Biology subjects

Domitrovic, T.

Publications and source records attributed to Domitrovic, T..

8 recordsLinked to original sources

bioRxiv2025

Translation efficiency changes at heat shock in Saccharomyces cerevisiae

Both eukaryotic and prokaryotic cells respond to heat shock by engaging multiple regulatory mechanisms that preserve proteostasis, including adjustments in transcription, translation, and mRNA metabolism. Although the transcriptional response to heat stress in Saccharomyces cerevisiae has been well characterized, the extent to which translation efficiency (TE) is remodeled and how this remodeling contributes to protein synthesis remains less understood. Here, analysis of previously published ribosome profiling and RNA-seq data revealed that in S. cerevisiae, the TE varies during heat shock, depending on both temperature and stress duration. In contrast, TE remained largely stable in Escherichia coli under comparable conditions or during activation of {sigma}32, the main regulator of heat shock response in bacteria. In yeast, TE modulation patterns correlated with changes in protein abundance and differed markedly between early and late stages of stress. At 10 min, transcripts with short 5'UTRs and high codon optimality tended to display higher TE, primarily due to the easier ribosome engagement in these transcripts. By 30 min, the TE pattern across transcripts had changed drastically, suggesting increased availability of newly synthesized stress-response transcripts. These findings support a model in which early TE adjustments depend on intrinsic mRNA properties, whereas shifts in mRNA accessibility and transcriptional output shape later changes. The results highlight TE remodeling as a dynamic component of the heat-shock response in yeast and distinguish this behavior from the more buffered response observed in bacteria.

microbiology↗

bioRxiv2023

Transcriptional profile of ribosome-associated quality control components and their associated phenotypes in mammalian cells

During protein synthesis, organisms detect translation defects that induce ribosome stalling and result in protein aggregation. The Ribosome-associated Quality Control (RQC) complex, comprising TCF25, LTN1, and NEMF, is responsible for identifying incomplete protein products from unproductive translation events, targeting them for degradation. Though RQC disruption causes adverse effects on vertebrate neurons, data regarding mRNA/protein expression and regulation across tissues are lacking. Employing high-throughput methods, we analyzed public datasets to explore RQC gene expression and phenotypes. Our findings revealed a widespread expression of RQC components in human tissues; however, silencing of RQC yielded only mild negative effects on cell growth. Notably, TCF25 exhibited elevated mRNA levels that were not reflected in protein content. We experimentally demonstrated that this disparity arises from post-translational protein degradation by the proteasome. Additionally, we observed that cellular aging marginally influences RQC expression, leading to reduced mRNA levels in specific tissues. Our results suggest the necessity of RQC expression in all mammalian tissues. Nevertheless, when RQC falters, alternative mechanisms seem to compensate, ensuring cell survival under non-stress conditions.

bioinformatics↗

bioRxiv2021

A systematic structural comparison of all solved small proteins (4-6 kDa) reveals the weight of disulfide bonds in proteins' foldability

Defensins are small proteins, usually ranging from 4 to 6 kDa, amphipathic, disulfide-rich, and with a small or even absent hydrophobic core. Since a hydrophobic core is generally found in globular proteins that fold in an aqueous solvent, the peculiar fold of defensins can challenge tertiary protein structure predictors. We performed a PDB-wide survey of small proteins (4-6 kDa) to understand the similarities of defensins with other small disulfide-rich proteins. We found no differences when we compared defensins with non-defensins regarding the proportion and exposition to the solvent of apolar, polar, and charged residues. Then we divided all small proteins (4-6 kDa) deposited in PDB into two groups, one group with at least one disulfide bond (bonded, defensins included) and another group without any disulfide bond (unbonded). The group of bonded proteins presented apolar residues more exposed to the solvent than the unbonded group. The ab initio algorithm for tertiary protein structure prediction Robetta was more accurate to predict unbonded than bonded proteins. Our work highlights one more layer of complexity for the tertiary protein prediction structure: small disulfide-rich proteins ability to fold even with a poor hydrophobic core.

biochemistry↗

bioRxiv2020

Natural variation of the cardiac transcriptome in humans

We investigated the gene-expression variation among humans by analyzing previously published mRNA-seq and ribosome footprint profiling (RP) of heart left-ventricles from healthy donors. We ranked the genes according to their coefficient of variation values and found that the top 5% most variable genes had special features compared to the rest of the genome, such as lower mRNA levels and shorter half-lives coupled to increased translation efficiency. We observed that these genes are mostly involved with immune response and have a pleiotropic effect on disease phenotypes, indicating that asymptomatic conditions contribute to the gene expression diversity of healthy individuals.

genomics↗

bioRxiv2019

Endogenous polybasic peptides cause slower movement of ribosomes but are not targets of the ribosome quality control complex (RQC)

