Abstract 3610: GW182 associates with circulating microvesicles and microRNA in human plasma

To date, microRNAs (miR) in human plasma have been discovered within circulating microvesicles (cMV), bound to Argonaute 2 and associated with HDL and LDL microvesicles. The protein GW182 shares an association with both multivesicular bodies and the Argonaute family of proteins. GW182 has the capacity to bind all human Argonaute proteins (1-4) and their associated miRs. In the cell, GW182 is associated with the membrane of multivesicular bodies and has the ability to congregate Argonaute-loaded RISC complexes. In addition, GW182 has been observed on the surface of purified exosomes. Here we investigated the relationship of GW182 with Argonaute and cMV in human plasma. A monoclonal antibody directed toward GW182 was used to capture the protein. This isolate also contained Argonaute proteins as determined by Western analysis. The co-precipitation of GW182 and Argonaute suggests that these two proteins retain their functional relationship in plasma. RNA was then isolated from precipitates for miR detection and analysis. The GW182-associated miR profile from human plasma contained individual miRs whose abundance either equaled or surpassed that of their matched Argonaute 2 immuno-precipitated miRs. This implies that GW182 maintains an association with the family of Argonaute proteins and a subset of cMV in human plasma. Phenotypic analysis of GW182-associated human plasma microparticles was performed using flow cytometry with a Beckman Coulter MoFlo XDP. A subpopulation of plasma-derived cMV was observed that co-expressed tetraspanins, GW182 and Argonaute 2. Since tetraspanins are transmembrane proteins highly associated with cMV, these results suggest that both GW182 and Argonaute can associate with cMV in human plasma. Thus, precipitation of GW182 enables the isolation and purification of miRs from human plasma including those bound to Argonaute 1-4 and a subset of cMV. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3610. doi:1538-7445.AM2012-3610