPI coated vesicles drive the recycling of resident Golgi proteins from older to younger cisternae, but the experimental evidence for the role of COPI has been ambiguous. We are now using new approaches to achieve rapid and selective inactivation of COPI and other key trafficking components in S. cerevisiae. The long-term goal is to elucidate the cascade of interactions that enables the cell to rebuild the Golgi apparatus with each round of transport.

56

Spatiotemporal Integration of Chemical and Mechanical Signals in Cell Migration.

G. Danuser1; 1Department of Cell Biology, Harvard Medical School, Boston, MA

Cell migration is driven by the self-organization of innumerable, coupled chemical and mechanical processes. Chemical signals activate and deactivate the assembly and disassembly of cytoskeleton polymer networks. Both assembly and disassembly generate mechanical forces that push parts of the cell forward and retract others. They also define the mechanical properties of the cytoskeleton, which in turn determine how forces translate into cell shape deformation and movements. Chemical signals modulate the activity of molecular motors that produce contractile forces. By pulling on the polymer networks, these same motors organize the architecture of the cytoskeleton. This affects again the mechanical properties of the cytoskeleton. Motors may even mediate polymer disassembly, inducing a secondary contractile response of the cytoskeleton. All these entangled mechanical outputs feed back into the activation of the upstream chemical signals, a process that can be referred to as cell intrinsic mechanotransduction. Decades of biochemical, genetic and molecular analyses have generated the parts lists for most of these chemical and mechanical processes as well as hypotheses of process interactions. However, there is still very limited understanding as to how the processes are spatially and temporally organized in a system with emergent properties. There are multiple challenges associated with addressing this question: First, the process hierarchies are transient and distributed over multiple time and length scales. Thus, new technology is required to monitor multiple processes working in parallel but at different time points and cellular locations; and analytical tools are needed to extract from these data the spatial, temporal, and functional linkages between processes. Second, because of the nested nonlinear interactions among processes, the stimulation or perturbation of a particular process component often propagates in complex ways, obscuring the relation between system response and component function. For the past ten years my lab has made attempts to implement approaches that tackle these issues. We have invested in the development of quantitative multi-parametric live cell imaging to acquire simultaneous measurements of cell morphodynamics, cytoskeleton dynamics, resulting intracellular forces, and chemical signals. We also have worked on multiplexing methods to couple these measurements across different experiments, allowing us to “stitch together” larger process models. Very recent work in the lab has focused on novel mathematical tools to predict complex nonlinear process hierarchies. Central to our approach is the use of minimally perturbing experimental strategies that avoid the ambiguities of nonlinear system responses induced by stronger interventions. This talk will highlight some of the most surprising discoveries we have made with this work and will give an outlook to the many challenges we are still facing in order to establish a comprehensive understanding of cell migration.

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Modeling cytoskeletal structures with Cytosim.

F. Nedelec1; 1Cell Biology and Biophysics, EMBL, Heidelberg, Germany

Living cells have a system of fibers and associated proteins, called the cytoskeleton, that provides an essential mechanical support during migration, polarization, division, etc. Cytoskeletal fibers form spontaneously by self-assembly of monomers, and multiple fibers can further self-organize into assemblies of cellular scale. In this talk, we will introduce cytosim, a simulation software with the aim to study systems of many fibers. We will explain how the user can easily configure the simulation with simple examples. We will also illustrate techniques that can be used to systematically explore the possibilities embedded in a cytoskeletal system.