Biology subjects

Zgoda, V.

Publications and source records attributed to Zgoda, V..

4 recordsLinked to original sources

bioRxiv2023

Interference Requirements of Type III CRISPR-Cas Systems from Thermus thermophilus

Among the diverse prokaryotic adaptive immunity mechanisms, the Type III CRISPR-Cas systems are the most complex. The multisubunit Type III effectors recognize RNA targets complementary to CRISPR RNAs (crRNAs). Target recognition causes synthesis of cyclic oligoadenylates that activate downstream auxiliary effectors, which affect cell physiology in complex and poorly understood ways. Here, we studied the ability of III-A and III-B CRISPR-Cas subtypes from Thermus thermophilus to interfere with plasmid transformation. We find that for both systems, requirements for crRNA-target complementarity sufficient for interference depend on the target transcript abundance, with more abundant targets requiring shorter complementarity segments. This result and thermodynamic calculations indicate that Type III effectors bind their targets in a simple bimolecular reaction with more extensive crRNA-target base pairing compensating for lower target abundance. Since the targeted RNA used in our work is non-essential for either the host or the plasmid, the results also establish that a certain number of target-bound effector complexes must be present in the cell to interfere with plasmid establishment. For the more active III-A system, we determine the minimal length of RNA-duplex sufficient for interference and show that the position of this minimal duplex can vary within the effector. Finally, we show that the III-A immunity is dependent on the HD nuclease domain of the Cas10 subunit. Since this domain is absent from the III-B system the result implies that the T. thermophilus III-B system must elicit a more efficient cyclic oligoadenylate dependent response to provide the immunity.

microbiology↗

bioRxiv2021

Temperature-induced reorganisation of Schistocephalus solidus (Cestoda) proteome during the transition to the warm-blooded host

The protein composition (proteome) of cestode Schistocephalus solidus was measured in an experiment simulating the transition of the parasite from a cold-blooded to a warm-blooded host. Infective S. solidus plerocercoids obtained from the three-spined stickleback Gasterosteus aculeatus were heated at 40 {degrees}C for 1 hour or cultured in vitro at 40 {degrees}C and 22 {degrees}C for 48 hours. In the short-term experiment, the content of only one tegument protein decreased after heating. After long-term heating, which triggered parasite sexual maturation, an increase in the content of ribosomal proteins, translation initiation factors and enzymes of the amino acid biosynthesis pathway was observed. The synthesis of certain gene products for carbohydrate metabolism, including glycolysis/gluconeogenesis, was found to be regulated in the parasite by temperature. Summary statementThe study focuses on the processes that determine the survival of parasites in warm-blooded animals, and thus demonstrates the potential for the application of proteomics in veterinary medicine.

biochemistry↗

bioRxiv2020

ROS in tobacco stigma exudate affect pollen proteome and provoke membrane hyperpolarization

1.ROS are known to be accumulated in stigmas of different species and can possibly perform different functions in plant reproduction. Here we confirm the assumption that they affect pollen by altering ion transport through the plasma membrane; as a more deferred effect, pollen proteome is modified. We detected ROS in stigma exudate, found hyperpolarization in exudate-treated growing pollen tubes and used flow cytometry of pollen protoplasts to compare the effects of fresh exudate and exogenous H2O2 on pollen tube plasmalemma. Exudate causes plasmalemma hyperpolarization similar to the one provoked by H2O2, which is abolished by catalase treatment and ROS quencher MnTMPP. Inhibitory analysis indicates the participation of Ca2+- and K+-conducting channels in the observed hyperpolarization, linking obtained data with previous patch-clamp studies in vitro. For a deeper understanding of pollen response to ROS we analyzed proteome alterations in H2O2-treated pollen grains. We found 50 unique proteins and 20 differently accumulated proteins that are mainly involved in cell metabolism, energetics, protein synthesis and folding. Thus, pollen is getting ready for effective resource usage, construction of cellular components and rapid growth. HighlightsO_LIThe active substance in stigma exudate is H2O2 C_LIO_LIH2O2 causes hyperpolarization mediated by the activation of cation channels. C_LIO_LIH2O2 affects pollen proteome; we found 50 unique proteins. C_LI

plant biology↗

bioRxiv2017

Distinct types of short open reading frames are translated in plant cells

Genomes contain millions of short (<100 codons) open reading frames (sORFs), which are usually dismissed during gene annotation. Nevertheless, peptides encoded by such sORFs can play important biological roles, and their impact on cellular processes has long been underestimated. Here, we analyzed approximately 70,000 transcribed sORFs in the model plant Physcomitrella patens (moss). Several distinct classes of sORFs that differ in terms of their position on transcripts and the level of evolutionary conservation are present in the moss genome. Over 5000 sORFs were conserved in at least one of ten plant species examined. Mass spectrometry analysis of proteomic and peptidomic datasets suggested that 584 sORFs located on distinct parts of mRNAs and long non-coding RNAs (lncRNAs) are translated, including 73 conservative sORFs. Translational analysis of the sORFs and main ORFs at a single locus suggested the existence of genes that code for multiple proteins and peptides with tissue-specific expression. Alternative splicing is likely involved in the excision of translatable sORFs from such transcripts. We identified a group of sORFs homologous to known protein domains and suggested they function as small interfering peptides. Functional analysis of candidate lncRNA-encoded peptides showed it to be involved in regulating growth and differentiation in moss. The high evolutionary rate and wide translation of sORFs suggest that they may provide a reservoir of potentially active peptides and their importance as a raw material for gene evolution. Our results thus open new avenues for discovering novel, biologically active peptides in the plant kingdom.

plant biology↗