1. OEB 100 – 11.01.24 Evolution in Action (OEB 100) Instructor: Christopher Marx Teaching fellow: Dipti Nayak Weekly meeting: NW B127 Time: Mondays 4 – 5:30 pm Laboratory: NW 152 Time: 24/7/365 open access Website: http://isites.harvard.edu/k77185 Please let me out of here: my barnacles and pigeons need me!

2. Integration of systems-level physiology and evolution genotype phenotype environment physiology Molecular, cell & developmental biology, biochem.

3. Integration of systems-level physiology and evolution genotype phenotype mutation environment genotype phenotype genotype phenotype ecological interactions selection & drift Evolutionary biology, ecology

4. Integration of systems-level physiology and evolution genotype phenotype mutation environment genotype phenotype genotype phenotype ecological interactions selection & drift physiology “Biology” Use adaptation of laboratory populations of microbes as a forum to explore how biological systems evolve. The “functional synthesis” (Dean & Thornton, 2007)

5. Could tackle all levels through various aspects of the project… genotype phenotype genotype phenotype genotype phenotype Sequence genomes; parallelism?; mutational trajectories? Assay external medium conditions Adaptive diversification? Competition assays; tradeoffs & recovery of past abilities? Global gene expression?

6. Model system: Methylobacterium M. extorquens is a plant epiphyte Model for C 1 growth (~50 years): C 1 compounds oxidized to formaldehyde Oxidation of formaldehyde to CO 2 Assimilation of C 1 units into biomass Requires ~100 genes for C 1 growth C 1 genes acquired via HGT Limited growth on multi-C compounds Genomes now available for 8 species including the one we’ll study (M. extorquens PA1) CH 3 -R HCHO CO 2 biomass

7. t=1t=2t=……. Ancestor Evolved isolates Selection largely for growth rate Living fossil record 1/64 dilution every 2 days (initially 4 days), N final = ~2x10 9 Experimental evolution You’ll use 48- well plates in an automated, robotic system

8. EvolvedFluor. ancestor T0T0 CompetitionFlow Cytometry Competitive growth T1T1 T1T1 fluorescence EvolvedFluor. ancestor T0T0 fluorescence (Lee et al., 2009. Evolution) Fitness assayed via competitions

9. Project: Using evolution to study bacterial resistance growth on the toxic chemical formaldehyde Methylobacterium

10. Living Cell Formaldehyde is a key metabolic intermediate…. Formaldehyde (1mM) But the cells cannot grow on it….

11. Evolution to Grow on Formaldehyde…..We now have 4 populations that can grow on 30 mM formaldehyde

12. Generic increase in stress Response? Has the ability to grow on Formaldehyde conferred a generic ability to withstand stress better? Students will subject the evolved and ancestral isolates to several stresses such as : High/Low Temperature Antibiotics Detergents UV exposure …… Genetic mechanisms underlying stress response will be analyzed. Sub-Projects

13. Coexistence of different genotypes? Genotype 1 : Can withstand high concentrations of Formaldehyde which leads to penalty in growth at lower concentrations. Genotype 2 : Cannot withstand high concentrations of Formaldehyde and grows faster at lower concentrations.

14. Tradeoffs between substrates? How has the ability to grow on formaldehyde affected growth on other compounds? We know that isolates from one population cannot grow on most carbon compounds it previously could. So there are MASSIVE tradeoffs? What are they? Can evolving these populations in an alternate environment help these isolates recover? (Lee et.al. Evolution 2009)

15. Genomics and Genetics Pyrosequencing evolved strains to determine genetic loci that have undergone mutations. Examining these loci in replicate populations to check for parallel mutational events. Reconstructing mutations in the ancestral background to determine the order of mutational events and the phenotypic change caused by individual mutations. Conducting Microarray/RNA sequencing to understand patterns of global gene expression in evolved and ancestral strains.

16. Formaldehyde-Induced Mutagenesis? Mutations arise spontaneously (one in ~300 cells), but at rates that can be affected by mutations (in DNA repair) or by the environment. From genome resequencing we’ll know if there was an elevated mutation rate. If so we can determine if formaldehyde specifically induces a higher mutation rate in the original strain, or whether the evolved strain has lowered (or increased) this rate.

17. From first time in 2009…

18. Last spring…

19. Undergraduate Diversity at the Evolution meetings 2011 We are pleased to announce a NSF-funded program for bringing talented and diverse undergraduates to the Evolution meetings this June 17-21 in Norman, Oklahoma. For the ninth year in a row we will fly a cohort of 25 undergraduates from throughout the US and Puerto Rico to present a poster at the meetings, receive mentoring from graduate students, postdocs and faculty, and participate in the Diversity Social as well as a career-oriented 'Undergraduate Futures in Evolutionary Biology' panel and discussion. The program covers the costs of travel, registration, food and accommodation at the meetings. The application deadline is 1 March, but admissions will be reviewed as they are received, and so the earlier you apply, the better. Applications are welcomed from all undergraduates, and the admissions goal is to create a diverse pool of students. An overview of the program and student eligibility can be found at: http://www.oeb.harvard.edu/faculty/edwards/community/application.html Apply online at: www.nescent.org/Evolution2011_application Applications consist of a short statement of interest, a letter of recommendation and the title and abstract of the poster to be presented. In addition, we will be soliciting names of graduate students, postdocs and faculty members who would like to serve as mentors during the meetings. Mentors take pairs of students and attend talks with them, introduce them to colleagues, network and generally make the meetings a welcoming place for them. Although costs are not covered for mentors it is an unusually rewarding experience. Contact Richard Kliman if you are interested in serving as a mentor.rmkliman@cedarcrest.edu For inquires contact one of the organizers: Scott Edwards - sedwards@oeb.harvard.edusedwards@oeb.harvard.edu Richard Kliman - rmkliman@cedarcrest.edurmkliman@cedarcrest.edu Jory Weintraub - jory@nescent.orgjory@nescent.org

20. Monday (1/31): Detailed description of project details and general fundamentals Read: Elena & Lenski, 2003. Nature Reviews Genetics. Evolution experiments with microorganisms: The dynamics and genetic bases of adaptation Lee et al., 2010. Nature. Bacterial charity work leads to population-wide resistance