It is generally accepted that bacterial chromosomes have a bias of Gn towards their leading strands, leading to
the so-called G+C skew, but for some genomes, including many Firmicutes, there is also a weaker A+T strand bias, with the leading strand containing more Tn than An (Rocha et al., 2000). G+C content has been related to
numerous properties, such as genome size, oxygen and nitrogen exposure, and specific habitats. On that note, intracellular bacteria on average have smaller genomes and are more A+T-rich (Wassenaar et al., 2009). The higher A+T content in small, intracellular bacteria has been attributed to a loss of repair genes, leading to an increase in the mutation rates from cytosine to thymine (Moran, 2002; Rocha and Danchin, 2002).
Bacterial chromosome replication starts from a single origin, the origin of replication (ori). Most bacteria show G+C and A+T skews around the origin of replication. However, two mycoplasma species were shown to be C+T-rich in the third codon position (McLean et al., 1998). Uneven distribution of nucleotides on the leading and lagging strands cannot always be reduced to a nucleotide skew. Circular chromosomes tend to have the ori and terminus of replication on exact opposite sides, which evens out the G+C skew to the extent that the bias is filtered out to near zero, resulting in the number of Gn being equal on both strands (Bohlin et al., 2010).
Replication mechanisms may influence the strand’s symmetry patterns, and as such, a lot can be learned by the examination thereof. However, horizontally transferred genes may also influence the strand symmetry, leading to changes in the polarity of skew plots (Xia, 2012).
The direction of the nucleotide skew switches at the origin of replication, so that the leading strand contains more Gn than Cn. However, in M. genitalium and M. pneumoniae, the direction of the skew was found to be
opposite to the direction for codon position 3 (McLean et al., 1998). Their genes are also arranged in such a way resulting in the transcriptional and replicational directions to be the same, a tendency that is more pronounced for highly expressed genes such as ribosomal protein genes (Brewer and Fangman, 1988). According to Karlin et al. (2003) most bacterial genomes contain a large cluster close to the ori that contains between 20% and 40% of all ribosomal protein genes. Many bacterial genomes’ rRNA operons are in the same direction as replication (Price et al., 2005; Rocha, 2002). Other genes that are involved in protein synthesis are encoded within or proximal to this large cluster, such as the tuf, fus, rpoA, rpoB and rpoC genes.
All of the replication origins that have been experimentally confirmed occur in intergenic regions (Gao and Zhang, 2008; Mackiewicz et al., 2004). The position of the ori is conserved among bacterial species, mostly found in close proximity to the dnaA gene. The dnaA gene encodes for the universally conserved DNA replication initiation protein DnaA (Ogasawara et al., 1991). The gene cluster rnpA-rpmH-dnaA-dnaN-recF-gyrB- gyrA that surrounds the ori and the dnaA gene is often conserved, with the ori located adjacent to the dnaA gene in an intergenic region (Briggs et al., 2012). The ori of M. genitalium and M. pneumoniae reside in an A+T-rich area between the dnaA and dnaN genes. The low G+C content Firmicutes generally have two unique essential genes, the dnaD and dnaB genes, in addition to the dnaA gene (Leonard and Grimwade, 2011; Soultanas, 2011). Several Mollicutes have only a single gene, the dnaD-like gene, combining both of the functions of the dnaD and dnaB genes (Briggs et al., 2012). Genomes with more An than Tn were shown to contain both a polC
and dnaE homologue, while the other genomes lack the polC homologue (Worning et al., 2006). The direction of the A+T skew is determined by the DNA polymerase-α subunit that replicates the leading strand (Worning et al., 2006).
3.1.6. DEFINITION OF ESSENTIAL GENES AND THE MINIMAL GENOME
Essential genes (EG) form one of the three groups of functional genes, with highly expressed genes (HEG) and horizontally transferred genes (HGT) being the other two, that play an important role in the survival and infectious ability of pathogens (Gao and Chen, 2010). EG represent the minimal gene set that is required for sustaining life of the simplest, self-reproducing bacteria. For some researchers they represent the gene set absolutely required for cell viability, under any conditions. Others use EG to refer to the gene set that contributes to the fitness of the organism and its competitive growth under favourable conditions, for instance in the absence of environmental stress and in the presence of essential nutrients (Mushegian and Koonin, 1996a). Most of the metabolic genes’ essentiality is relative to the metabolic context or specific conditions (Gerdes et al., 2003; Koonin, 2003). These genes therefore represent the part of the bacterial genome that one would suspect to be relatively constant, since a change in the expression profile or change in the phase variability would cause functional differences and lead to a loss of essentiality. Some of the known functional genes are drug targets, therefore further identification and investigation into important EG might provide further drug targets, lead to development of broad-spectrum antibiotics, and aid in developing species-specific antibiotics (Gao and Chen, 2010). It will further enable quicker annotation of unknown genome sequences through comparative studies and homology searches (Lin and Zhang, 2011).
One would expect EG to be conserved, as is the case in bacteria (Gerdes et al., 2003; Jordan et al., 2002). EG are more conserved than non-EG, however, conserved genes are not necessarily essential (Fang et al., 2005). In addition to this, the fact that certain genes are conserved among bacteria, does not necessarily imply that the genes’ essentiality is also conserved, i.e. extrapolation of essentiality is not possible among bacteria (Zalacain et al., 2003).