title: New tools to analyze overlapping coding regions.
abstract: BACKGROUND
  Retroviruses transcribe messenger RNA for the overlapping Gag and Gag-Pol polyproteins, by using a programmed -1 ribosomal frameshift which requires a slippery sequence and an immediate downstream stem-loop secondary structure, together called frameshift stimulating signal (FSS). It follows that the molecular evolution of this genomic region of HIV-1 is highly constrained, since the retroviral genome must contain a slippery sequence (sequence constraint), code appropriate peptides in reading frames 0 and 1 (coding requirements), and form a thermodynamically stable stem-loop secondary structure (structure requirement).
  
  
  RESULTS
  We describe a unique computational tool, RNAsampleCDS, designed to compute the number of RNA sequences that code two (or more) peptides p,q in overlapping reading frames, that are identical (or have BLOSUM/PAM similarity that exceeds a user-specified value) to the input peptides p,q. RNAsampleCDS then samples a user-specified number of messenger RNAs that code such peptides; alternatively, RNAsampleCDS can exactly compute the position-specific scoring matrix and codon usage bias for all such RNA sequences. Our software allows the user to stipulate overlapping coding requirements for all 6 possible reading frames simultaneously, even allowing IUPAC constraints on RNA sequences and fixing GC-content. We generalize the notion of codon preference index (CPI) to overlapping reading frames, and use RNAsampleCDS to generate control sequences required in the computation of CPI. Moreover, by applying RNAsampleCDS, we are able to quantify the extent to which the overlapping coding requirement in HIV-1 [resp. HCV] contribute to the formation of the stem-loop [resp. double stem-loop] secondary structure known as the frameshift stimulating signal. Using our software, we confirm that certain experimentally determined deleterious HCV mutations occur in positions for which our software RNAsampleCDS and RNAiFold both indicate a single possible nucleotide. We generalize the notion of codon preference index (CPI) to overlapping coding regions, and use RNAsampleCDS to generate control sequences required in the computation of CPI for the Gag-Pol overlapping coding region of HIV-1. These applications show that RNAsampleCDS constitutes a unique tool in the software arsenal now available to evolutionary biologists.
  
  
  CONCLUSIONS
  Source code for the programs and additional data are available at http://bioinformatics.bc.edu/clotelab/RNAsampleCDS/ .
authors.affiliation: Biology Department, Boston College, 140 Commonwealth Avenue, Chestnut Hill MA, 02467, USA.
authors.forename: Amir H
authors.initials: AH
authors.lastname: Bayegan
authors.affiliation: Biology Department, Boston College, 140 Commonwealth Avenue, Chestnut Hill MA, 02467, USA.
authors.forename: Juan Antonio
authors.initials: JA
authors.lastname: Garcia-Martin
authors.affiliation: Biology Department, Boston College, 140 Commonwealth Avenue, Chestnut Hill MA, 02467, USA. clote@bc.edu.
authors.forename: Peter
authors.initials: P
authors.lastname: Clote
mesh_terms: D001483:Base Sequence; D003062:Codon / Q000235:genetics* / Q000378:metabolism; D019295:Computational Biology / Q000295:instrumentation / Q000379:methods*; D015658:HIV Infections / Q000821:virology; D015497:HIV-1 / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D056510:Position-Specific Scoring Matrices; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics*; D016364:Reading Frames; D012984:Software
chemical_list: D003062:Codon; D012367:RNA, Viral
keywords: Frameshift stimulating signal; HCV; HIV-1; Overlapping coding region; Ribosomal frameshift