## 2 Transcriptional Regulation in Integrons
### Interplay between Integrase and Gene Cassette Expression in Class 1 Integrons
#### 2.6.1 Integrase Activity Is Linked and Inversely Correlated to Gene Cassette Expression
However, the occurrence of multiple gene cassette arrangements was observed only when the most active integrase variant IntI1R32_H39 was present [26]. In this case, IntI1 preferentially excised the cassettes immediately preceding _catB9_ rather than excising _catB9_ and reinserting it closer to Pc, since it is less cost-effective considering that only one recombination event takes place. Additionally, although the dynamics of cassette shuffling were higher when IntI1 was found derepressed or overexpressed, such recombination events under selective pressure were observed even when IntI1 was weakly expressed in response to LexA repression [116]. Interestingly, in all rearrangements observed in that study, the _catB9_ was brought to the second cassette array position, downstream of _aac(6')-Ib_. This could be due to the lower efficiency of the integrase in catalyzing excision of _aac(6')-Ib_ in the first position, which involves _attI X attC_ recombination [23], or to the length and structure of the folded bottom strand of the _aac(6')-Ib_ and _catB9 attC_ sites, which could be poorly recognized by IntI1 [117]. A recent study established another interesting interplay between cassettes expression and integrase activity. As extensively discussed above in the previous topic, the _intI1_ expression is tightly regulated in class 1 integrins by LexA repressor [17]. The LexA protein is absent in _Acinetobacter baumannii_, suggesting that _intI1_ expression in this species is probably constitutive. However, a recent study demonstrated that class 1 integrins identified in _Acinetobacter_ predominantly harbor the PCS configuration, the strongest cassette promoter variant [43]. As already mentioned, the PCS configuration determines the least efficient integrase variant (IntI1R32_N39) [26] and, consequently, the lowest fitness cost for bacteria. Therefore, this finding suggested that the PCS prevalence in _A. humantii_ could be associated with the reduced fitness burden to evade the deleterious activity of integrase. Additionally, this study also verified that despite the strong nature of the PCS promoter, its transcription efficiency was weaker in _A. humantii_ than in _E. coli_. Although _A. baumannii_ and _E. coli_ RNA polymerases and \(\sigma^{70}\) factors share considerable conservation, such inter-species differences in Pc activity could result from differences in their transcriptional machinery [118]. Therefore, this regulatory mechanism represents an adaptive response designed to reduce the fitness cost and contribute to integrand stabilization in this species [43]. 6.2 Transcription Interference between P\({}_{\text{intI1}}\) and Cassette Promoters in Class 1 Integons  
It is already known that, in the case of class 1 integrins, the Pc promoter is found embedded in the _intI1_ gene and that Pc and P\({}_{\text{intI1}}\) drive transcription in opposite orientations [2; 3], with the corresponding transcription initiation sites separated by 100 bp. Thus, when both P\({}_{\text{intI1}}\) and Pc promoters are active, convergent transcription takes place, resulting in transcriptional interference due to sense-antisense mRNA interactions (Figure 8). In fact, it was verified that P\({}_{\text{intI1}}\) is more efficient in driving _intI1_ transcription in the absence of an active Pc promoter. A previous study demonstrated that P\({}_{\text{intI1}}\) is susceptible to transcription interference, which preferentially occurs in the presence of the strong PCS. Due to the high transcription levels that PcS evokes, this is the only Pc variant able to influence negatively _intI1_ expression even in the absence of LexA repression [101]. These findings indicated that the _intI1_ is less transcribed in the presence of strong Pc promoter versions, which is expected in transcription interference events, where the activity of a promoter is negatively influenced by that of a stronger one [119].