Protein synthesis translates information from messenger RNAs into functional proteomes. Because of the finite nature of the resources required by the translational machinery, both the overall protein synthesis activity of a cell and activity on individual mRNAs are controlled by the allocation of limiting resources. Upon introduction of heterologous sequences into an organism—for example for the purposes of bioprocessing or synthetic biology—limiting resources may also become overstretched, thus negatively affecting both endogenous and heterologous gene expression. In this study, we present a mean-field model of translation in Saccharomyces cerevisiae for the investigation of two particular translational resources, namely ribosomes and aminoacylated tRNAs. We firstly use comparisons of experiments with heterologous sequences and simulations of the same conditions to calibrate our model, and then analyse the behaviour of the translational system in yeast upon introduction of different types of heterologous sequences. Our main findings are that: competition for ribosomes, rather than tRNAs, limits global translation in this organism; that tRNA aminoacylation levels exert, at most, weak control over translational activity; and that decoding speeds and codon adaptation exert strong control over local (mRNA specific) translation rates.

SummaryWe have recently cloned an oligopeptide transport gene from Candida albicans denoted OPT1. This gene showed significant sequence similarity to three open reading frames (ORFs) with no previously established function: isp4 from Schizosaccharomyces pombe and Saccharomyces cerevisiae YJL212C and YPR194C, identified during the genome project. The S. pombe gene isp4 was originally identified by Sato et al. as a gene that was upregulated through nitrogen starvation induction of meiosis. However, an isp4⌬ strain exhibited a wild-type phenotype with respect to sexual differentiation. We have found that the same isp4⌬ strain is deficient in tetrapeptide transport activity as measured by its resistance to toxic tetrapeptides, by its inability to accumulate a radiolabelled tetrapeptide and by the inability to use tetrapeptides as a sole source of an amino acid to satisfy an auxotrophic requirement. Similarly, we found that the ORF YPR194C from S. cerevisiae encodes an oligopeptide transporter. Sequence analyses as well as physiological evidence has led us to propose that the proteins encoded by isp4 and the genes identified from S. cerevisiae and C. albicans comprise a new group of transporters specific for small oligopeptides, which we have named the OPT family.

Bi-phasic or diauxic growth is often observed when microbes are grown in a chemically defined medium containing two sugars (for example glucose and lactose). Typically, the two growth stages are separated by an often lengthy phase of arrested growth, the so-called lag-phase. Diauxic growth is usually interpreted as an adaptation to maximise population growth in multi-nutrient environments. However, the lag-phase implies a substantial loss of growth during the switch-over. It therefore remains unexplained why the lag-phase is adaptive. Here we show by means of a stochastic simulation model based on the bacterial PTS system that it is not possible to shorten the lag-phase without incurring a permanent growth-penalty. Mechanistically, this is due to the inherent and well established limitations of biological sensors to operate efficiently at a given resource cost. Hence, there is a trade-off between lost growth during the diauxic switch and the long-term growth potential of the cell. Using simulated evolution we predict that the lag-phase will evolve depending on the distribution of conditions experienced during adaptation. In environments where switching is less frequently required, the lag-phase will evolve to be longer whereas, in frequently changing environments, the lag-phase will evolve to be shorter.

We have identified an oligopeptide transporter in the yeast Saccharomyces cerevisiae which mediates the uptake of tetra-and pentapeptides, including the endogenous opioids leucine enkephalin (Tyr-Gly-Gly-Phe-Leu) and methionine enkephalin (Tyr-Gly-Gly-Phe-Met). The transporter is encoded by the gene OPT1. Yeast expressing OPT1 can utilize enkephalins to satisfy amino acid auxotrophic requirements for growth. The transport of radiolabeled leucine enkephalin exhibits saturable kinetics, with a K m of 310 M. Transport activity is optimum at acidic pH and sensitive to reagents which uncouple oxidative phosphorylation, suggesting an energy dependence on the proton gradient. Growth, transport, and chromatographic data indicate that leucine enkephalin is not hydrolyzed in the extracellular medium and as such is translocated intact across the cell membrane. The system is specific for tetra-and pentapeptides and can be inhibited by the opioid receptor antagonists naloxone and naltrexone. To date, this is the first example of a eukaryotic transport system which can use enkephalins as a substrate, opening the possibility that a homologue exists in higher eukaryotes.

In this paper, we describe a recent trend in the introductory computer science curriculum which advocates conceptualizing computation primarily as coordinated concurrent activities [8], [9], [10]. Consistent with this philosophy is the focus on the event-driven model of computation [7]. While one can utilize these approaches with any thread and/or event supporting object-oriented language (e.g. Java) in a desktop programming environment, they become particularly worthwhile when used in conjunction with physical robots. This paper argues the case for the benefits of this approach and provides sample exercises that illustrate the use of this pedagogy using Lego Mindstorms RCX bricks programmed in Java for use in introductory programming.