Affiliation: Southern Illinois University Edwardsville
Keywords: e. synaptic function and organization; k. small GTPases
Chromodomain Helicase DNA-Binding Domain (CHD) proteins are chromatin remodelers that are critical for early neuronal development and the expression of genes important for synaptic plasticity. Mutations in CHD7/8 can lead to a variety of disorders in humans such as CHARGE syndrome, a neurodevelopmental disorder, and autism spectrum disorders. Mutations in kismet (kis), the Drosophila ortholog of CHD7/8, lead to an accumulation of cell adhesion molecules and the transmembrane protein amyloid precursor protein-like (APPL) at the synapse as well as reductions in endocytosis, synaptic transmission, and larval locomotion. Kismet regulates rab11 expression. Rab11 is a GTPase that traffics vesicles from the recycling endosome to the plasma membrane. Because of its role in membrane trafficking, Rab11 might facilitate some of these phenotypic changes observed in kismet mutants.
To investigate this possibility, we expressed constitutively active or dominant negative isoforms of Rab11 and examined Fasciclin II (FasII), a cell adhesion molecule. Similar to kis mutants, constitutively active Rab11 increased synaptic FasII. Expression of dominant negative Rab11, however, decreased FasII protein levels. When rab11 was knocked down, we found a decrease in APPL, which directly contrasts kis mutants that show an increase in APPL levels. In addition, when Rab11 dominant negative and constitutive active isoforms were expressed, we saw no change in endocytosis or endocytic proteins Endophilin A (EndoA) and Dynamin (Dyn) unlike kis mutants, which show reductions in each. Kis affects genes, possibly rab11, that localize synaptic proteins to phosphatidylinositol-4,5- P2 (PIP2), a phospholipid that helps organize proteins into lipid rafts in the plasma membrane. When rab11 was knocked down, we did find a reduction in PIP2 levels at the synapse. These data may suggest that while Rab11 might only play a minor role in contributing to kis mutant phenotypes, Rab11 may be able to restore kis mutant phenotypes. Therefore, we will express wildtype Rab11 in kis mutants to determine whether Rab11 can restore the synaptic levels of FasII and APPL.