EXPERIMENTAL EVOLUTION OF Vibrio cholerae SHOWS INTERPLAY BETWEEN MOTILITY AND GROWTH INDEPENDENT OF CHROMOSOMAL POSITION OF RIBOSOMAL GENES.

LETICIA LAROTONDA; MARIE EVE VAL; COMERCI, DIEGO J.; MAZEL D; SOLER BISTUE, ALFONSO

Los Cocos

Congreso; Congreso Samige 2022; 2022

SAMIGE

It is unclear how gene order within the chromosome influences bacterial evolution. The genomiclocation of genes encoding the flow of genetic information is biased towards the replicationorigin (oriC) in fast-growing bacteria. Under optimal conditions, fast-growing bacteria overlapreplication rounds, a process called multi-fork replication. Hence, genes close to the oriC benefitfrom a relatively higher dose during exponential growth. In previous work we aimed at studyingthe role of chromosomal location of genes encoding the flow of genetic information on cellphysiology. We relocated the S10-spec-α locus (S10), the main ribosomal protein gene locus,and rplKAJL-rpoBC (rpoBC) locus, encoding the catalytic core of the RNA polymerase, to differentchromosomal positions in the fast-growing pathogen V. cholerae. We found that growth rate(GR) and fitness inversely correlated the distance between each locus and oriC. To gain insightinto the evolutionary effect of RP genomic position, we evolved strains bearing S10 at its currentlocation or far from oriC for 1000 generations. Although all populations increased their GR alongthe experiment the differences between the latter and the former persisted. Deep-sequencingshowed that populations fixed on average 1 mutation each 100 generations mainly at geneslinked to flagellum biosynthesis regulation, lipopolysaccharide synthesis, chemotaxis, biofilmand quorum sensing. We selected fast-growing clones displaying a ~10% GR increment findingthat they harbored inactivating mutations at, among other sites, flagellum master regulatorsflrAB. Using a novel multiple genome editing technique called Multiplex Genome Editing byNatural Transformation we introduced these single mutations into naïve V. cholerae strains. Weobserved a ~10% increase in growth rate demonstrating the role of these mutations in GRincrease. Therefore, along the evolution V. cholerae increases its GR independently of theposition of genes encoding ribosomal genes. The lack of suppressor mutations compensatingS10 genomic location demonstrates that gene order influences the evolution of bacteriallineages in the long run.