# The _E. coli_ Global Regulator DksA Reduces Transcription during T4 Infection
## 3 Results
### Deletion of dksA Results in a Shorter Latent Period in a T4 motA\({}^{am}\) Infection
This phenomenon arises in part because the effect of the _motA_ knockdown on Pm activation is promoter specific [40; 42; 55]. In addition, most middle genes are transcribed by both Pm's and upstream Pe's (Figure 3). Therefore, the effect of T4_motA\({}^{am}\)_ on a particular middle gene also depends on how much transcription from a Pe extends into this gene. While the mechanism for this elongation has not been fully elucidated, it appears to be dependent on processes that stabilize RNA rather than by anti-termination [56; 57]. Even though the 3\({}^{\prime}\) ends of many of these transcripts have been identified (indicated as grey bars in Figure 3), these ends are thought to represent the ends of the stable RNA rather than transcription termination sites. Differential expressions determined by RNA-seq analysis were compared to RT-qPCR assays of 68 genes using RNA isolated 5 min after infection and by primer extension analyses using RNA isolated at 1 and 5 min after infection. One min after infection, early promoters are active, but middle transcription has just initiated [53; 54]. A comprehensive summary of our RNA-seq and RT-qPCR analyses is shown in Figure 3. Overall, we observed a strong correlation between the results of the RNA-seq and RT-qPCR analyses (Figure 55a). Not surprisingly, our RNA-seq/RT-qPCR analyses were consistent with the results from previous studies of T4_motA\({}^{am}\)_ infections. Expression of 44% of early genes increased \(\geq\)2-fold, while expression of 79% of late genes decreased \(\geq\)2-fold (Figure 3). (Genes are classified according to the expression patterns observed in a previous global microarray analysis of a T4 wt infection [58].) Expression of middle genes was mixed, and middle genes that show significant MotA dependence varied in the magnitude of their dependence. To confirm that changes in gene expression arose from defects in Pm transcription, primer extensions were performed for several middle genes (Figure 56a). As expected, this analysis confirmed previous results indicating that the _motA\({}^{am}\)_ mutation results in a significant decrease in Pm activation. In addition, we also observed lower transcription in the T4_motA\({}^{am}\)_ infection that maps to one early promoter, Pe148.6 (Figure 56b). This result is surprising because the classification of this promoter as a Pe is confirmed by its activity 1 min after infection (Figure 4b, lanes 5-8) and because there is no recognizable Pm sequence at this site. However, reduced transcription from Pe148.6 has also been observed in a T4_motA\({}^{am}\)_ infection of another cell line [40]. Figure 3: _Cont._  
Figure 3: Cont. Figure 3: Genomic map of bacteriophage T4 correlated with gene expression data from RNA-seq. Graphical representation of the T4 genome showing the location of annotated genes and promoters as given in [53]. The fold changes from the RNA-seq analyses determined for infections of T4 wt/B606 \(d\)s/a (wt/\(d\)s/a), T4 mt0_A_A_A_A_B  
Overall, these analyses provided a \"proof of principle\" that our RNA-seq analyses correctly reflected the state of T4 transcription 5 min post-infection. However, this study also significantly extended previous work because one can now investigate the effect of the \(motA\) knockdown on each gene throughout the T4 genome. Interestingly, our results indicate that there did not appear to be any general correlation between the function of a middle gene and its dependence on MotA.