Unraveling a tangled skein: Evolutionary analysis of the bacterial gibberellin biosynthetic operon

Abstract

Running title: Evolution of the bacterial gibberellin operon 12 13ABSTRACT 14Gibberellin (GA) phytohormones are ubiquitous regulators of growth and developmental processes 15 in vascular plants. The convergent evolution of GA production by plant-associated bacteria, including both 16 symbiotic, nitrogen-fixing rhizobia and phytopathogens, suggests that manipulation of GA signaling is a 17 powerful mechanism for microbes to gain an advantage in these interactions. Although homologous 18 operons encode GA biosynthetic enzymes in both rhizobia and phytopathogens, notable genetic 19 heterogeneity and scattered operon distribution in these lineages suggests distinct functions for GA in varied 20 plant-microbe interactions. Therefore, deciphering GA operon evolutionary history could provide crucial 21 evidence for understanding the distinct biological roles for bacterial GA production. To further establish 22The clustering of bacterial biosynthetic genes within operons allows for the controlled co-33 expression of functionally-related genes under a single promoter, and the opportunity for these genes to be 34 mobilized and co-inherited as a complete metabolic unit via horizontal gene transfer (HGT) [1, 2]. Because 35 operons are responsible for many fundamental biosynthetic pathways in bacteria, analysis of the genetic 36 structure of complex operons can provide important clues regarding the selective pressures driving the 37 evolution of bacterial metabolism, and can also give insight into the occurrences and mechanisms of HGT. 38The ability for bacteria to produce gibberellin (GA), a ubiquitous plant hormone, is imparted by a 39 GA biosynthetic operon (GA operon; Figure 1), which is found in both nitrogen-fixing rhizobia and 40 phytopathogenic bacteria [3][4][5]. While the diterpenoid GA phytohormones act as endogenous signaling 41 molecules for growth and development in vascular plants [6], plant-associated fungi and bacteria have 42 convergently evolved the ability to produce GA as a mechanism for host manipulation [4,[7][8][9]. The 43 phenomenon of GA production by plant-associated microbes has important biological implications, as 44 perturbation in GA signaling can lead to extreme phenotypic changes in plants. For example, production of 45 GA by the rice pathogen Gibberella fujikuroi leads to dramatic elongation and eventual lodging of rice 46 crops [10], and impaired GA metabolism is responsible for the semi-dwarf crop phenotypes associated with 47 aforementioned genes other than cyp115, is widely distributed in symbiotic nitrogen-fixing rhizobia from 59 the alphaproteobacteria class (α-rhizobia) [15], and biochemical characterization of GA operon genes in 60 several α-rhizobia, including B. diazoefficiens, Sinorhizobium fredii, and Mesorhizobium loti, has 61 demonstrated that this core operon is responsible for biosynthesis of GA9, the penultimate intermediate to 62the bioactive phytohormone GA4 [3, 4,[16][17][18]. While exclusively found in plant-associated bacteria [19], 63 the GA operon exhibits scattered d...