Origin and evolution of the RIG-I like RNA helicase gene family

Abstract

Background: The DExD/H domain containing RNA helicases such as retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are key cytosolic pattern recognition receptors (PRRs) for detecting nucleotide pathogen associated molecular patterns (PAMPs) of invading viruses. The RIG-I and MDA5 proteins differentially recognise conserved PAMPs in double stranded or single stranded viral RNA molecules, leading to activation of the interferon system in vertebrates. They share three core protein domains including a RNA helicase domain near the C terminus (HELICc), one or more caspase activation and recruitment domains (CARDs) and an ATP dependent DExD/H domain. The RIG-I/MDA5 directed interferon response is negatively regulated by laboratory of genetics and physiology 2 (LGP2) and is believed to be controlled by the mitochondria antiviral signalling protein (MAVS), a CARD containing protein associated with mitochondria.

Results: The DExD/H containing RNA helicases including RIG-I, MDA5 and LGP2 were analysed in silico in a wide spectrum of invertebrate and vertebrate genomes. The gene synteny of MDA5 and LGP2 is well conserved among vertebrates whilst conservation of the gene synteny of RIG-I is less apparent. Invertebrate homologues had a closer phylogenetic relationship with the vertebrate RIG-Is than the MDA5/LGP2 molecules, suggesting the RIG-I homologues may have emerged earlier in evolution, possibly prior to the appearance of vertebrates. Our data suggest that the RIG-I like helicases possibly originated from three distinct genes coding for the core domains including the HELICc, CARD and ATP dependent DExD/H domains through gene fusion and gene/domain duplication. Furthermore, presence of domains similar to a prokaryotic DNA restriction enzyme III domain (Res III), and a zinc finger domain of transcription factor (TF) IIS have been detected by bioinformatic analysis.

Conclusion: The RIG-I/MDA5 viral surveillance system is conserved in vertebrates. The RIG-I like helicase family appears to have evolved from a common ancestor that originated from genes encoding different core functional domains. Diversification of core functional domains might be fundamental to their functional divergence in terms of recognition of different viral PAMPs.

Figure 1

Comparative analysis of gene synteny of RIG-I (A), MDA5 (B) and LGP2 (C) in vertebrate genomes. The gene symbols are used according to the human genome map (Build 36.3, ).

Figure 2

Genomic location of predicted RIG-I/MDA5 homologues in sea anemone Nematostella vectensis. Exons and gene transcriptional orientation are indicated by blank boxes and arrows respectively. The accession numbers of predicted genes are given.

Figure 3

Phylogenetic tree analysis of DExD/H box RNA helicases. Multiple alignment of the full length protein sequences of known or predicted RIG-I, MDA5, LGP2, eIF4A and DICER was generated by CLUSTALW and used for construction of a phylogenetic tree using the neighbour-joining method within the Mega3.1 programme. The bootstrap values of the branches were obtained by testing the tree 10,000 times and values over 50% percent marked. The sequences used for tree construction are listed in Table 1. Others are as follows: monkey MDA5, NP_001040588; mouse MDA5, EDL26991; pig_MDA5, NP_001093664; mouse LGP2, NP_084426; cow LGP2, NP_001015545; dog LGP2, XP_860567; mouse RIG-I, Q6Q899; pig RIG-I, Q9GLV6; mouse DICER1, EDL18787.

Figure 4

Schematic of key functional domains of RIG-I, MDA5, LGP2, DICER and eIF4A predicted by the Pfam HMM programme.

Figure 5

Multiple alignment (A) and structural modelling (B) of Res III. The domain sequences were predicted by the Pfam HMM programme and aligned using the CLUSTALW programme. The 3-dimensional structures of the domains were generated using the 3D-jigsaw comparative modelling programme and the VAST search programme and visualised by the Cn3D programme (Version 4.1). Identical (*) and similar (:, .) residues are shown below the alignment. The conserved signature "DECH" is boxed. Amino acid position of the domain position is listed in Table 2. Predicted α-helices (green), β-strands (purple or dark blue) and loops (light blue) are marked in both the alignment and the 3D structures respectively.

Figure 6

Multiple alignment (A) and structural modelling (B) of TFIIS domains. The domain sequences were predicted by the Pfam HMM programme and aligned using the CLUSTALW programme. The 3-dimensional structures of the domains were generated using the 3D-jigsaw comparative modelling programme and the VAST search programme and visualised by the Cn3D programme (Version 4.1). Identical (*) and similar (:, .) residues are shown below the alignment. Cysteines (C1–4) involved in zinc (Zn) binding are indicated by arrows in the alignment and the 3D structures. Amino acid position of the domain position is listed in Table 2. Predicted α-helices (green), β-strands (purple or dark blue) and loops (light blue) are marked in both the alignment and the 3D structures respectively.

Figure 7

Hypothetical evolutionary origin of RIG-I, MDA5 and LGP2.