Biological entities are multicomponent systems where each part is directly or indirectly dependent on the others. In effect, a change in a single component might have a consequence on the functioning ...of its partners, thus affecting the fitness of the entire system. In this article, we provide a few examples of such complex biological systems, ranging from ant colonies to a population of amino acids within a single-polypeptide chain. Based on these examples, we discuss one of the central and still challenging questions in biology: how do such multicomponent consortia co-evolve? More specifically, we ask how telomeres, nucleo-protein complexes protecting the integrity of linear DNA chromosomes, originated from the ancestral organisms having circular genomes and thus not dealing with end-replication and end-protection problems. Using the examples of rapidly evolving topologies of mitochondrial genomes in eukaryotic microorganisms, we show what means of co-evolution were employed to accommodate various types of telomere-maintenance mechanisms in mitochondria. We also describe an unprecedented runaway evolution of telomeric repeats in nuclei of ascomycetous yeasts accompanied by co-evolution of telomere-associated proteins. We propose several scenarios derived from research on telomeres and supported by other studies from various fields of biology, while emphasizing that the relevant answers are still not in sight. It is this uncertainty and a lack of a detailed roadmap that makes the journey through the jungle of biological systems still exciting and worth undertaking.
Abstract
In virtually every eukaryotic species, the ends of nuclear chromosomes are protected by telomeres, nucleoprotein structures counteracting the end-replication problem and suppressing ...recombination and undue DNA repair. Although in most cases, the primary structure of telomeric DNA is conserved, there are several exceptions to this rule. One is represented by the telomeric repeats of ascomycetous yeasts, which encompass a great variety of sequences, whose evolutionary origin has been puzzling for several decades. At present, the key questions concerning the driving force behind their rapid evolution and the means of co-evolution of telomeric repeats and telomere-binding proteins remain largely unanswered. Previously published studies addressed mostly the general concepts of the evolutionary origin of telomeres, key properties of telomeric proteins as well as the molecular mechanisms of telomere maintenance; however, the evolutionary process itself has not been analyzed thoroughly. Here, we aimed to inspect the evolution of telomeres in ascomycetous yeasts from the subphyla Saccharomycotina and Taphrinomycotina, with special focus on the evolutionary origin of species-specific telomeric repeats. We analyzed the sequences of telomeric repeats from 204 yeast species classified into 20 families and as a result, we propose a step-by-step model, which integrates the diversity of telomeric repeats, telomerase RNAs, telomere-binding protein complexes and explains a propensity of certain species to generate the repeat heterogeneity within a single telomeric array.
To assess the immediate responses of the yeast cells to telomere defects, we employed the auxin‐inducible degron (AID) enabling rapid depletion of essential (Rap1, Tbf1, Cdc13, Stn1) and ...non‐essential (Est1, Est2, Est3) telomeric proteins. Using two variants of AID systems, we show that most of the studied proteins are depleted within 10–30 min after the addition of auxin. As expected, depletion of essential proteins yields nondividing cells, provided that the strains are cultivated in an appropriate carbon source and at temperatures lower than 28°C. Cells with depleted Cdc13 and Stn1 exhibit extension of the single‐stranded overhang as early as 3 h after addition of auxin. Notably, prolonged incubation of strains carrying AID‐tagged essential proteins in the presence of auxin resulted in the appearance of auxin‐resistant clones, caused at least in part by mutations within the OsTIR1 gene. Upon assessing the length of telomeres in strains carrying AID‐tagged non‐essential telomeric proteins, we found that the depletion of Est1 and Est3 leads to auxin‐dependent telomere shortening. However, the EST3‐AID strain had slightly shorter telomeres even in the absence of auxin. Furthermore, a strain with the AID‐tagged version of Est2 (catalytic subunit of telomerase) not only had shorter telomeres in the absence of auxin but also did not exhibit auxin‐dependent telomere shortening. Our results demonstrate that while AID can be useful in assessing immediate cellular responses to telomere deprotection, each strain must be carefully evaluated for the effect of AID‐tag on the properties of the protein of interest.
Auxin‐inducible degron (AID) is a powerful tool to study early events associated with telomere deprotection. However, one has to be cautious about potential drawbacks such as rapid formation of auxin‐resistant clones, dependency on cultivation conditions, or interference of the AID tag with protein activity.
Take‐away
Auxin‐inducible degron (AID) allows rapid depletion of telomere‐associated proteins.
The AID system is sensitive to temperature and carbon source.
Long‐term cultivation in the presence of auxin causes appearance of auxin‐resistant clones.
The C‐terminal AID tag interferes with function of some of the proteins.
Changes in telomere status are noticeable within hours after depletion of a telomeric factor.
Games and play are proven to be the most efficient means for children to learn. We should make greater efforts to use this tool for university teaching.
Abstract The main aim of this article is to provide a short overview of the research that gradually culminated in the concept of cognitive biology. To a certain extent it can be compared to a ...prelude: as the Merriam Webster dictionary defines it, “a musical section or movement introducing the theme or chief subject (as of a fugue or suite) or serving as an introduction to an opera or oratorio.” At first glance, this may seem to downplay the importance of the pre-cognitive biology period, but the opposite is true. Some preludes (consider, for example, La Traviata or Carmen ) are as rich and beautiful as the acts that follow. We believe this is the case with the story culminating in the birth of cognitive biology, and we present it here in the form of an annotated (and somewhat virtual) interview with its leading actor, Ladislav Kováč.
•We provide a list of yeast mitochondrial protein kinases and phosphatases•972 (~70%) of proteins in yeast mitochondria contain a total of >9,400 phospho-sites.•Phosphorylation affects a large ...repertoire of mitochondrial functions.•Phosphorylation of mitochondrial DNA encoded proteins is rare.•Many proteins are phosphorylated prior to their import into the organelle.
