Architecture and secondary structure of an entire HIV-1 RNA genome

Abstract

Single-stranded RNA viruses encompass broad classes of infectious agents and cause the common cold, cancer, AIDS and other serious health threats. Viral replication is regulated at many levels, including the use of conserved genomic RNA structures. Most potential regulatory elements in viral RNA genomes are uncharacterized. Here we report the structure of an entire HIV-1 genome at single nucleotide resolution using SHAPE, a high-throughput RNA analysis technology. The genome encodes protein structure at two levels. In addition to the correspondence between RNA and protein primary sequences, a correlation exists between high levels of RNA structure and sequences that encode inter-domain loops in HIV proteins. This correlation suggests that RNA structure modulates ribosome elongation to promote native protein folding. Some simple genome elements previously shown to be important, including the ribosomal gag-pol frameshift stem-loop, are components of larger RNA motifs. We also identify organizational principles for unstructured RNA regions, including splice site acceptors and hypervariable regions. These results emphasize that the HIV-1 genome and, potentially, many coding RNAs are punctuated by previously unrecognized regulatory motifs and that extensive RNA structure constitutes an important component of the genetic code. The secondary structure of a complete HIV-1 RNA genome has been determined, based on analysis of authentic HIV RNA extracted from infectious virions. Secondary structures within single-stranded viral RNA genomes are known to serve functional and regulatory roles, but until now there has been no comprehensive analysis of total RNA for any virus. SHAPE technology (high-throughput selective 2′-hydroxyl acylation analysed by primer extension) was used to characterize all structures formed by the HIV-1 RNA genome. Numerous highly structured motifs were discovered, and functions can be inferred for many of these motifs. Importantly, the presence of RNA structural elements was found to influence the translation of proteins and to facilitate proper protein folding. These results emphasize that the HIV-1 genome does have a structure, elements of which are critical to viral fitness. Insights from this work could lead to a better understanding of HIV-1 biology and to new antiretroviral interventions. Single-stranded RNA viruses are responsible for the common cold, cancer, AIDS and other serious health threats. The genomes of these viruses form conserved secondary structures that have functional and regulatory roles, but most potential regulatory elements in viral RNA genomes remain uncharacterized. Here however, the structure of an entire HIV-1 genome at single nucleotide resolution is reported.