## 4 Discussion
### The Respiratory Chain
The EcoCyc database reports 106 transcriptional units controlled by IHF including several genes analyzed in this study like _ompC_ (controlled negatively), the TCA cycle and the glyoxylate shunt (_sucABCD_, controlled negatively; _aceBAK_ and _glcB_ controlled positively), fermentative and acetate utilization (_pflB_ and _acs_, controlled negatively), respiratory genes (_nuoABCFMN_, _ndh, marG_, all controlled negatively), and Fnr (controlled positively) [4]. The DNA-binding transcriptional repressor IcIR (coded by _icIR_) is involved in adaptive responses in bacteria. In _E. coli_, this repressor is reported to control the expression of genes involved in the glyoxylate shunt positively [41], such as _aceBAK_ (included in this study). FadR, the DNA-binding transcriptional dual regulator encoded by _fadR_, is the fatty acid degradation regulator that negatively controls fatty acid and lipid metabolisms [42] and acetate metabolism [43]. Acetate transport, oxidation, and incorporation into macromolecules were observed to increase (5X) in a _fadR\({}^{-}\)_ mutant during growth on succinate as a carbon source, associated with an increased level of glyoxylate shunt in this mutant [43; 44]. As FrdR activates the expression of _icIR_, it has been proposed as an indirect upregulation of _aceBAK_ genes involved in the glyoxylate shunt [11; 44]. Finally, the DNA-binding transcriptional dual regulator Fnr (encoded by _fnr_) is a primary transcription regulator controlling the transition from aerobic to anaerobic growth through the control of more than 140 genes [4], including acid resistance, chemotaxis, cell structure, and several biosynthetic processes [45]. There are 142 transcriptional units controlled with Fnr, including the negatively controlled TCA cycle genes _aceEFB_, _sdhCDAC_, _sucABCD_, and _lpd_ genes, and the respiratory genes _cyoBCDE_ and _marG_. Additionally, Fnr positively controls the expression of ArcA [4]. As the cellular concentration of Fnr is the same under both aerobic and aerobic growth conditions, its activity is regulated by oxygen concentration [46]. In addition to the direct role as a transcriptional factor, Fnr can act indirectly through the increased expression of ArcA and the target genes [45]. Fnr acts as a dimmer; in the presence of oxygen, the dimeric structure is converted to a monomeric form, reducing its ability to bind target DNA. Additionally, depending on the growth conditions, the proposed number of targets for Fnr and other global regulators varies (e.g., growing on a minimal M9 medium supplemented with glucose vs. growing in LB medium) [45]. The expression of genes encoding the sigma factors _rpoD_, _rpoE_, _rpoH_, _rpoN_, and _rpoS_ showed an increased expression in both PB12 mutants compared to the wild-type strain. The genes _rpoA_, _rpoC_, and _rpoZ_ encoding the RNA polymerase subunits \(\alpha\), \(\beta^{\prime}\), and \(\omega\), respectively, were found unchanged in both strains (_rpoA_), unregulated only in PB12 (_rpoC_) and upregulated in both strains (_rpoZ_) (Table 3). The increased transcription of _rpoS_ and _rpoD_ has been proposed to indicate the exposition to nutritional stress in wild-type _E. coli_[47; 48]. In strains lacking the _crr_ gene, like the PB12 mutants, the absence of the EII\({}^{\text{Glc}}\) enzyme led to higher levels of RpoS even during exponential growth, allowing the transcription of several central carbon metabolism genes. This condition led to the proposal of \(\alpha^{38}\) as a second vegetative sigma factor [4]. _rpoD_ was the unique sigma factor more highly expressed in the AGD\({}^{+}\) than in PB12, showing an increment of 75% in the relative transcription level.