Protein phosphorylation is one of the best-known post-translational modifications occurring in all domains of life. In eukaryotes, protein phosphorylation affects all cellular compartments including mitochondria. High-throughput techniques of mass spectrometry combined with cell fractionation and biochemical methods yielded thousands of phospho-sites on hundreds of mitochondrial proteins. We have compiled the information on mitochondrial protein kinases and phosphatases and their substrates in Saccharomyces cerevisiae and provide the current state-of-the-art overview of mitochondrial protein phosphorylation in this model eukaryote. Using several examples, we describe emerging features of the yeast mitochondrial phosphoproteome and present challenges lying ahead in this exciting field.
Abstract
One powerful strategy of how to increase the complexity of cellular proteomes is through posttranslational modifications (PTMs) of proteins. Currently, there are ∼400 types of PTMs, the ...different combinations of which yield a large variety of protein isoforms with distinct biochemical properties. Although mitochondrial proteins undergoing PTMs were identified nearly 6 decades ago, studies on the roles and extent of PTMs on mitochondrial functions lagged behind the other cellular compartments. The application of mass spectrometry for the characterization of the mitochondrial proteome as well as for the detection of various PTMs resulted in the identification of thousands of amino acid positions that can be modified by different chemical groups. However, the data on mitochondrial PTMs are scattered in several data sets, and the available databases do not contain a complete list of modified residues. To integrate information on PTMs of the mitochondrial proteome of the yeast Saccharomyces cerevisiae, we built the yeast mitochondrial posttranslational modification (y-mtPTM) database (http://compbio.fmph.uniba.sk/y-mtptm/). It lists nearly 20,000 positions on mitochondrial proteins affected by ∼20 various PTMs, with phosphorylated, succinylated, acetylated, and ubiquitylated sites being the most abundant. A simple search of a protein of interest reveals the modified amino acid residues, their position within the primary sequence as well as on its 3D structure, and links to the source reference(s). The database will serve yeast mitochondrial researchers as a comprehensive platform to investigate the functional significance of the PTMs of mitochondrial proteins.
Many fungal species utilize hydroxyderivatives of benzene and benzoic acid as carbon sources. The yeast Candida parapsilosis metabolizes these compounds via the 3-oxoadipate and gentisate pathways, ...whose components are encoded by two metabolic gene clusters. In this study, we determine the chromosome level assembly of the C. parapsilosis strain CLIB214 and use it for transcriptomic and proteomic investigation of cells cultivated on hydroxyaromatic substrates. We demonstrate that the genes coding for enzymes and plasma membrane transporters involved in the 3-oxoadipate and gentisate pathways are highly upregulated and their expression is controlled in a substrate-specific manner. However, regulatory proteins involved in this process are not known. Using the knockout mutants, we show that putative transcriptional factors encoded by the genes OTF1 and GTF1 located within these gene clusters function as transcriptional activators of the 3-oxoadipate and gentisate pathway, respectively. We also show that the activation of both pathways is accompanied by upregulation of genes for the enzymes involved in β-oxidation of fatty acids, glyoxylate cycle, amino acid metabolism, and peroxisome biogenesis. Transcriptome and proteome profiles of the cells grown on 4-hydroxybenzoate and 3-hydroxybenzoate, which are metabolized via the 3-oxoadipate and gentisate pathway, respectively, reflect their different connection to central metabolism. Yet we find that the expression profiles differ also in the cells assimilating 4-hydroxybenzoate and hydroquinone, which are both metabolized in the same pathway. This finding is consistent with the phenotype of the Otf1p-lacking mutant, which exhibits impaired growth on hydroxybenzoates, but still utilizes hydroxybenzenes, thus indicating that additional, yet unidentified transcription factor could be involved in the 3-oxoadipate pathway regulation. Moreover, we propose that bicarbonate ions resulting from decarboxylation of hydroxybenzoates also contribute to differences in the cell responses to hydroxybenzoates and hydroxybenzenes. Finally, our phylogenetic analysis highlights evolutionary paths leading to metabolic adaptations of yeast cells assimilating hydroxyaromatic substrates.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
ABSTRACT
The 3-oxoacyl-CoA thiolases catalyze the last step of the fatty acid β-oxidation pathway. In yeasts and plants, this pathway takes place exclusively in peroxisomes, whereas in animals it ...occurs in both peroxisomes and mitochondria. In contrast to baker's yeast Saccharomyces cerevisiae, yeast species from the Debaryomycetaceae family also encode a thiolase with predicted mitochondrial localization. These yeasts are able to utilize a range of hydroxyaromatic compounds via the 3-oxoadipate pathway the last step of which is catalyzed by 3-oxoadipyl-CoA thiolase and presumably occurs in mitochondria. In this work, we studied Oct1p, an ortholog of this enzyme from Candida parapsilosis. We found that the cells grown on a 3-oxoadipate pathway substrate exhibit increased levels of the OCT1 mRNA. Deletion of both OCT1 alleles impairs the growth of C. parapsilosis cells on 3-oxoadipate pathway substrates and this defect can be rescued by expression of the OCT1 gene from a plasmid vector. Subcellular localization experiments and LC-MS/MS analysis of enriched organellar fraction-proteins confirmed the presence of Oct1p in mitochondria. Phylogenetic profiling of Oct1p revealed an intricate evolutionary pattern indicating multiple horizontal gene transfers among different fungal groups.
The Candida parapsilosis OCT1 gene encodes mitochondrial thiolase catalysing the last step of the 3-oxoadipate pathway.
PDF
