Biology subjects

Asamoto, C. K.

Publications and source records attributed to Asamoto, C. K..

2 recordsLinked to original sources

bioRxiv2020

Enzyme specific isotope effects of the Nap and Nar nitrate reductases

Dissimilatory nitrate reduction (DNR) to nitrite is the first step in denitrification, the main process through which bioavailable nitrogen is removed from ecosystems. DNR fractionates the stable isotopes of nitrogen (14N, 15N) and oxygen (16O, 18O) and thus imparts an isotopic signature on residual pools of nitrate in many environments. Data on the relationship between the resulting isotopic pattern in oxygen versus nitrogen isotopes (18{varepsilon} / 15{varepsilon}) suggests systematic differences exist between marine and terrestrial ecosystems that are not fully understood. DNR can be catalyzed by both cytosolic (Nar) and periplasmic (Nap) nitrate reductases, and previous work has revealed differences in their 18{varepsilon} / 15{varepsilon} isotopic signatures. In this study, we thus examine the 18{varepsilon} / 15{varepsilon} of six different nitrate-reducing microorganisms that encode Nar, Nap or both enzymes, as well gene deletion mutants of the enzymes catalytic subunits (NarG and NapA) to test the hypothesis that enzymatic differences alone could explain the environmental observations. We find that the distribution of the 18{varepsilon} / 15{varepsilon} fractionation ratios of all examined nitrate reductases form two distinct, non-overlapping peaks centered around a 18{varepsilon} / 15{varepsilon} proportionality of 0.55 and a 18{varepsilon} / 15{varepsilon} proportionality of 0.91, respectively. All Nap reductases studied to date cluster around the lower proportionality (0.55) and none exceed a 18{varepsilon} / 15{varepsilon} proportionality of 0.68. Almost all Nar reductases, on the contrary, cluster tightly around the higher proportionality (0.91) with no values below a 18{varepsilon} / 15{varepsilon} proportionality of 0.84 with the notable exception of the Nar reductases from the genus Bacillus which fall around 0.62 and thus closely resemble the isotopic fingerprints of the Nap reductases. Our findings confirm the existence of two remarkably distinct isotopic end-members in the dissimilatory nitrate reductases that could indeed explain differences in coupled N and O isotope fractionation between marine and terrestrial systems, and almost but not fully match reductase phylogeny.

microbiology↗

bioRxiv2016

Natural Genetic Variation Modifies Gene Expression Dynamics at the Protein Level During Pheromone Response in Saccharomyces cerevisiae

Heritable variation in gene expression patterns plays a fundamental role in trait variation and evolution, making understanding the mechanisms by which genetic variation acts on gene expression patterns a major goal for biology. Both theoretical and empirical work have largely focused on variation in steady-state mRNA levels and mRNA synthesis rates, particularly of protein-coding genes. Yet in order for this variation to affect higher order traits it must lead to differences at the protein level. Variation in protein-specific processes including protein synthesis rates and protein decay rates could amplify, mask, or even reverse effects transmitted from the transcript level, but the extent to which this happens is unclear. Moreover, mechanisms that underlie protein expression variation under dynamic conditions have not been examined. To address this challenge, we analyzed how mRNA and protein expression dynamics covary between two strains of Saccharomyces cerevisiae during mating pheromone response. Although divergent steady-state mRNA expression levels explained divergent steady-state protein levels for four out of five genes in our study, the same was true for only one out of five genes for expression dynamics. By integrating decay rate and allele-specific protein expression analyses, we resolved that expression divergence for Fig1p was caused by genetic variation acting in trans on protein synthesis rate, expression divergence for Ina1p was caused by cis-by-trans epistatic effects on transcript level and protein synthesis rate, and expression divergence for Fus3p and Tos6p were caused by divergence in protein synthesis rates. Our study demonstrates that steady-state analysis of gene expression is insufficient to understand the impact of genetic variation on gene expression variation. An integrated and dynamic approach to gene expression analysis - comparing mRNA levels, protein levels, protein decay rates, and allele-specific protein expression - allows for a detailed analysis of the genetic mechanisms underlying protein expression divergences.

genetics↗