Functional effects of ZBP1 in mammary and neuronal cells and tissues

Abstract

RNA localization and subsequent regulation of expression occur in many cell types. This allows for variations in regional protein contents and specialization. Localization requires information in the RNA's sequence. Cis-acting sequences called the "zipcode", are recognized by trans-acting factors such as Zipcode Binding Protein-1 (ZBP1). ZBP1 binds to a portion of the 3' untranslated region of beta-actin mRNA to form ribonucleoprotein complexes that move and are anchored on the cytoskeleton. While ZBP1 has been shown to regulate mRNA localization and translation in cultured cells, we hypothesized that such fundamental regulation would be evident in tissues in the living animal. Because of ZBP1's role in cell polarity, migration, and localization we selected two well-characterized tissue models to investigate: a tumor where cell migration leads to metastasis, and the brain where highly polarized neurons require mRNA localization to distal synaptic locations.;We tested the first model by stably expressing a ZBP1 fusion protein in an unpolarized, metastatic tumor cell line. ZBP1 expression increased beta-actin mRNA localization and cell polarity while reducing metastasis. ZBP1 expressing cells also exhibited reduced chemotaxis, extending previous work that suggested different motility phenotypes result from differences in beta-actin localization.;We hypothesized that neuronal specific expression of ZBP1 would affect RNA localization in brain tissue and hence behavior. We examined striatal behaviors, where our transgenic mice express ZBP1 under the CaMKII promoter. We studied ZBP1's effects on cocaine-induced conditioned place preference. ZBP1 expression reduces the ability of mice to develop a preference in this assay, suggesting that ZBP1 may play an important role in addictive behavior, and providing insight into potential therapies for drug dependence and relapse. Since ZBP1 was expressed as a fusion protein with a FLAG tag, it was used to immunoprecipitate RNAs complexed with ZBP1. These RNA targets were analyzed using oligonucleotide microarrays, revealing specific mRNAs important in behavior.;The results of these investigations indicated essential systemic roles for ZBP1 that result from its ability to direct mRNA translation to cellular regions where the protein function is required.