RNA Thermosenser Controls Fraction 1 Antigen Expression In Yersinia Pestis

Yersinia pestis,the causative agent of plague,is usually transmitted by flea bites.There remain many natural epidemic foci of plague.Y.pestis is regarded as a bioterrorism agent and biological warfare agent.In addition,emergence of multi-drug resistant Y.pestis strains greatly threatens public health and national biosafety.Y.pestis contains three virulent plasmids:pCD1,and the newly obtained pMT1 and pPCP1 during evolution of Y.pestis from its ancestor Yersinia pseudotubeculosis.pMT1 encodes the murine toxin Ymt as well as fraction 1(F1)antigen.F1 antigen,a critical immunoprotective antigen,accumulates on cell surface to form an amorphous capsule which is unique to.pestis.F1 antigen encoding gene cluster was acquired in the early step of evolution of Y.pestis.F1 antigen is important for.pestis to inhibit phagocytosis by macrophage,and might promote transmission via flea vector.The expression of F1 antigen is tightly controlled,and temperature serves as a critical signal.FI antigen is not expressed in the temperature of flea host(less than 30?)but highly expressed in the temperature of mammalian host(37?).Bacterial RNA thermosenser is a temperature-sensing RNA seeondary structure folded by 5' end of mRNA,preventing access of the ribosome to the Shine-Dalgamo(SD)sequence at lower temperatures,as temperature increases the structure unfolds and facilitates mRNA translation at elevated temperatures,and regulates gene expression at the translational level.In this study5 we found that FI antigen expression is regulated by temperature at the transcriptional level via the transcription activator CaflR.Further study showed Caf1R auto-regulates its transcription and its expression is eontrolled by an RNA thermosenser.We found that caf1R mRNA 5' UTR and the 5' end of the coding sequence forms a secondary structure of an RNA thermosenser to sense temperature and regulate caf1R expression.When the regulatory sequence was fused to egfp,eGFP was highly expressed at 37?,but not at 26?.Mutations of the base-pairs in the region of the SD sequence and start codon predicted to stabilize the structure inhibited eGFP expression at 37?.However,when the mutations destabilized the structure,eGFP expression improved at 26? and was comparable to that at 37?,destroying the temperature-dependent induction effect.Complementary mutations recovered the induction effect.In vitro translation showed that the regulatory sequence renders temperature-dependent translation efficiency of eGFP.Further chemical probing demonstrated that temperature and site mutations influence RNA folding of the region of the SD sequence and start codon significantly.Our results show caflR RNA thermosenser represents a new regulatory element to eontrol gene expression at the translational level.These results not only enrich our knowledge about RNA thermosenser but also facilitate the functional study on FI antigen and promote the Y.pestis vaccine development based on F1 antigen.