## 4 Discussion
### M-Mutations
RcpC, encoded in the prototypical _tad_ locus of _Aggregatibacter actinomycetemontans_, and CpaB in _Caulobacter crescentus_ and all its known homologues contain two \(\beta\)-clip motifs involved in a variety of functions, including binding carbohydrate moieties to assemble structures such as Flp pili [46; 57]. It has also been proposed that one or both \(\beta\)-clip motifs glycoslate pilin subunits to facilitate their assembly. As indicated by [PERSON] et al. [56], a rcpC/cpaB gene is absent in the tad loci of Gram-positive bacteria, which also holds true for _M. luteus_[15]. This does not indicate that such genes cannot still be present at a different genomic locus and belong to a different transcriptional unit. The same applies for _tadZ/cpaE_, although this gene is indeed present in _tad_ clusters of other actinobacteria [56]. This cytosolic soluble protein is an essential component of the Tad secretion system in many organisms, and its particular molecular architecture combines components from the bacterial cytoskeleton (MinD/ParA/Soj ATPases) and two-component signal transduction response regulators. Studies in _A. actinomycetemontans_ and _C. crescentus_ showed that they probably function as localization factors [58; 59]. In _C. crescentus_, CpaE mediates positioning of the pilus secretion protein CpaC, as well as the histidine kinase PleC. Proteins containing PAS-domains such as the one encoded by Mlut_14670 are usually modular, functioning as sensors that detect a variety of stimuli and regulate, in response, the activity of effector (output) domains with functions such as catalysis or DNA binding [60]. In certain organisms, such as _P. aeruginosa_[61], _A. actinomycetemontans_, _A. aphrophilus_[57] and _C. crescentus_[62; 63], the environment-dependent modulation of pilin production through two-component regulatory systems of this type seems to be a common feature. Considering how close together Mlut_14660 and Mlut_14650 are, it is difficult to assess whether mutations in the 5\({}^{\prime}\) regions of these genes, which also may encompass their regulatory sequences, affect one gene or the other, or both, and in which way. To obtain more information on this matter, the starting point of each transcript unit was determined. It seems that there is a divergent overlap of the transcriptional units and 5\({}^{\prime}\)UTRs (97 bp apart) of Mlut_14660 and the Mlut_14650/40/30 operon structure, including the promoter or operator sequences of each unit and maybe extending into part of the coding regions. This type of arrangement [64] benefit the hypothesis of transcriptional interference (TI) as a way of gene regulation of Mlut_14660 and the operon Mlut_14650/40/30. TI was defined as the suppressive influence of one transcriptional process, directly and in cis on a second transcriptional process by [PERSON] [65]. A model such as this one was also proposed for the divergently overlapping _comEA/EC_ and _comER_ genes in _B. subtilis_[16]. In a more recent work [66], it has been shown that _comER_ plays an important role in biofilm formation and sporulation, and that competence has a negative correlation with biofilm formation [67], supporting the stated hypothesis. When analyzing if the gene Mlut_14660 and the operon Mlut_14650/40/30 and their specific genetic context is conserved in other bacteria, we found that they are present and have the same or similar surroundings on the chromosomes of several different Actinobacteria genera (see Figure 5; only selection of chromosomal regions of related genera and species with similar gene arrangements depicted). In the proposed model (Figure 7), the expression of one of the transcripts may generate a negative superhelical turn in the opposite sense, impeding the expression of the other transcript. The steric hindrance derived from the RNA polymerase bound to the promoter can also play a role, or a mix of both phenomena. Data published recently showed that the Mlut_14650/40/30 operon is among the 25 most highly expressed _M.