Highly positively charged protein segments are known to result in poor translation efficiency. This effect is explained by ribosome stalling caused by electrostatic interactions between the nascent peptide and the negatively charged ribosome exit tunnel, leading to translation termination followed by protein degradation mediated by the RQC complex. These polybasic sequences are mainly studied in the context of artificial reporter systems. Examples of endogenous yeast proteins targeted by the RQC complex are Rqc1, a protein essential for RQC function, and Sdd1. Both contain polybasic sequences that are thought to activate the RQC, leading to protein down-regulation. Here, we investigated whether the RQC complex regulates other endogenous proteins with polybasic sequences. We show by bioinformatics, ribosome profiling data analysis, and western blot that endogenous proteins containing polybasic sequences similar to, or even more positively charged than those of Rqc1 and Sdd1, are not targeted by the RQC complex suggesting that endogenous polybasic sequences are not sufficient to induce this type of regulation. Finally, our results also suggest that Rqc1 is regulated post-translationally by the E3 component of the RQC complex Ltn1, in a manner independent of the RQC complex.

biochemistry↗

bioRxiv2019

The distribution, conservation and correlation with translational efficiency of the first five codons in Escherichia coli genes

Translation initiation is a critical step in the regulation of protein synthesis, and it is subjected to different control mechanisms, such as 5 UTR secondary structure and initiation codon context, that can influence the rates at which initiation and consequentially translation occur. For some genes, translation elongation also affects the rate of protein synthesis. With a GFP library containing nearly all possible combinations of nucleotides from the 3rd to the 5th codon positions in the protein coding region of the mRNA, it was previously demonstrated that some nucleotide combinations increased GFP expression up to four orders of magnitude. While it is clear that the codon region from positions 3 to 5 can influence protein expression levels of artificial constructs, its impact on endogenous proteins is still unknown. Through bioinformatics analysis, we identified the nucleotide combinations of the GFP library in Escherichia coli genes and examined the correlation between the expected levels of translation according to the GFP data with the experimental measures of protein expression. We observed that E. coli genes were enriched with the nucleotide compositions that enhanced protein expression in the GFP library, but surprisingly, it seemed to affect the translation efficiency only marginally. Nevertheless, our data indicate that different enterobacteria present similar nucleotide composition enrichment as E. coli, suggesting an evolutionary pressure towards the conservation of short translational enhancer sequences.

bioinformatics↗

bioRxiv2019

Icosahedral viruses defined by their positively charged domains: a signature for viral identity and capsid assembly strategy

Capsid proteins often present a positively charged arginine-rich region at the N and/or C-termini that for some icosahedral viruses has a fundamental role in genome packaging and particle stability. These sequences show little to no conservation at the amino-acid level and are structurally dynamic so that they cannot be easily detected by common sequence or structure comparison. As a result, the occurrence and distribution of positively charged protein domain across the viral and the overall protein universe are unknown. We developed a methodology based on the net charge calculation of discrete segments of the protein sequence that allows us to identify proteins containing amino-acid stretches with an extremely high net charge. We observed that among all organisms, icosahedral viruses are especially enriched in extremely positively charged segments (Q [≥] +17), with a distinctive bias towards arginine instead of lysine. We used viral particle structural data to calculate the total electrostatic charge derived from the most positively charged protein segment of capsid proteins and correlated these values with genome charge arising from the phosphates of each nucleotide. We obtained a positive correlation (r = 0.91, p-value < 0001) for a group of 17 viral families, corresponding to 40% of all families with icosahedral structures described so far. These data indicated that unrelated viruses with diverse genome types adopt a common underlying mechanism for capsid assembly and genome stabilization based on R-arms. Outliers from a linear fit pointed to families with alternative strategies of capsid assembly and genome packaging.\n\nSignificance StatementViruses can be characterized by the existence of a capsid, an intricate proteinaceous container that encases the viral genome. Therefore, capsid assembly and function are essential to viral replication. Here we specify virus families with diverse capsid structure and sequence, for each capsid packing capacity depends on a distinctive structural feature: a highly positively charged segment of amino acids residues, preferentially made of arginine. We also show that proteins with the same characteristics are rarely found in cellular proteins. Therefore, we identified a conserved viral functional element that can be used to infer capsid assembly mechanisms and inspire the design of protein nanoparticles and broad-spectrum antiviral treatments.

bioinformatics↗

bioRxiv2018

Codon stabilization coefficient as an index for prediction of codon bias and its potential applications.

Different methods of mRNA half-life measurements are available, but genome-wide measurements of mRNA half-life in yeast showed a weak correlation between the methods. Moreover, when we compared mRNA half-life determined by these methods with other cellular measurements such as mRNA and protein abundance low correlation was found. To clarify this matter, we analyzed mRNA half-life datasets from nine different groups to determine the most accurate method of measurement. Since codon optimality is one of the significant determinants of mRNA stability, we used the codon stabilization coefficient (CSC) as a reference for mRNA half-life measurement accuracy. After CSC calculation for each dataset, we find strong positive correlations between the CSC from some datasets with other parameters that reflect codon optimality such as tRNA abundance and ribosome residence time. By the use of CSC parameter, we observed that most genes contain non-optimal codons and that codon bias exists toward optimized and non-optimized genes. We also observed that stretches of non-optimal are not randomly distributed since it causes impacts on translation.

bioinformatics↗