Function of profilins in the tripartite synapse- from structural plasticity to functional modulation

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For a long time cell biologists have been motivated to understand how the process of cellular self-organization generates dynamic, robust and elaborate structures that organize cells. In particular there is a fundamental gap between our understanding of individual molecules and our understanding of how these molecules function collectively to form living neurons. The biological importance of establishing order over diverse length scales and timescales, as well as the challenges of understanding how systems of self-organizing molecules carry out cellular functions, is perhaps best illustrated by studies of the cytoskeleton. And here the actin cytoskeleton is of upmost important. It is crucially involved in motility in all animal cells and particular in neurons and astrocytes, where it is mandatory in processes of motility. Neurons react to different activity levels with structural changes e.g. neuronal differentiation, dendritic and axonal maintenance and plastic adaption of dendrites, axons and synapses. Modulators of the microfilament system mediate signals from the pre- or postsynaptic membrane to the actin cytoskeleton and by this means change function and structure of specific neuronal compartments. In this context the actin binding protein profilin could be a mayor player. In the central nervous system, two isoforms of profilin, PFN 1 and PFN 2a, are co-expressed. Recent studies focusing on the cellular role of profilin 1 and 2a in neurons and glia cells are showing that both isoforms possess overlapping as well as isoform specific functions. Furthermore, the findings obtained by knocking out only one of the isoforms could not exclude that each isoform could compensate the loss of the other one. This hypothesis was supported via the acute downregulation of each isoform by RNA interference which leads to results that are not completely in line with the knock out studies. Therefore we will here address the functional role of profilins in astrocytes and in neurons by its acute inactivation. By this means we will study the formation and modulation of the tripartite synapse and thereby their impact on neuronal function. To avoid compensatory effects of each other profilin we will knock out both isoforms simultaneously in a cell-type specific manner by a CRISPR/Cas9 mediated approach. We will investigate the function of the profilin isoforms in the translation of neuronal activity into cytoskeletal reorganization, specifically in processes of structural and functional synaptic plasticity via modern imaging and electrophysiological methods. We will in addition use behavioral tests in order to explore the possible role of profilin isoforms in hippocampus dependent learning tasks.