Figure 2.

Fitness contributions of TFBs across diverse environments reveal their complex and overlapping functions. Growth assays were performed in high throughput by tracking cell density at OD₆₀₀ using the Bioscreen C instrument as described in Materials and methods. We determined the maximum growth rates (fitness) from smooth spline fitted growth curves after depositing cell density measurements into a database with relevant meta-information and associated plate layout information. Maximum growth rate of each TFB knockout was normalized to appropriate controls and log₂ ratios were reported as normalized maximum growth rates or fitness (Supplementary Table S3). (A) Distinct trends in fitness contribution of TFBs across specific environmental gradients. The condition-specific fitness trends (normalized maximum growth rate) of each TFB knockout strain can be viewed as evidence for complex patterns of subfunctionalizations. (B) Relative order of fitness contributions of TFBs changes with environmental context. Fitness of each TFB knockout was subtracted from fitness of the parent to obtain degree of fitness contributed by that TFB in each environment (plotted on the y axis as ‘TFB Fitness’). Statistical significance of fitness differences among pairs of TFBs was calculated using t-test (Supplementary Figure S1). Starting with the lowest fitness contributing TFB on the left boxplots of the TFBs are rank ordered with increasing fitness contributions going rightward. The different orderings of the TFBs in these rank-ordered plots demonstrate how TFBs take turns in assuming a primary role across the 17 environmental conditions. (C) Distribution of different clades of TFBs across all of the 11 fully sequenced halophilic archaeal genomes. Clade membership of TFBs was assigned based on similarity to H. salinarum NRC-1 family members. Numbers in parenthesis indicate total number of TFB proteins in each species. While TFBf- and TFBc/TFBg-like proteins are present in all archaea, TFBb/TFBd- and TFBa/TFBe-like proteins are limited to certain species (Supplementary Table S1).