Molecular mechanisms for the evolution of gene structure and organization: the histidine case

Abstract

The origin and evolution of metabolic pathways represented a crucial event occurred during molecular and cellular evolution, since they rendered the primordial cells less dependent on the exogenous supply of abiotically formed molecules. The evolution of genes, comprising the increase of their size and complexity, is the result of different molecular mechanisms including substitutions and indels mutations, gene duplication, gene fusion, gene elongation, and horizontal transfer of external DNA and its homologous recombination in the host genome. One of the most studied metabolic pathways, which shows a plethora of gene structures and organizations, is histidine biosynthesis and analyses of the structure of his genes revealed that these different molecular mechanisms played an important role in shaping this metabolic route. In this context, this thesis aimed to explore the molecular mechanisms that shaped metabolic pathways during evolution, using the histidine biosynthesis as a model and aiming to experimentally simulate these events in the microorganism Escherichia coli. Through bioinformatic analyses, genome editing techniques and directed evolution experiments, we i) studied the structure and organization of histidine biosynthetic genes in the Bacteroidota-Rhodothermota-Balneolota-Chlorobiota superphylum, highlighting a high variety of genes structures and organizations and allowing to suggest a possible hypothesis for the assembly in operons of his genes during this taxonomic group evolution; ii) analyzed the compartmentalization of histidine biosynthetic enzymes in E. coli, demonstrating the in vivo interaction between HisF and HisH enzymes; iii) investigated the evolutionary molecular mechanisms of gene elongation, frameshift mutation and homologous recombination using the hisF gene as a model, simulating its possible early evolution; iv) explored the HisF involvement in different cellular processes in the bacterial world, suggesting its central role in cellular metabolism. Results obtained from the proposed analyses could represent a further step towards the understanding of metabolic pathways evolution.