Search Results (23)

Due to its antioxidant and immunomodulatory properties, using passion fruit seed oil (PFSO) is a promising strategy to mitigate the effects of heat stress in laying hens, potentially optimizing the absorption of essential minerals such as selenium (Se) and zinc (Zn). Therefore, this study investigated the profile of selenium- and zinc-binding proteins (Se/Zn-BPs) in the hepatic proteome of Lohmann White laying hens (26 weeks old, n = 96) subjected to heat stress and whose diet was supplemented with 0.9% PFSO, using a metalloproteomic approach that combined two-dimensional electrophoresis (2D PAGE), graphite furnace atomic absorption spectrometry (GFAAS), and liquid chromatography–tandem mass spectrometry (LC-MS/MS). The experimental design was a 2 × 2 factorial (temperature: thermoneutral/stress × diet: control/PFSO) design. After 84 days, liver samples were collected and subjected to metalloproteomic analyses. GFAAS analysis showed higher concentrations of Zn and Se in the protein pellets and in 11 specific protein spots of the supplemented groups (thermoneutral/PFSO and stress/PFSO). LC-MS/MS analysis identified 56 Se/Zn-BPs, with a predominance of heat shock chaperones (HSPs) and proteins involved in energy metabolism. In conclusion, PFSO supplementation modulates Se and Zn absorption, promoting a mineral balance that optimizes immune and antioxidant defense processes. This mechanism can lead to a positive impact on the health and productive performance of laying hens under heat stress. Full article

Background: Primary ciliary dyskinesia (PCD) is a rare genetic disorder characterized by recurrent sinopulmonary infections due to motile cilia defects. The disease is genetically heterogeneous, with abnormalities in structural ciliary proteins. Zinc finger MYND-type containing 10 (ZMYND10) is essential for the assembly of outer dynein arms (ODA), with chaperones like Glucose-regulated protein 78 (GRP78) facilitating protein folding. This study investigates ZMYND10 and Dynein axonemal heavy chain 5 (DNAH5) mutations in individuals with PCD. Methods: Eight individuals aged 14–22 with clinical PCD symptoms and confirmed DNAH5 mutations were included. We analyzed the correlation between DNAH5 abnormalities and preassembly/chaperone proteins using immunofluorescence labeling. Nasal swabs were double-labeled (DNAH5–β-tubulin, β-tubulin–ZMYND10, β-tubulin–GRP78) and examined via fluorescence microscopy. Serum metabolomics and proteomics were also assessed. Results: The corrected total cell fluorescence (CTCF) levels of DNAH5, ZMYND10, and GRP78 were significantly different between PCD individuals and controls. Metabolomic analysis showed reduced valine, leucine, and isoleucine biosynthesis, with increased malate and triacylglycerol biosynthesis, malate-aspartate and glycerol phosphate shuttles, and arginine/proline metabolism, suggesting mitochondrial and ER stress. Conclusions: The altered expression of DNAH5, ZMYND10, and GRP78, along with metabolic shifts, points to a complex link between ciliary dysfunction and cellular stress in PCD. Further studies are needed to clarify the underlying mechanisms. Full article

Viral protein X (Vpx) is a unique accessory protein encoded by the genome of the human immunodeficiency virus type 2 (HIV-2) and lineages of the simian immunodeficiency virus of sooty mangabeys. So far, counteracting the cellular restriction factor SAMHD1 and mediating the efficient translocation of viral pre-integration complex have been recognized as key functions of Vpx; however, a thorough exploration of its effects on the cellular transcriptome and cytokine milieu has not yet been undertaken. In this study, we carried out the transcriptomic analysis of THP-1 cells and determined differential gene expressions induced by HIV-2 Vpx, utilizing vectors coding for the wild-type and K68-R70 functionally restricted proteins. Significantly altered genes were then validated and quantified through real-time quantitative PCR (qPCR); additionally, replication-competent virions were also used to confirm the findings. Moreover, we analyzed the effect of Vpx expression on the secretion of key cytokines in the medium of transfected cells. Our findings revealed that wild-type HIV-2 Vpx can significantly alter the expression of genes coding for helicases, zinc finger proteins, chaperons, transcription factors and proteins involved in DNA methylation. Differentially altered genes were involved in negative regulation of viral processes, the type I interferon-signaling pathway, DNA-template transcription, elongation, the positive regulation of interferon beta production and the negative regulation of innate immune response. Importantly, Vpx was also found to decrease the expression of HIV-1 Tat, possibly through the downregulation of a crucial splicing factor, required for the maturation of Tat. Additionally, studies on cellular cytokine milieu showed that this accessory protein induced key proinflammatory cytokines. Our study provides important information about the complex role played by HIV-2 Vpx in priming and taming the cellular environment to allow for the establishment of the infection. Full article

