Stress-Survival Gene Identification From an Acid Mine Drainage Algal Mat Community

Tuesday, 16 December 2014
Jesica Urbina-Navarrete1,2, Kosuke Fujishima1, Ivan G Paulino-Lima1, Brooke Rothschild-Mancinelli3 and Lynn J Rothschild1, (1)NASA Ames Research Center, Biospheric Science Brance, Moffett Field, CA, United States, (2)University of California Santa Cruz, Microbiology and Environmental Toxicology, Santa Cruz, CA, United States, (3)University of Edinburgh, Edinburgh, United Kingdom
Microbial communities from acid mine drainage environments are exposed to multiple stressors to include low pH, high dissolved metal loads, seasonal freezing, and desiccation. The microbial and algal communities that inhabit these niche environments have evolved strategies that allow for their ecological success. Metagenomic analyses are useful in identifying species diversity, however they do not elucidate the mechanisms that allow for the resilience of a community under these extreme conditions. Many known or predicted genes encode for protein products that are unknown, or similarly, many proteins cannot be traced to their gene of origin. This investigation seeks to identify genes that are active in an algal consortium during stress from living in an acid mine drainage environment. Our approach involves using the entire community transcriptome for a functional screen in an Escherichia coli host. This approach directly targets the genes involved in survival, without need for characterizing the members of the consortium.

The consortium was harvested and stressed with conditions similar to the native environment it was collected from. Exposure to low pH (< 3.2), high metal load, desiccation, and deep freeze resulted in the expression of stress-induced genes that were transcribed into messenger RNA (mRNA). These mRNA transcripts were harvested to build complementary DNA (cDNA) libraries in E. coli. The transformed E. coli were exposed to the same stressors as the original algal consortium to select for surviving cells. Successful cells incorporated the transcripts that encode survival mechanisms, thus allowing for selection and identification of the gene(s) involved. Initial selection screens for freeze and desiccation tolerance have yielded E. coli that are 1 order of magnitude more resistant to freezing (0.01% survival of control with no transcript, 0.2% survival of E. coli with transcript) and 3 orders of magnitude more resistant to desiccation (0.005% survival of control cells with no transcripts, 5% survival of cells with transcript).

This work is transformative because genetic functions can be selected without having prior knowledge of the genes or of the organisms involved. Work continues to identify the genes responsible for tolerance to extreme conditions and the bio-mechanisms involved.