T Domain analysis and phylogenetic tree of Argonaute household of proteins.
T Domain evaluation and phylogenetic tree of Argonaute family of proteins.(A) Amino acid alignment of Argonaute protein of Bombyx mori, Tribolium castaneum, Drosophila melanogaster and Sf.Indicated sequences were aligned employing CLUSTALX.The Endoxifen E-isomer hydrochloride Formula conserved amino acid residues Asp, Asp and His (DDH) triad residues are marked with red.(B) Domain architecture of Argonaute proteins.The similarity in the domain architectures of BmAgo and SfAgo suggests that they are orthologous as well as the organisation of PAZPiwi domain is very conserved inside the insects.(C) The PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/21323637 phylogenetic tree was based on consideration of two different class of Argonaute proteins, miRNA class (Argonaute) and siRNA class (Argonaute).Sf genome consists of Ago even though Tribolium, Bombyx and Drosophila have each Ago and Ago.proteins; Aubergine and Piwi were identified from Drosophila ovaries and genome wide survey of Tribolium also identified a PiwiAub class Argonaute involved in transcriptional silencing as well as TcAgo .We could determine a single Aubergine protein homolog associated to the PiwiAubergine subfamily proteins in Sf cells (Table) (Figure B, C).As Sf cells are developed from ovaries of pupal stage Spodoptera frugiperda, it could possess an active piRNA biogenesis pathway and SfAubergine could induce Piwiassociated RNA silencing that may be reminiscent of Drosophila and Bombyx mori.Spodoptera frugiperda auxilliary RNAi components `DEAD’ box containing RNA helicase domain proteinsDExDHbox RNA helicase family members of enzymes functions as RNA chaperons because of the presence of a distinctive RNA recognition motifs over an ATP dependent component .Conserved DExHbox helicase, predominantly Dicer and Agorelated helicase genes have been shown to become required for RNAi in C.elegans and human .ATPdependent subunits p and p of DEAD box helicase distinguishes primiRNAs and induces their processing by means of Drosha complex in microRNA biogenesis pathway .Figure Domain structure and phylogeny of piRNA class elements.(A) and (B) domain search finds both Sf Argonaute and Aubergine like protein have conserved lineage of PAZ and Piwi domains.(C) A phylogenetic tree was produced utilizing the fulllength sequences of piRNA class proteins.Siwi represents silkmoth Piwi.Ghosh et al.BMC Genomics , www.biomedcentral.comPage ofWe identified three distinct classes of DEAD box RNA helicases associated to siRNA pathway in Sf genome.One of the probable ATPdependent RNA helicase resembles DpbA (DDXDBP subfamily) of Drosophila melanogaster and DDX of Homo sapiens that also includes a close homology with yet another DEAD box helicase family members protein of Bombyx mori in turn (Figure A, B).Another member in the DEAD helicase that belongs to Vasa DDX RNA binding subfamily was identified.Vasa RNA helicases are recognized to recruit Aubergine which implicates transposable element silencing in Drosophila .A third Helicase from the set of putative RNAi aspects was identified to be analogous with RNA helicase Pitchoune of Drosophila and Tribolium and DDX of Bombyx mori and Homo sapiens, which belongs to DDXHAS DEAD box helicase subfamily.Presence of a Cterminal DUF (domain of unknown function) besides theATPbinding and Helicase domains confers a typical function, which characterizes SfDDXlike protein under DDXHAS subfamily (Figure).With the 3 RNAi related helicases identified in Sf genome, knocking down the DDXHAS mRNA offered the highest reversion level of gfp, thereby.RNA processing moduleA variety of candidate RNAi genes encode proteins with known or predi.