Conserved RNA secondary structures and long-range interactions in Hepatitis C Viruses

Supporting Data

Markus Fricke, Nadia Dünnes, Margarita Zayas, Ralf Bartenschlager, Michael Niepmann, Manja Marz

Data

Initial data
• Alignment from HCV DB (http://hcv.lanl.gov/content/sequence/NEWALIGN/align.html)
• Decryption of names and fasta header
• All HCV from NCBI and HCV DB FASTA
• All HCV from NCBI and HCV DB Alignment
• All HCV from NCBI and HCV DB Tree (nexus format)

Selected data
• [txt] List with chosen HCV from NCBI and HCV DB
• [fasta] Fasta with chosen HCV from NCBI and HCV DB
• [stk|html] MAFFT Alignment with chosen HCV from NCBI and HCV DB

  LocARNA

  • LocARNA Alignment 0-358 (+)
  • LocARNA Alignment 0-365 (-)
  • LocARNA Alignment 330-520 (+)
  • LocARNA Alignment 9399-9820 (+)
  • LocARNA Alignment 10000-10101 (+)
  • LocARNA Alignment 10000-10101 (-)

GTF Annotation
• MAFFT HCV-Alignemnt Annotation (gtf) annotated according to con1 isolate (AJ238799)

[Dat. 1]
• Variable region (Stokholm alignment). Each sequences is folded seperatly (#=GR).

Color-code

Colors are used to indicate conserved base pairs: from red (conservation of only one base-pair type) to purple (all six base-pair types are found); from dark (all sequences contain this base pair) to light colors (1 or 2 sequences are unable to form this base pair).

[Fig. 1]

A	B	C
(A) Alignment based secondary structure prediction of the 5’ UTR with 86 isolates. The alignment was calculated by LocARNA, the consensus sequence and structure by RNAalifold. The structure was forced to build the already known IRES structure. Blue - Forced to be unpaired. Magenta - Forced to be paired.	(B) RNAalifold base pairing probability matrix of the 5' UTR,based on 86 isolates. The upper half of the matrix shows all base pair probabilities lower 10^−6. The lower half shows the base pair probabilities of the MFE structure.	(C) Part of the SHAPE analysis of Pang 2011.
D
(D) Alignment based secondary structure prediction of the 5’ UTR with 86 isolates (unbiased). The alignment was calculated by LocARNA, the consensus sequence and structure by RNAalifold.

[Fig. 2]

A	B
(A) Alignment based secondary structure prediction of the 5’ UTR and the beginning of the core region with 77 isolates. The alignment was calculated by LocARNA, the consensus sequence and structure by RNAalifold. The pseudo knot is forced to be unpaired (blue).	(B) RNAalifold base pairing probability matrix of the 5' UTR and the beginning of the core region with 77 isolates. The upper half of the matrix shows all base pair probabilities lower 10^−6. The lower half shows the base pair probabilities of the MFE structure.
C
(C) Secondary structure prediction of the first 177\,nt of the core region based on an alignment with 106 isolates.

[Fig. 3]

A	B
(A) Consensus sequence and secondary structure of the CRE, VR and X-tail, based on 19 isolates.	(B) RNAalifold base pairing probability matrix of the CRE, VR and X-tail region, based on 19 isolates. The upper half of the matrix shows all base pair probabilities ≤ 10 −6 . The lower half shows the base pair probabilities of the MFE structure.
C	D
(C) Consensus sequence and secondary structure of the CRE, VR without constraints, based on 96 isolates.	(D) RNAalifold base pairing probability matrix of the CRE, VR, based on 96 isolates. The upper half of the matrix shows all base pair probabilities ≤ 10 −6 . The lower half shows the base pair probabilities of the MFE structure.

[Fig. 4]

A

RNAalifold base pairing probability matrix of the X-Tail, based on 19 isolates. The upper half of the matrix shows all base pair probabilities ≤ 10 −6 . The lower half shows the base pair probabilities of the MFE structure.

[Fig. 5]

RNAalifold base pairing probability matrix of the 5' end of the minus strand, based on 19 isolates. The upper half of the matrix shows all base pair probabilities lower 10^−6. The lower half shows the base pair probabilities of the MFE structure.

[Fig. 6]

Extended long-range interaction of LRI 14 with 104 isolates.

[Fig. 7]

A

B

C

RNAalifold base pairing probability matrix of a artifical 5'UTR / CRE (A), 5' UTR / X-Tail 1 (B) and 5'UTR / X-Tail 2 (C). The alignments are concatinated for each region from at least three consecutive non-pairing nucleotides, added with two flanking nucleotides, separated with `NNN'. Based on each matrix we picked the most probable long range interactions, recalculate the selected interactions with LocARNA and filtered the results manually.

Newly added for the Reviewers

[Fig. 8]

A	B
C	RNAalifold consensus structure of the CRE region (96 isolates) based on LocaRNA (A) on DAFS (B) and Lara (C) to compare the different secondary structure based alignment tools.

[Fig. 9]

PETcofold probability matrix of the 5’UTR and the CRE-region based on 80 isolates.