## 6 Misincorporation and Conflicts between Transcription and Replication  
Despite different, and sometimes antagonistic activities of SCBFs, some SCBFs appear to complement each other in one important instance--in preventing conflicts between the replication and transcription machinery [66]. In rapidly growing and dividing bacterial cells replication and transcription have to proceed at the same time, and both machineries operate on the same genomic DNA [67]. As a consequence, collisions between RNAP and replication forks are highly likely to occur. Two types of collisions are possible; head to head or co-directional. Head to head collisions appear to be more detrimental to cell viability [68, 69, 70] since ribosomal operons and other highly transcribed genes are encoded on the leading DNA strand, and therefore transcribed codirectionally with replication fork progression [71]. However, codirectional collisions are still inevitable because the DNA polymerase moves at least 10 times faster than RNAP [72]. Resolution of both types of conflict _in vivo_ requires fork restart machinery in \(B\). _subtilis_[45]. Stalled/backtracked RNAPs present a particular challenge to replication fork progress; recently, backtracking was linked to genomic instability in _E. coli_ following the observation that codirectional collisions between replication forks and backtracked RNAPs resulted in double strand breaks [44]. Inability to resolve the conflicts between the two machineries often leads to chromosomal rearrangements and deletions [73, 74, 75]. Gre factors and DKsA have been linked with the prevention of collisions between replication forks and elongating RNAPs in _E. coli_[66, 76, 77]. This function appears to be redundant between GreA and DksA; deletion of DksA results in replication fork stalling in starved cells, and overexpression of GreA alleviates this effect [76]. The involvement of Gre factors in collision prevention is understandable since Gre factors can resolve backtracked elongation complexes. However, it remained unclear how the DksA factor, which does not change the catalytic properties of the RNAP active center, may reduce interference of transcription with replication. It has been hypothesised that DksA, with or without ppGpp, may prevent collisions with replication forks by displacing stalled elongation complexes, or by preventing RNAP backtracking [76]. However, neither of these activities has been observed _in vitro_[29, 63, 78]. In fact, DksA weakly inhibits transcription elongation _in vitro_[29, 78]. As mentioned above, backtracking and pausing of transcription can be caused by misincorporation of a wrong nucleotide. Interestingly, it has recently been observed that DksA inhibits incorporation of erroneous substrates into RNA both _in vitro_ and _in vivo_[29]. Further, ppGpp potentiated this function of DksA. The exact mechanism of transcriptional fidelity increase by DksA/ppGpp remains unknown. By binding in the vicinity of the active center, DksA could disfavour binding of non-cognate substrates or influence the process of TL folding, which is the major determinant of accuracy of NTP selection [79]. In addition, ppGpp, which was recently shown to bind a substantial distance from the active center, proximal to the \(\omega\) subunit [80, 81], may affect misincorporation allosterically by changing the overall configuration of the elongation complex. A second proposed binding site for ppGpp in the active center [82] may also be functional and play a role in reducing misincorporation. Therefore, although DksA/ppGpp cannot stop movement of RNAP backward _per se_, they can potentially prevent one of the causes of backtracking (Figure 2). Sequence-dependent pausing seems not to involve many backtracked pauses since deletion of either Gre factor in _E. coli_ does not have a significant effect on the pattern of pausing [83].