2015 Annual Science Report
University of Montana, Missoula Reporting | JAN 2015 – DEC 2015
Project 3: Consequences of recA Duplication for Recombination, Genome Stability and Fitness
Homologous recombination (HR) – the exchange of genetic information between similar DNA molecules – is an ancient process that is central to the emergence of biological complexity, diversity and stability. Yet, it must be tightly regulated, as it is likewise an important source of destabilizing genomic rearrangements. Despite the importance of HR, we still have a poor understanding of the balance of these creative, stabilizing and destabilizing contributions to organismal fitness, complexity and genome evolution. We are using the extraordinary genome evolutionary dynamics and duplicated copies of the HR gene recA in the cyanobacterium Acaryochloris as a model to gain novel insights on the fitness consequences that emerge from the interplay between HR-mediated maintenance of genome stability, selectively favored gene duplications and non-adaptive genomic rearrangements.
We have made substantial progress during the first year of the award period, detailed below.
(1) Development of an E. coli model for the characterization of Acaryochloris RecAs. A major question regarding the molecular evolutionary origins of biological diversity is what happens to a generalist enzyme like RecA upon duplication. We have developed tools for investigating the functional divergence of Acaryochloris RecAs using the genetic model bacterium E. coli. We constructed plasmids to specifically induce the expression of recA genes from Acaryochloris in E. coli in the presence of the sugar rhamnose. We can detect both RNA transcripts (Fig. 1) and protein for Acaryochloris RecAs, indicating that the genes are successfully expressed. We next tested whether the Acaryochloris genes could complement (i.e., rescue) a number of defects observed for an E. coli mutant that lacks its own copy of recA. We observed that no Acaryochloris recA could complement the well-characterized sensitivity of the mutant to ultraviolet radiation or mutagens such as mitomycin C; however, this appears to be a general property of cyanobacterial RecAs that we tested. Interestingly, they could complement the general growth defect of the mutant, which requires recombinational repair activity, to different degrees (Fig. 2), indicating that: (1) Acaryochloris recAs have indeed functionally diverged; and (2) all copies have recombinase activity, consistent with our model that multiple recA copies contribute to the extraordinary genome instability of Acaryochloris through enhanced recombination.
PROJECT INVESTIGATORS:Scott Miller
PROJECT MEMBERS:Amy Gallagher
RELATED OBJECTIVES:Objective 4.2
Production of complex life.
Environment-dependent, molecular evolution in microorganisms
Biochemical adaptation to extreme environments
Effects of environmental changes on microbial ecosystems
Adaptation and evolution of life beyond Earth