RNA structure regulates HIV-1 reverse transcription initiation
Abstract/Contents
- Abstract
- Reverse transcription of the human immunodeficiency virus-1 (HIV-1) RNA genome into double-stranded DNA is an essential, highly regulated step in the viral lifecycle and a common target of antiretroviral drugs. The reaction is catalyzed by viral reverse transcriptase (RT) that is packaged into an infectious virion along with two copies of viral genomic RNA (vRNA) and host transfer RNA lysine 3 (tRNALys3), which serves as the primer for the initiation of reverse transcription. Initiation is distinct from elongation as it is slow and non-processive, with discrete stalling points along the vRNA sequence. Studies over several decades have revealed regions of RNA structure and sequence complementarity between the vRNA template and tRNALys3 primer that appear to regulate reverse transcription initiation. Despite this body of work, the structural basis of RT function during initiation has remained a mystery. Using cryo-electron microscopy, we have determined the structures of several reverse transcriptase initiation complexes representative of key initiation intermediates. In all structures, the primer binding site (PBS) helix formed between tRNALys3 and vRNA lies in the cleft of RT and is extended by additional pairing interactions. We find that during early stages of initiation, the 5′ end of the tRNA refolds and stacks on the PBS to create a long helical structure, while the remaining viral RNA forms two helical stems positioned above the RT active site. Structures of subsequent steps in the initiation process feature a dramatic refolding of the tRNA and vRNA that may govern the transition from initiation into the processive elongation phase. Newly obtained data on the reverse transcriptase initiation complex reconstituted using naturally post-transcriptionally modified tRNALys3 further supports the hypothesis that refolding of vRNA and tRNALys3 does not occur during early stages of initiation. Our results illustrate how RNA structure in the initiation complex alters RT conformation to decrease activity, highlighting a potential target for drug action.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2019; ©2019 |
| Publication date | 2019; 2019 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Larsen, Kevin |
|---|---|
| Degree supervisor | Puglisi, Elisabetta |
| Degree supervisor | Puglisi, Joseph D |
| Thesis advisor | Puglisi, Elisabetta |
| Thesis advisor | Puglisi, Joseph D |
| Thesis advisor | Brünger, Axel T |
| Degree committee member | Brünger, Axel T |
| Associated with | Stanford University, Biophysics Program. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Kevin Larsen. |
|---|---|
| Note | Submitted to the Biophysics Program. |
| Thesis | Thesis Ph.D. Stanford University 2019. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2019 by Kevin Larsen
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