Bipartite promoters and RNA editing of paramyxoviruses and filoviruses

Abstract

A primary property of paramyxovirus bipartite promoters is to ensure that their RNA genomes are imprinted with a hexamer phase via their association with nucleoproteins, in part because this phase as well the editing sequence itself controls mRNA editing. The question then arises whether a similar mechanism operates for filoviruses that also contain bipartite promoters that are governed by the "rule of six," even though these genomes need not, and given Ebola virus biology, cannot always be of hexamer genome length. This review suggests that this is possible and describes how it might operate, and that RNA editing may play a role in Ebola virus genome interconversion that helps the virus adapt to different host environments.

Keywords: Ebola virus; RNA editing; bipartite promoters; genome interconversion; rule of six.

FIGURE 1.

(A) Paramyxovirus bipartite promoters. Schematic diagram of N-RNA 3′ ends and bipartite promoter sequences of SeV (representative of genera Respirovirus and Morbilivirus) and hPIV2 (genus Rubulavirus). The N subunits and the hexamer positions within each subunit are numbered from the genome 3′ end. Critical regions PE1 and PE2 indicate conserved elements of the bipartite promoters. The top right box displays the 6-nt “in” or “out” topology in the context of the nucleocapsid. On the diagrams, nucleotides in white point “out” whereas nucleotides in black point “in.” The strictly conserved nucleotides of the PE2 tripartite repeats are underlined. (B) Filovirus bipartite promoters. Schematic diagram of N-RNA 3′ ends and bipartite promoter sequences of EBOV 7U/6n + 5 and 8U/6n + 0; 6n + X referring to overall genome length. The 3′-terminal N of the 7U/6n + 5 genome is shown associated with only 5 nt and this genome 3′ end would start at hex2. An asterisk here denotes the empty hex1 (Fig. 2). On the diagrams, nucleotides in white point “out” whereas nucleotides in black point “in.” Little is known about EBOV PE1, and the underlined dinucleotides here could represent the critical PE1 elements of EBOV that differ in overall length by 1 nt needed for the initiation of RNA synthesis, as their precise distance from the strictly conserved uridine of PE2 (underlined) is maintained. The top right box displays the 6-nt “in” or “out” topology in the context of the nucleocapsid.

FIGURE 2.

Organization and replication of 7U/6n + 5 and 8U/6n + 0 EBOV genomes. The salient features of the EBOV cis-acting sequences PE1, PE2 (only a single repeat is shown for clarity) and the editing site for 7U/6n + 5 and 8U/6n + 0 genomes are shown (not drawn to scale). The proposed hexamer phases (large boxes) assume that genomes are assembled concomitantly with their synthesis, starting at their precise 5′ ends, 6 nt/protomer, thereby determining hexamer phase throughout. When genomes are less than 6n + 0 long, the 3′-terminal protomer contains less than 6 nt, as indicated (the asterisk represents an empty space). Nucleotide sequences that point out are in red; those that point in are in black. The underlined dinucleotides of PE1 could represent critical promoter elements of both EBOV needed for the initiation of RNA synthesis, as their precise distance from the strictly conserved uridine of PE2 (underlined) is maintained. The replication product (antigenome) of the 7U/6n + 5 genome is shown below. The critical RNA editing pause site is proposed to be the seventh U from the start (3′ end) of the U run (in red). The addition of one U to this U run in 8U/6n + 0 moves the proposed pause site by one hexamer position upstream (highlighted with dotted lines). Hexamer phase may then affect the editing phenotype differently in 7U and 8U genomes, promoting their interconversion.

FIGURE 3.

Interconversion of EBOV 7U/8U genomes. Models of the RNAs in and around RdRp's synthesis chamber at the editing pause site, the seventh uridine from the start of the U run (highlighted in black). The nascent product strand is shown below, 5′–3′, and the genome template strand (3′–5′) is on top; the separations in the sequences indicate hexamer phase. The seventh U (in bold black) is proposed as the site where RdRp pauses, forming a 7 bp RNA:RNA hybrid. Hybrid realignments that lead to adenosine insertions or deletions in the transcript can occur as indicated. See text for detail.

FIGURE 4.

Possible ways in which hexamer phase can affect EBOV RNA editing. Cartoon of U7/6n + 5 and U8/6n + 0 RNA synthesis elongation complexes when the active site of RdRp (internal oval) is paused at the proposed editing site; the seventh U of the U run, in red (vertical dashed lines). The eighth U of 8U/6n + 0 is also highlighted. The nascent transcript is shown below the template (bent arrow in green). During RNA synthesis, as RdRp moves (left to right) down the template, N protomers are continuously being separated from their associated nucleotides as the template RNA enters the template entry channel (on right side), and Ns reassociate with their cognate hexa-nucleotides as the template exits RdRp via its product exit channel (left side). These events must be highly coordinated, and these N:RNA interactions can affect the pace at which RdRp's active site arrives at, and remains at the pause site, before continuing strictly templated RNA synthesis. The template RNAs exiting RdRp are colored differently in U7/6n + 5 and U8/6n + 0 as they would be in different hexamer phases. The protomers of the RNA-free N chain are depicted crossing RdRp's top surface from right to left, and their hexamer positions are indicated. Possible protomer interactions with this surface will occur in a hexamer phase-dependent manner as schematized, and could affect how the nascent transcript 3′ end realigns with the template slippery sequence during the pause, as depicted in Figure 3.