Background: Tomatoes are self-pollinating plants, and successful fruit set depends on the production of functional pollen within the same flower. Our previous studies have shown that the ‘Black Vernissage’ tomato variety exhibits greater resilience to heat stress in terms of pollen productivity compared to the ‘Micro-Tom’ variety. Pollen productivity is determined by meiotic activity during microsporogenesis and the development of free microspores during gametogenesis. This study focused on identifying heat stress (HS)-induced proteomes in pollen mother cells (PMCs) and microspores. Methods: Tomato plants were grown under two temperature conditions: 26 °C (non-heat-treated control) and 37 °C (heat-treated). Homogeneous cell samples of meiotic PMCs (prior to the tetrad stage) and free microspores were collected using laser capture microdissection (LCM). The heat-induced proteomes were identified using tandem mass tag (TMT)–quantitative proteomics analysis. Results: The enrichment of the meiotic cell cycle in PMCs and the pre-mitotic process in free microspores confirmed the correlation between proteome expression and developmental stage. Under HS, PMCs in both tomato varieties were enriched with heat shock proteins (HSPs). However, the ‘Black Vernissage’ variety exhibited a greater diversity of HSP species and a higher level of enrichment compared to the ‘Micro-Tom’ variety. Additionally, several proteins involved in gene expression and protein translation were downregulated in PMCs and microspores of both varieties. In the PMC proteomes, the relative abundance of proteins showed no significant differences between the two varieties under normal conditions, with very few exceptions. However, HS induced significant differential expression both within and between the varieties. More importantly, these heat-induced differentially abundant proteins (DAPs) in PMCs are directly involved in meiotic cell division, including the meiosis-specific protein ASY3 (Solyc01g079080), the cell division protein kinase 2 (Solyc11g070140), COP9 signalosome complex subunit 1 (Solyc01g091650), the kinetochore protein ndc80 (Solyc01g104570), MORC family CW-type zinc finger 3 (Solyc02g084700), and several HSPs that function in protecting the fidelity of the meiotic processes, including the DNAJ chaperone (Solyc04g009770, Solyc05g055160), chaperone protein htpG (Solyc04g081570), and class I and class II HSPs. In the microspores, most of the HS-induced DAPs were consistently observed across both varieties, with only a few proteins showing significant differences between them under heat stress. These HS-induced DAPs include proteases, antioxidant proteins, and proteins related to cell wall remodeling and the generation of pollen exine. Conclusions: HS induced more dynamic proteomic changes in meiotic PMCs compared to microspores, and the inter-varietal differences in the PMC proteomes align with the effects of HS on pollen productivity observed in the two varieties. This research highlights the importance of the cell-type-specific proteomics approach in identifying the molecular mechanisms that are critical for the pollen developmental process under elevated temperature conditions. Full article

As areas of application of terahertz (THz) radiation expand in science and practice, evidence is accumulating that this type of radiation can affect not only biological molecules directly, but also cellular processes as a whole. In this study, the transcriptome in cells of the thermophilic bacterium Geobacillus icigianus was analyzed immediately after THz irradiation (0.23 W/cm², 130 μm, 15 min) and at 10 min after its completion. THz irradiation does not affect the activity of heat shock protein genes and diminishes the activity of genes whose products are involved in peptidoglycan recycling, participate in redox reactions, and protect DNA and proteins from damage, including genes of chaperone protein ClpB and of DNA repair protein RadA, as well as genes of catalase and kinase McsB. Gene systems responsible for the homeostasis of transition metals (copper, iron, and zinc) proved to be the most sensitive to THz irradiation; downregulation of these systems increased significantly 10 min after the end of the irradiation. It was also hypothesized that some negative effects of THz radiation on metabolism in G. icigianus cells are related to disturbances in activities of gene systems controlled by metal-sensitive transcription factors. Full article

