EFFECTS OF CASKIN SCAFFOLD PROTEINS ON THE MOTILITY OF DENDRITIC FILOPODIA
CASK-interactive proteins, Caskin1 and Caskin2, are multidomain neuronal scaffold proteins. We have recently shown that Caskin proteins play an important role in hippocampal learning and memory formation via regulating AMPA receptor turnover. Our results also proved that deletion of both Caskin proteins in Caskin dKO mice led to the formation of smaller spine heads and reduced PSD length, raising the possibility that Caskin proteins regulate dendritic protrusion formation and maturation, as well. Dendritic filopodia are transient protrusions on dendritic shafts, which get stabilized upon establishing connections with nearby axons, leading to the formation of new synapses. In order to investigate whether Caskin1 is involved in the early stabilisation of dendritic filopodia, we performed dendritic filopodia motility analyses in Caskin dKO cultured hippocampal neurons. Cultures were transfected with V5-tagged wild-type and point mutant Caskin1 constructs to reveal the importance of upstream Caskin1 regulators. Motility of dendritic filopodia was followed by fluorescent live cell imaging using a semi-automated Image/Fiji plugin, Dendritic Filopodia Motility Analyzer (DFMA). Our results indicate that Caskin1 overexpression increased the motility of dendritic filopodia in a phosphorylation-dependent manner. Mimicking the PKA/PKC-mediated phosphorylation on the T1065 site by expressing the T1065E point mutant abolished this effect while blocking the 296 and 336 tyrosin-phosphorylation within the SH3 domain of Caskin1 further increased filopodial motility. These data indicate that Caskin1 may play a role in growth factor receptor-dependent signalling and synapse stabilisation. Supported by the National Brain Research Program (2017-1.2.1-NKP-2017-00002) and the VEKOP-2.3.3-15-2016-00007 grant provided by NRDIO, Hungary.