# The _E. coli_ Global Regulator DksA Reduces Transcription during T4 Infection
## 3 Results
### Reduction of Early Promoter Activity during T4 wt Infection in the Presence of DksA
For example, in regions whose pre-replicative RNA levels were severely depressed in the T4_motA\({}^{am}\)_/wt infection (the region containing genes 43, 44, 62, and 45 (map units 32,662 to 27,197) and the region containing genes 33, _rnh_, 59, and 32 (map units 150,727 to 146,948)), the level of RNA from the T4_motA\({}^{am}\)_ infection in the absence of DksA increases from 1.5 to 7-fold over that seen in the T4_motA\({}^{am}\)_ infection in the presence of DksA. These are the middle genes that are most severely affected by the _motA_ knock-down. Because the absence of DksA also increases the level of _motA_ transcription in the _motA\({}^{am}\)_ background by \(\sim\)2-fold (Figure 3) and the _motA\({}^{am}\)_ mutation is leaky, some of this increase could arise from an increase in MotA protein, which could then lead to more Pm activation. However, it might also arise from the increase in transcription from Pe's, which then extends into downstream middle genes. To observe early and middle transcription separately, we used primer extension analyses. We found that for the region containing map units 35,014 to 30,342 increased transcription from Pe35.3 in the absence of DksA was particularly helpful in counter-acting the effect of the _motA_ knock-down since we observed no significant increase in middle transcription from Pm46 or Pm45 (Figures S6 and S8). Likewise, in the _rnh-32_ region (map units 150,727 to 146,948), we observed that the overall increase in transcription resulted from a significant increase in transcription from Pe148.6 rather than an increase in Pm_dksA_ (Figure 4 and Figure S6b). In this case, a low level of Pe148.6 RNA was observed in the T4_motA\({}^{am}\)_ infection of B606 wt at 5 min (Figure 4). However, the level of Pe148.6 RNA returned to that seen in the T4 wt/B606 wt infection when the _motA_ knockdown was combined with the absence of DksA (Figure 4 and Figure S6b). Unlike the middle genes detailed above, the transcript level of gene 55 (late sigma factor; map units 40,157 to 39,600) was not significantly reduced by the _motA_ knockdown, yet gene 55 expression increased even more in a T4_motA\({}^{am}\)_ infection in the absence of DksA (Figure 3). Primer extensions indicate that increased transcription of 55 is coming from Pe40.4 rather than from Pm55 (Figures S6, S8c and S8d). In addition, primer extensions revealed that Pe40.4 RNA is processed at map unit 40,902. This position contains the consensus sequence recognized by RegB, a T4-encoded sequence-specific endonuclease that inactivates T4 early transcripts shortly after infection [(59; 60)]. The transcript levels near (Figure S6b) and downstream (Figure S8d) from Pe40.4 were similar, indicative of regulation at or near the promoter either directly or indirectly by DksA. No novel primer extension products were observed in any of the tested regions in the absence of DksA (Figure 4 and Figure S8), indicating that the deletion of DksA does not generate new transcription starts. As genes in regions transcribed by Pe35.3 (map units 35,662 to 27,197), Pe40.4 (map units 41,225 to 39,600), and Pe148.6 (map units 150,727 to 146,948) are required for replication and late transcription, it is not surprising that an increase in transcription from these promoters in the absence of DksA would improve the growth of the T4_motA\({}^{am}\)_ phage in B606 \(\Delta\)dksA_ relative to B606 wt.