In collaboration with Dr. Faklers laboratory (Institute of Physiology, University of Freiburg, Freiburg, Germany), we continue the investigation of the effects of Olfm1, Olfm2, Olfm3, and Vwc2 null mutations on the composition and activity of the AMPA receptor complex. We demonstrated that although elimination of Olfm1 leads to profound changes in the composition of the AMPA receptor complex in both brain and retina, Olfm1 does not affect AMPAR conductance and desensitization properties in HEK293 cells transfected with corresponding expression constructs in vitro. We demonstrated that mouse and zebrafish Vwc2 and Vwc2l were co-precipitated with Olfm1 from lysates of COS7 cells transiently transfected with corresponding expression constructs indicating direct interaction of Olfm1 with Vwc2 and Vwc2l. Injection of vwc2l mRNA into zebrafish eggs and subsequent overexpression during early development resulted in severe developmental defects, mainly in the posterior larvae. These defects were remarkably reduced when the vwc2l mRNA was injected into olfm1 knockout eggs, indicating that the effects of vwc2l over- or mis-expression may be modulated by Olfm1. Overexpression of vwc2l may affect TNF signaling as suggested by the results of RNAseq analysis of zebrafish larva heads 24 hours post-fertilization. TNF signaling is known to be involved in neuronal degeneration in brain and eye in various diseases and in particular, during glutamate excitotoxicity. Moreover, glia-derived soluble TNF was shown to be a major inducer of RGC death through the activation of a Ca2+-permeable AMPA receptor subtype. Another potential target of vwc2l overexpression is SREBP, a transcription factor responsible for synthesis of cholesterol and other lipids. Cholesterol synthesis is known to be important for maintaining healthy neuronal membrane structure in axons and dendrites as well as lipid rafts in synapses. Our data suggest that vwc2l has multiple functions in both neural and non-neural tissues. The characterization of Vwc2 and Vwc2l effects on the AMPA receptor activity is currently under investigation.