In light of expected climate change, it is important to seek nature-based solutions that can contribute to the protection of our planet as well as to help overcome the emerging adverse changes. In an agricultural context, increasing plant resistance to abiotic stress seems to be crucial. Therefore, the scope of the presented research was focused on the application of botanical extracts that exerted positive effects on model plants growing under controlled laboratory conditions, as well as plants subjected to sorbitol-induced osmotic stress. Foliar spraying increased the length and fresh mass of the shoots (e.g., extracts from Taraxacum officinale, Trifolium pratense, and Pisum sativum) and the roots (e.g., Solidago gigantea, Hypericum perforatum, and Pisum sativum) of cabbage seedlings grown under stressful conditions, as well as their content of photosynthetic pigments (Pisum sativum, Lens culinaris, and Hypericum perforatum) along with total phenolic compounds (Hypericum perforatum, Taraxacum officinale, and Urtica dioica). The antioxidant activity of the shoots measured with the use of DDPH (Pisum sativum, Taraxacum officinale, Urtica dioica, and Hypericum perforatum), ABTS (Trifolium pratense, Symphytum officinale, Valeriana officinalis, Pisum sativum, and Lens culinaris), and FRAP (Symphytum officinale, Valeriana officinalis, Urtica dioica, Hypericum perforatum, and Taraxacum officinale) assays was also enhanced in plants exposed to osmotic stress. Based on these findings, the most promising formulation based on Symphytum officinale was selected and subjected to transcriptomic analysis. The modification of the expression of the following genes was noted: Bol029651 (glutathione S-transferase), Bol027348 (chlorophyll A-B binding protein), Bol015841 (S-adenosylmethionine-dependent methyltransferases), Bol009860 (chlorophyll A-B binding protein), Bol022819 (GDSL lipase/esterase), Bol036512 (heat shock protein 70 family), Bol005916 (DnaJ Chaperone), Bol028754 (pre-mRNA splicing Prp18-interacting factor), Bol009568 (heat shock protein Hsp90 family), Bol039362 (gibberellin regulated protein), Bol007693 (B-box-type zinc finger), Bol034610 (RmlC-like cupin domain superfamily), Bol019811 (myb_SHAQKYF: myb-like DNA-binding domain, SHAQKYF class), Bol028965 (DA1-like Protein). Gene Ontology functional analysis indicated that the application of the extract led to a decrease in the expression of many genes related to the response to stress and photosynthetic systems, which may confirm a reduction in the level of oxidative stress in plants treated with biostimulants. The conducted studies showed that the use of innovative plant-based products exerted positive effects on crops and can be used to supplement current cultivation practices. Full article

Ribosomal proteins are fundamental components of the ribosomes in all living cells. The ribosomal protein uS5 (Rps2) is a stable component of the small ribosomal subunit within all three domains of life. In addition to its interactions with proximal ribosomal proteins and rRNA inside the ribosome, uS5 has a surprisingly complex network of evolutionarily conserved non-ribosome-associated proteins. In this review, we focus on a set of four conserved uS5-associated proteins: the protein arginine methyltransferase 3 (PRMT3), the programmed cell death 2 (PDCD2) and its PDCD2-like (PDCD2L) paralog, and the zinc finger protein, ZNF277. We discuss recent work that presents PDCD2 and homologs as a dedicated uS5 chaperone and PDCD2L as a potential adaptor protein for the nuclear export of pre-40S subunits. Although the functional significance of the PRMT3–uS5 and ZNF277–uS5 interactions remain elusive, we reflect on the potential roles of uS5 arginine methylation by PRMT3 and on data indicating that ZNF277 and PRMT3 compete for uS5 binding. Together, these discussions highlight the complex and conserved regulatory network responsible for monitoring the availability and the folding of uS5 for the formation of 40S ribosomal subunits and/or the role of uS5 in potential extra-ribosomal functions. Full article

The bioavailability of copper (Cu) in human cells may depend on a complex interplay with zinc (Zn) ions. We investigated the ability of the Zn ion to target the human Cu-chaperone Atox1, a small cytosolic protein capable of anchoring Cu(I), by a conserved surface-exposed Cys-X-X-Cys (CXXC) motif, and deliver it to Cu-transporting ATPases in the trans-Golgi network. The crystal structure of Atox1 loaded with Zn displays the metal ion bridging the CXXC motifs of two Atox1 molecules in a homodimer. The identity and location of the Zn ion were confirmed through the anomalous scattering of the metal by collecting X-ray diffraction data near the Zn K-edge. Furthermore, soaking experiments of the Zn-loaded Atox1 crystals with a strong chelating agent, such as EDTA, caused only limited removal of the metal ion from the tetrahedral coordination cage, suggesting a potential role of Atox1 in Zn metabolism and, more generally, that Cu and Zn transport mechanisms could be interlocked in human cells. Full article

