Dead Affects MRNA Levels In Escherichia Coli

DeaD(previously named CsdA)is a member of the DEAD-box family of RNA helicases in Escherichia coli.It is a cold shock-inducing protein.DeaD assembles with ribonuclease E into RNA degradosome involved in mRNA turnover under low temperature conditions,and is also involved in ribosome assembly.Our laboratory found that DeaD affected the cell cycle processes,but the mechanism behind is not known.The study firstly measured the amount of total protein per cell and the amount of DnaA,the replication initiation protein,per cell by Western blotting and RT-PCR.DnaA increased in a single ?deaD cell,indicating that the early initiation of DNA replication may be related to the increased amount of DnaA in ?deaD cells.The bacterial two-hybrid experiments showed that there was no interaction between DnaA and DeaD,indicating that DeaD doesn't affect DNA replication initiation through interacting with DnaA directly.In order to further explore the mechanism of DeaD-mediated effect on the initiation of replication,the total cellular RNA per cell was measured.The result show that the total amount of RNA per cell increases in?dead cell.In vitro experiment showed that DeaD degraded RNAs in the presence of ATP at 25?.These results suggest that DeaD may affect the level of transcription.The RNA-seq technique was used to determine the changes in gene expression of?deaD and wild-type cells before and after rifampicin treatment.The results suggest that DeaD may degrade the mRNA of genes coding subunits of DNA polymerase,RNA polymerase,stress proteins,proteins involved in glucose and amino acids metabolism.It is likely that the earlier initiation of DNA replication in ?dea D cells could be due to the up-regulation of the genes described above.In addition,the expressions of flagella,respiratory related genes and ABC transporter genes were down-regulated in ?dead cells,suggesting that DeaD may protect mRNAs of the genes mentioned above at low temperature.In summary,the DeaD protein is likely to degrade or protect mRNA as a RNase and also affect gene expression as a transcription factor.