Function of Cytoplasmic FMR1-interacting protein 2 (CYFIP2) in spinal motor neuron morphology during mouse embryonic development

Abstract

The development of the nervous system is a tightly regulated spatio-temporal process that involves actin cytoskeletal dynamics. The remodelling of the neuronal cytoskeleton is important for dendritic and axonal guidance, neurite elongation, dendritic arborization and synapse assembly in both the central and peripheral nervous system. Cytoplasmic FMR1-interacting protein 2 (CYFIP2) is a component of the WAVE regulatory complex and essential in regulating ARP2/3-dependent branched actin networks. CYFIP2 is highly expressed in the mouse CNS and the complete deletion of CYFIP2 leads to a perinatal lethal phenotype. However, the exact cellular role of CYFIP2 in mouse embryonic development is still not fully understood. <br /> We show in this study that CYFIP2 is a key component in peripheral nervous system development. The Cyfip2^-/- mouse model fails to properly innervate the diaphragm leading to respiratory failure of newborn mice. CYFIP2 is essential for axonal growth properties that are needed for stereotyped phrenic motor neuron branching pattern. The formation of focal adhesion sites in the spinal motor neuron growth cone requires proper interaction between actin adaptor proteins (i.e., vinculin), scaffold proteins (i.e., FAK), and CYFIP2-mediated ARP2/3 actin polymerization. In addition, CYFIP2 modulation of tyrosine phosphorylation signaling might be required to stabilize focal adhesion sites mediated by the ECM-integrin adhesion pathway. The structural composition of the presynaptic neuromuscular junction is also highly dependent on CYFIP2. <br /> The function of CYFIP2 in axonal elongation is cell autonomous in spinal motor neurons. However, A-to-I RNA editing of CYFIP2 in neurons (CYFIP2-K320E) could be a potential mechanism used to initiate neurite protrusion. Ultrastructural analysis demonstrated CYFIP2 is an important factor regulating the actin-microtubule crosstalk in spinal motor neuron growth cones and ensuring proper axonal outgrowth. Biochemical and histological assays showed CYFIP2 deletion did not alter the steady-state F-actin level in neurons but was mainly required for morphological properties. Furthermore, other actin nucleation/elongation factors, such as the CYFIP1-dependent WRC and MENA/VASP, did not compensate for the loss of CYFIP2. Proteomic analysis revealed novel CYFIP2 interaction partners which could help elucidate potential guidance signaling functions and cell adhesion mechanisms.