Aptamer tools for the study of HIV capsid structure, function, and interactions.

About This Grant

HIV remains a significant problem worldwide. The CA protein of HIV is involved in several critical replication events, including Gag oligomerization and viral assembly, maturation, reverse transcription (RT), trafficking to the nucleus via interaction with host factors, nuclear import, integration, and evasion of host immune responses. CA’s numerous roles are facilitated by its ability to adopt distinct structural forms at different steps of replication. Despite significant advances in our understanding of the role of CA in replication, there are many unresolved questions regarding CA structural dynamics during viral assembly and post-entry replication steps, CA-host factor interactions, and the impact of these interactions on virus biology. The genetic fragility of CA, a lack of tools for specific CA structural forms, and difficulty of examining CA-host interactions in cells represent significant barriers to the resolution of these questions. To address these challenges, have developed novel RNA aptamer tools capable of discriminating among distinct CA structural forms. Aptamers are uniquely well-suited to the study of CA, as they bind targets with high specificity, discriminate among different conformations of the same protein, can be expressed in or delivered to cells, and are amenable to a variety of different modifications. Further, they have significant applications for biosensing due to their dynamic conformational variability, for which we provide proof-of-concept. This proposal will 1) elucidate the molecular basis for aptamer-CA interactions, 2) leverage aptamer programmability and specificity for the development of cutting-edge molecular tools, and) determine the biological significance of aptamer-CA interactions to inform novel therapeutic targets. If successful, this work will provide a complete panel of CA structure form-specific aptamers, including those that bind pentamer or in the presence of LEN and/or host factors, provide aptamer-CA interaction maps that define aptamer-targeted epitopes along with the biological significance of these interactions, develop aptamer-based biosensor technology for detection of CA structural forms in specific cellular compartments, and resolve key questions regarding proposed CA structure form-specific host interactions. Importantly, this work will set the stage for future innovations in CA structure form-specific detection, including high throughput mapping of accessible CA epitopes using aptamer barcoding, multiplexed detection of CA structural forms using aptamer- based biosensors, in-home diagnostics with improved sensitivity, and cost-effective, aptamer-based detection methods to measure CA structural forms or detect viral resistance, among others. Collectively, this study will provide exciting new molecular tools and insights into CA, informing future drug design strategies, as well as biological interactions and mechanisms.