(390e) Higher Order Gene Deletion Mutants Give Insights On Latent Pathway Activation

It has been well established that deletion of metabolic genes causes a local redistribution of fluxes which results in a sub-optimal growth rate of the mutants¹. However, the mutants can adaptively evolve to achieve the optimal FBA predicted growth rates². Observations on temporal flux profiles of single gene deletion mutants have shown that latent pathways are transiently activated during the course of adaptive evolution³. The reason for the activation of these pathways, however, still remains speculative. Recently, it has been proposed that the presence of these pathways adversely affects the growth rates during the sub-optimal growth phase⁴. Furthermore, it is not well understood whether the activation of latent pathways is a suboptimal response to the genetic perturbations or whether it is a global non-specific response.

We attempted to understand the phenomenon using higher order gene deletion mutants, which showed different growth phenotypes on different carbon sources. Two Escherichia coli mutants, one with three gene deletions (LMSE2) and one with five gene deletions (LMSE5), were used in the study. The strategic gene deletions in the two mutants were expected to give insights on the latent pathways activation under different media conditions. Using the genome-scale model iAF1260, the latent pathways for both the mutants were identified as the reactions that were activated during the sub-optimal growth phase (predicted using MOMA) and but not active during optimal growth phase (predicted using FBA)⁴. Abilities of the mutants to recover on different carbon sources after gene deletions, was used to predict the activation of the latent pathways in these mutants.

Our experimental observations suggest that latent pathways are deterministic, intermediate states that are specifically recruited to support growth. In addition to this, our results indicate that the ability of the mutants to return to the original growth state plays a major role in the recruitment of latent pathways and hence, determines the mutant’s survival.  It has been previously suggested that mutants do not recover in the event of major flux re-routing. Our results are in agreement with this observation and show that in E. coli, the recovery of the mutant strongly correlates to the ability of the mutants to restore the original flux distribution, with the help of transient activation of latent pathways.

References

1. Segrè, D., Vitkup, D. & Church, G. M. Analysis of optimality in natural and perturbed metabolic networks. Proc. Natl. Acad. Sci. U. S. A. 99, 15112-15117 (2002).

2. Fong, S. S. & Palsson, B. Ø. Metabolic gene-deletion strains of Escherichia coli evolve to computationally predicted growth phenotypes. Nat. Genet. 36, 1056-1058 (2004).

3. Fong, S. S., Nanchen, A., Palsson, B. O. & Sauer, U. Latent pathway activation and increased pathway capacity enable Escherichia coli adaptation to loss of key metabolic enzymes. J. Biol. Chem. 281, 8024-8033 (2006).

4. Cornelius, S., Lee, J. & Motter, A. Dispensability of Escherichia coli's latent pathways. Proc. Natl. Acad. Sci. U. S. A. 108, 3124-3129 (2011).