Thermus thermophilus trigger factor (TtTF) is a zinc-dependent molecular chaperone whose folding-arrest activity is regulated by Zn²⁺. However, little is known about the mechanism of zinc-dependent regulation of the TtTF activity. Here we exploit in vitro biophysical experiments to investigate zinc-binding, the oligomeric state, the secondary structure, and the thermal stability of TtTF in the absence and presence of Zn²⁺. The data show that full-length TtTF binds Zn²⁺, but the isolated domains and tandem domains of TtTF do not bind to Zn²⁺. Furthermore, circular dichroism (CD) and nuclear magnetic resonance (NMR) spectra suggested that Zn²⁺-binding induces the partial structural changes of TtTF, and size exclusion chromatography-multi-angle light scattering (SEC-MALS) showed that Zn²⁺ promotes TtTF oligomerization. Given the previous work showing that the activity regulation of E. coli trigger factor is accompanied by oligomerization, the data suggest that TtTF exploits zinc ions to induce the structural change coupled with the oligomerization to assemble the client-binding site, thereby effectively preventing proteins from misfolding in the thermal environment. Full article

Low body weight at birth has been shown to be a risk factor for future metabolic disorders, as well as stress response abnormalities and depression. We showed that low-birthweight rats had prolonged high blood corticosterone levels after stress exposure, and that an increase in Gas5 lncRNA, a decoy receptor for glucocorticoid receptors (GRs), reduces glucocorticoid responsiveness. Thus, we concluded that dampened pituitary glucocorticoid responsiveness disturbed the glucocorticoid feedback loop in low-birthweight rats. However, it remains unclear whether such glucocorticoid responsiveness is suppressed solely in the pituitary or systemically. The expression of Gas5 lncRNA increased only in the pituitary, and the intact induction of expression of the GR co-chaperone factor Fkbp5 against dexamethasone was seen in the liver, muscle, and adipose tissue. Intervention with a methyl-modulator diet (folate, VB₁₂, choline, betaine, and zinc) immediately before or one week after delivery reversed the expression level of Gas5 lncRNA in the pituitary of the offspring. Consequently, it partially normalized the blood corticosterone levels after restraint stress exposure. In conclusion, the mode of glucocorticoid response in low-birthweight rats is impaired solely in the pituitary, and intervention with methyl-modulators ameliorates the impairment, but with a narrow therapeutic time window. Full article

Assembly of photosynthetic complexes is sensitive to changing light intensities, drought and pathogens, each of which induces a redox imbalance that requires the assistance of specific chaperones to maintain protein structure. Here we report a thylakoid membrane-associated DnaJ-like protein, ZnJ6 (Cre06.g251716.t1.2), in Chlamydomonas reinhardtii. The protein has four CXXCX(G)X(G) motifs that form two zinc fingers (ZFs). Site-directed mutagenesis (Cys > Ser) eliminates the ability to bind zinc. An intact ZF is required for ZnJ6 stability at elevated temperatures. Chaperone assays with recombinant ZnJ6 indicate that it has holding and oxidative activities. ZnJ6 is unable to reduce the disulfide bonds of insulin but prevents its aggregation in a reducing environment. It also assists in the reactivation of reduced denatured RNaseA, possibly by its oxidizing activity. ZnJ6 pull-down assays revealed interactions with oxidoreductases, photosynthetic proteins and proteases. In vivo experiments with a C. reinhardtii insertional mutant (∆ZnJ6) indicate enhanced tolerance to oxidative stress but increased sensitivity to heat and reducing conditions. Moreover, ∆ZnJ6 has reduced photosynthetic efficiency shown by the Chlorophyll fluorescence transient. Taken together, we identify a role for this thylakoid-associated DnaJ-like oxidizing chaperone that assists in the prevention of protein misfolding and aggregation, thus contributing to stress endurance, redox maintenance and photosynthetic balance. Full article

Many bacteria require ATP binding cassette (ABC) transporters for the import of the essential metal zinc from limited environments. These systems rely on a periplasmic or cell-surface solute binding protein (SBP) to bind zinc with high affinity and specificity. AztABCD is one such zinc transport system recently identified in a large group of diverse bacterial species. In addition to a classical SBP (AztC), the operon also includes a periplasmic metallochaperone (AztD) shown to transfer zinc directly to AztC. Crystal structures of both proteins from Paracoccus denitrificans have been solved and suggest several structural features on each that may be important for zinc binding and transfer. Here we determine zinc binding affinity, dissociation kinetics, and transfer kinetics for several deletion mutants as well as a crystal structure for one of them. The results indicate specific roles for loop structures on AztC and an N-terminal motif on AztD in zinc binding and transfer. These data are consistent with a structural transfer model proposed previously and provide further mechanistic insight into the processes of zinc binding and transfer. Full article

