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A Rapidly Evolving Polybasic Motif Modulates Bacterial Detection by Guanylate Binding Proteins.

ABSTRACT: Cell-autonomous immunity relies on the rapid detection of invasive pathogens by host proteins. Guanylate binding proteins (GBPs) have emerged as key mediators of vertebrate immune defense through their ability to recognize a diverse array of intracellular pathogens and pathogen-containing cellular compartments. Human and mouse GBPs have been shown to target distinct groups of microbes, although the molecular determinants of pathogen specificity remain unclear. We show that rapid diversification of a C-terminal polybasic motif (PBM) in primate GBPs controls recognition of the model cytosolic bacterial pathogen Shigella flexneri By swapping this membrane-binding motif between primate GBP orthologs, we found that the ability to target S. flexneri has been enhanced and lost in specific lineages of New World primates. Single substitutions in rapidly evolving sites of the GBP1 PBM are sufficient to abolish or restore bacterial detection abilities, illustrating a role for epistasis in the evolution of pathogen recognition. We further demonstrate that the squirrel monkey GBP2 C-terminal domain recently gained the ability to target S. flexneri through a stepwise process of convergent evolution. These findings reveal a mechanism by which accelerated evolution of a PBM shifts GBP target specificity and aid in resolving the molecular basis of GBP function in cell-autonomous immune defense.IMPORTANCE Many infectious diseases are caused by microbes that enter and survive within host cells. Guanylate binding proteins (GBPs) are a group of immune proteins which recognize and inhibit a variety of intracellular pathogenic microbes. We discovered that a short sequence within GBPs required for the detection of bacteria, the polybasic motif (PBM), has been rapidly evolving between primate species. By swapping PBMs between primate GBP1 genes, we were able to show that specific sequences can both reduce and improve the ability of GBP1 to target intracellular bacteria. We also show that the ability to envelop bacteria has independently evolved in GBP2 of South American monkeys. Taking the results together, this report illustrates how primate GBPs have adapted to defend against infectious pathogens.

SUBMITTER: Kohler KM

PROVIDER: S-EPMC7240152 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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A Rapidly Evolving Polybasic Motif Modulates Bacterial Detection by Guanylate Binding Proteins.

Kohler Kristin M KM Kutsch Miriam M Piro Anthony S AS Wallace Graham D GD Coers Jörn J Barber Matthew F MF

mBio 20200519 3

Cell-autonomous immunity relies on the rapid detection of invasive pathogens by host proteins. Guanylate binding proteins (GBPs) have emerged as key mediators of vertebrate immune defense through their ability to recognize a diverse array of intracellular pathogens and pathogen-containing cellular compartments. Human and mouse GBPs have been shown to target distinct groups of microbes, although the molecular determinants of pathogen specificity remain unclear. We show that rapid diversification ...[more]

PMID: 32430466

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Project description:Here we focus on identifying conserved elements that are required for Chaserr function, a lncRNA that we recently characterized as being essential for mouse viability (https://doi.org/10.1038/s41467-019-13075-8). Chaserr is highly conserved in different species, and is located in close proximity (<2kb) to the transcription start site of CHD2. It acts in concert with the CHD2 protein to maintain proper Chd2 expression levels. Using the LncLOOM framework we identified AUGG enriched conserved elements which are located in the last exon of mouse Chaserr. In order to identify proteins that potentially bind to these conserved regions, we used in vitro transcription to generate the following biotinylated RNAs; along with their antisense controls: 1) The WT sequence, containing the first 322 nt of the last exon of Chaserr (bases 764–1086 of the NR_037601 isoform). An antisense control fragment was also prepared, labelled as AWT 2) The mutant conserved (MC) sequence, that contained AUGG→UACC mutations in four conserved motifs that were found using LncLOOM. An antisense control fragment was also prepared, labelled as AMC 3) The muatant all (MA) sequence in which all seven of the AUGG sites in the last exon were mutated to UACC. An antisense control fragment was also prepared, labelled as AMA 4) We also included a No Probe Control, which did not contain an RNA fragment. The fragments were incubated with N2a cell lysates followed by RNA-pulldown and Mass Spec analysis. The experiment was performed in triplicate. A total of 938 proteins were found to associate with the WT sequence: 74 of these were enriched ≥3-fold compared to the antisense sequence.We identified 9 proteins that had ≥2-fold higher recovery when using the WT sequence compared to both mutants. Included in these proteins was DHX36, which showed the highest enrichment compared to the mutated sequences. We validated the association of DHX36 with Chaserr through RNA immunoprecipitation experiments.

Dataset Information

A Rapidly Evolving Polybasic Motif Modulates Bacterial Detection by Guanylate Binding Proteins.

Publications

A Rapidly Evolving Polybasic Motif Modulates Bacterial Detection by Guanylate Binding Proteins.

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