Convergent genetic adaptation of Escherichia coli in minimal media leads to pleiotropic divergence.

Adaptation in an environment can either be beneficial, neutral or disadvantageous in another. To test the genetic basis of pleiotropic behaviour, we evolved six lines of E. coli independently in environments where glucose and galactose were the sole carbon sources, for 300 generations. All six lines in each environment exhibit convergent adaptation in the environment in which they were evolved. However, pleiotropic behaviour was observed in several environmental contexts, including other carbon environments. Genome sequencing reveals that mutations in global regulators rpoB and rpoC cause this pleiotropy. We report three new alleles of the rpoB gene, and one new allele of the rpoC gene. The novel rpoB alleles confer resistance to Rifampicin, and alter motility. Our results show how single nucleotide changes in the process of adaptation in minimal media can lead to wide-scale pleiotropy, resulting in changes in traits that are not under direct selection.

Rapid evolution of pre-zygotic reproductive barriers in allopatric populations.

IMPORTANCE: A population diversifies into two or more species-such a process is known as speciation. In sexually reproducing microorganisms, which barriers arise first-pre-mating or post-mating? In this work, we quantify the relative strengths of these barriers and demonstrate that pre-mating barriers arise first in allopatrically evolving populations of yeast, Saccharomyces cerevisiae. These defects arise because of the altered kinetics of mating of the participating groups. Thus, our work provides an understanding of how adaptive changes can lead to diversification among microbial populations.

Determination of the Mating Efficiency of Haploids in Saccharomyces cerevisiae.

Saccharomyces cerevisiae is a widely used model organism in genetics, evolution, and molecular biology. In recent years, it has also become a popular model organism to study problems related to speciation. The life cycle of yeast involves both asexual and sexual reproductive phases. The ease of performing evolution experiments and the short generation time of the organism allow for the study of the evolution of reproductive barriers. The efficiency with which the two mating types (a and α) mate to form the a/α diploid is referred to as the mating efficiency. Any decrease in the mating efficiency between haploids indicates a pre-zygotic barrier. Thus, to quantify the extent of reproductive isolation between two haploids, a robust method to quantify the mating efficiency is required. To this end, a simple and highly reproducible protocol is presented here. The protocol involves four main steps, which include patching the haploids on a YPD plate, mixing the haploids in equal numbers, diluting and plating for single colonies, and finally, calculating the efficiency based on the number of colonies on a drop-out plate. Auxotrophic markers are employed to clearly make the distinction between haploids and diploids.

Public good-driven release of heterogeneous resources leads to genotypic diversification of an isogenic yeast population.

Understanding the basis of biological diversity remains a central problem in evolutionary biology. Using microbial systems, adaptive diversification has been studied in (a) spatially heterogeneous environments, (b) temporally segregated resources, and (c) resource specialization in a homogeneous environment. However, it is not well understood how adaptive diversification can take place in a homogeneous environment containing a single resource. Starting from an isogenic population of yeast Saccharomyces cerevisiae, we report rapid adaptive diversification, when propagated in an environment containing melibiose as the carbon source. The diversification is driven due to a public good enzyme α-galactosidase, which hydrolyzes melibiose into glucose and galactose. The diversification is driven by mutations at a single locus, in the GAL3 gene in the S. cerevisiae GAL/MEL regulon. We show that metabolic co-operation involving public resources could be an important mode of generating biological diversity. Our study demonstrates sympatric diversification of yeast starting from an isogenic population and provides detailed mechanistic insights into the factors and conditions responsible for generating and maintaining the population diversity.

Molecular Architectonics of Cyclic Dipeptide Amphiphiles and Their Application in Drug Delivery.

Assembly and co-assemblies of peptide amphiphiles through specific noncovalent forces expand the space of molecular architectonics-driven construction of diverse nanoarchitectures with potential biological applications. In this work, cyclic dipeptide amphiphiles (CDPAs) of cyclo(Gly-Asp) with varying lengths of alkyl chains (C8-C18) were synthesized, and their molecular organization was studied. The noncovalent interactions of the components, CDP and alkyl chain, drive the molecular self-assembly of CDPAs into well-defined and diverse nanoarchitectures such as nanotubes, nanospheres, nano/microsheets, and flowers. The co-assembly of CDPAs with biological molecules such as nucleosides was studied to ascertain their utility as potential drug delivery vehicles. Mechanical properties of these nanoarchitectures in nanoindentation study established them as robust in nature. A temperature-dependent NMR study confirmed the formation of stable co-assembly of CDPAs, primarily driven by the intermolecular hydrogen bonding interactions. Computational modeling of oligomers of CDPAs and their co-assembly with nucleosides/nucleotides reveal the molecular level interactions and driving force behind such assemblies. CDPAs exhibit good biocompatibility and cytocompatibility, as revealed by the cellular studies which substantiated their suitability for drug delivery applications. The co-assembly of CDPA with an anticancer drug 5-bromo-2'-deoxyuridine (BrdU) was studied as a drug delivery platform and cytotoxicity was successfully assessed in HeLa cells. Computational modeling of the oligomers of CDPAs and their co-assembly with the drug molecule was performed to understand the molecular level interactions and driving force behind the assemblies. Our findings reveal the design strategy to construct diverse structural architectures using CDP as the modular building unit and specific molecular interactions driven co-assembly for potential application as drug delivery carrier.