In mammals, adipose tissue is an active secretory tissue that responds to mild hypothermia and as such is a genuine model to study molecular and cellular adaptive responses to cold-stress. A recent study identified a mammal-specific protein of the endoplasmic reticulum that is strongly induced in the inguinal subcutaneous white adipocyte upon exposure to cold, calsyntenin 3β (CLSTN3β). CLSTN3β regulates sympathetic innervation of thermogenic adipocytes and contributes to adaptive non-shivering thermogenesis. The calcium- and zinc-binding S100B is a downstream effector in the CLSTN3β pathways. We review, here, the literature on the transcriptional regulation of the S100b gene in adipocyte cells. We also rationalize the interactions of the S100B protein with its recognized or hypothesized intracellular (p53, ATAD3A, CYP2E1, AHNAK) and extracellular (Receptor for Advanced Glycation End products (RAGE), RPTPσ) target proteins in the context of adipocyte differentiation and adaptive thermogenesis. We highlight a chaperon-associated function for the intracellular S100B and point to functional synergies between the different intracellular S100B target proteins. A model of non-classical S100B secretion involving AHNAK/S100A10/annexin2-dependent exocytosis by the mean of exosomes is also proposed. Implications for related areas of research are noted and suggestions for future research are offered. Full article

The unfolded protein response (UPR) is an adaptive response to intrinsic and external stressors, and it is mainly activated by the accumulation of misfolded proteins at the endoplasmic reticulum (ER) lumen producing ER stress. The UPR signaling network is interconnected with autophagy, the proteolytic machinery specifically devoted to clearing misfolded proteins in order to survive bioenergetic stress and/or induce cell death. Oncosuppressor TP53 may undergo inactivation following missense mutations within the DNA-binding domain (DBD), and mutant p53 (mutp53) proteins may acquire a misfolded conformation, often due to the loss of the DBD-bound zinc ion, leading to accumulation of hyperstable mutp53 proteins that correlates with more aggressive tumors, resistance to therapies, and poorer outcomes. We previously showed that zinc supplementation induces mutp53 protein degradation by autophagy. Here, we show that mutp53 (i.e., Arg273) degradation following zinc supplementation is correlated with activation of ER stress and of the IRE1α/XBPI arm of the UPR. ER stress inhibition with chemical chaperone 4-phenyl butyrate (PBA) impaired mutp53 downregulation, which is similar to IRE1α/XBPI specific inhibition, reducing cancer cell death. Knockdown of mutp53 failed to induce UPR/autophagy activation indicating that the effect of zinc on mutp53 folding was likely the key event occurring in ER stress activation. Recently discovered small molecules targeting components of the UPR show promise as a novel anticancer therapeutic intervention. However, our findings showing UPR activation during mutp53 degradation indicate that caution is necessary in the design of therapies that inhibit UPR components. Full article

Zinc (II) ions (hereafter simplified as zinc) are important for the structural and functional activity of many proteins. For Cu, Zn superoxide dismutase (Sod1), zinc stabilizes the native structure of each Sod1 monomer, promotes homo-dimerization and plays an important role in activity by “softening” the active site so that copper cycling between Cu(I) and Cu(II) can rapidly occur. Previously, we have reported that binding of Sod1 by its copper chaperone (Ccs) stabilizes a conformation of Sod1 that promotes site-specific high-affinity zinc binding. While there are a multitude of Sod1 mutations linked to the familial form of amyotrophic lateral sclerosis (fALS), characterizations by multiple research groups have been unable to realize strong commonalities among mutants. Here, we examine a set of fALS-linked Sod1 mutations that have been well-characterized and are known to possess variation in their biophysical characteristics. The zinc affinities of these mutants are evaluated here for the first time and then compared with the previously established value for wild-type Sod1 zinc affinity. Ccs does not have the same ability to promote zinc binding to these mutants as it does for the wild-type version of Sod1. Our data provides a deeper look into how (non)productive Sod1 maturation by Ccs may link a diverse set of fALS-Sod1 mutations. Full article