Structural Alterations in a Component of Cytochrome c Oxidase and Molecular Evolution of Pathogenic Neisseria in Humans

Three closely related bacterial species within the genus Neisseria are of importance to human disease and health. Neisseria meningitidis is a major cause of meningitis, while Neisseria gonorrhoeae is the agent of the sexually transmitted disease gonorrhea and Neisseria lactamica is a common, harmless commensal of children. Comparative genomics have yet to yield clear insights into which factors dictate the unique host-parasite relationships exhibited by each since, as a group, they display remarkable conservation at the levels of nucleotide sequence, gene content and synteny. Here, we discovered two rare alterations in the gene encoding the CcoP protein component of cytochrome cbb3 oxidase that are phylogenetically informative. One is a single nucleotide polymorphism resulting in CcoP truncation that acts as a molecular signature for the species N. meningitidis. We go on to show that the ancestral ccoP gene arose by a unique gene duplication and fusion event and is specifically and completely distributed within species of the genus Neisseria. Surprisingly, we found that strains engineered to express either of the two CcoP forms conditionally differed in their capacity to support nitrite-dependent, microaerobic growth mediated by NirK, a nitrite reductase. Thus, we propose that changes in CcoP domain architecture and ensuing alterations in function are key traits in successive, adaptive radiations within these metapopulations. These findings provide a dramatic example of how rare changes in core metabolic proteins can be connected to significant macroevolutionary shifts. They also show how evolutionary change at the molecular level can be linked to metabolic innovation and its reversal as well as demonstrating how genotype can be used to infer alterations of the fitness landscape within a single host

Cytochrome c4 is required for siderophore expression by Legionella pneumophila, whereas cytochromes c1 and c5 promote intracellular infection

A panel of cytochrome c maturation (ccm) mutants of Legionella pneumophila displayed a loss of siderophore (legiobactin) expression, as measured by both the chrome azurol S assay and a Legionella-specific bioassay. These data, coupled with the finding that ccm transcripts are expressed by wild-type bacteria grown in deferrated medium, indicate that the Ccm system promotes siderophore expression by L. pneumophila. To determine the basis of this newfound role for Ccm, we constructed and tested a set of mutants specifically lacking individual c-type cytochromes. Whereas ubiquinol-cytochrome c reductase (petC) mutants specifically lacking cytochrome c1 and cycB mutants lacking cytochrome c5 had normal siderophore expression, cyc4 mutants defective for cytochrome c4 completely lacked legiobactin. These data, along with the expression pattern of cyc4 mRNA, indicate that cytochrome c4 in particular promotes siderophore expression. In intracellular infection assays, petC mutants and cycB mutants, but not cyc4 mutants, had a reduced ability to infect both amoebae and macrophage hosts. Like ccm mutants, the cycB mutants were completely unable to grow in amoebae, highlighting a major role for cytochrome c5 in intracellular infection. To our knowledge, these data represent both the first direct documentation of the importance of a c-type cytochrome in expression of a biologically active siderophore and the first insight into the relative importance of c-type cytochromes in intracellular infection events