## 3 Post-Transcriptional and Translational Regulation in Integrons
### The Role of attC Site on Integro-Associated Gene Cassette Translation  
The cassette position effect clearly interferes with the gene cassette translation efficiencies, and several pieces of evidence pointed to the influence of _attC_ sites in this phenomenon and in the differential manifestation of antibiotic resistance. For example, previous studies demonstrated that streptomycin resistance decreased differentially depending on which gene cassette (_aacC1_ or _aacA4_) preceded _aadA2_[2]. In addition, _aacA4_ was weakly expressed when preceded by _bla_IMP cassettes [35,54]. [PERSON] et al. [59] have extensively studied the interference of _attC_ sites on downstream cassette translation. They showed that, contrary to what was previously proposed [2], the second structures formed in mRNA due to intra-pairing of the _attC_ palindromic region have no influence on transcription termination of downstream gene cassettes. Actually, these stem-loop structures prevent the ribosome movement throughout the mRNA, directly interfering, at the translational level, in the expression of more distal genes regarding Pc [4; 59]. It was demonstrated that hairpin destabilization of _attC_ showed to enhance the translation of 3\({}^{\prime}\) gene cassettes. Considering that the _attC_ sites differ from one another in both sequence and length [6], distinct cassettes (with distinct _attC_ sites) would modulate the expression of downstream genes to different extents. In fact, [PERSON] et al. [59] observed that gene expression was differentially affected in response to the length and composition of upstream _attC_ sites, corroborating previous findings in which the bottom strand of some lengthy _attC_ sites presented more complex secondary structures [66]. For example, it had been verified that the levels of _aac(6\({}^{\prime}\))-lb\({}_{7}\)_ translation, the second cassette in the array _bla_IMP-8_-aac(6\({}^{\prime}\))-lb\({}_{7}\)-catB4_, varied significantly in response to alterations in stem-loop structures and/or shortening of IMP-8 _attC_ site [59]. However, these stem-loop barriers may be relieved by the occurrence of a short ORF within the _attC_ site, which would increase the processivity of the translational complex, favoring expression of downstream cassettes via a mechanism of translational termination-reinitiation similar to that described for ORF-11. In termination-reinitiation, the ribosome is not released from the preceding gene cassette stop codon and starts translation at the initiation codon of the short ORF encoded in its _attC_ site. This event prevents the stem-loop formation and allows ribosomal scanning along with the transcript, reinitiating translation of the following cassette [59; 60; 126]. As mentioned above, the expression of fused cassettes occurs by a translation coupling mechanism. Since these fusions are usually formed as a result of total or partial loss of the _attC_ site, the lack of stem-loop structures would no longer constitute a physical barrier to ribosome progression, increasing expression of downstream gene cassettes [66]. [PERSON] et al. [98] identified the ORF-11 and its TIR at the _att11_ region of a class 1 inte-gon harboring the _gcu14_ followed by the fused cassette _bla_GES-1_/aacA4_. They verified that transforms harboring this gene cassette array were resistant to \(\beta\)-lactams and aminoglycos, suggesting that both _bla_GES-1 and _aacA4_ were expressed. The _gcu14_ was a TIR-deficient gene cassette, and they proposed that it was expressed by translation coupling with ORF-11. An SD sequence was found 10 bp upstream of the _bla_GES-1_/aacA4_ and the lack of stem-loop structures, due to loss of GES-1 _attC_ region that led the fusion (Figure 7), probably enhanced the chances of _aacA4_ translation.