Small Heat Shock Protein Research Articles

Enhanced Acetate Tolerance and Recombinant Protein Accumulation in Escherichia coli by Transgenic Expression of a Heat Shock Protein from Carrot (Daucus carota L.).

In Escherichia coli (E. coli) culture, acetate accumulates as an undesirable by-product of aerobic fermentation on glucose and inhibits cell growth and recombinant protein production. We examined whether the heterologous expression of a eukaryotic heat shock protein (Hsp) can confer tolerance to acetate in E. coli. Transgenic cell lines (TCLs) heterologously expressing a small heat shock protein (sHsp) from carrot (Daucus carota L.), DcHsp17.7, were exposed to heat, sodium acetate, and alkaline conditions. The cell growth and cell viability were examined by measuring O.D.600 and colony-forming units (CFU), respectively. The His-tagged recombinant alcohol dehydrogenase (ADH) gene cloned in a pET11a expression vector was introduced into TCL1 and expressed by isopropyl β-D-1-thiogalactopyranoside treatment. After purifying using Ni-NTA affinity chromatography, its accumulation levels were examined using SDS-PAGE in the presence of acetate. TCLs constitutively expressing DcHsp17.7 showed improved growth, cell density, and cell viability under the stress conditions of heat, acetate, and alkaline compared to an empty vector control line. In acetate stress conditions, TCL1 accumulated more cellular proteins (approximately 130%) than the control. The recombinant ADH accumulated to a higher level in TCL1 (2.2-fold at 16 °C) than the control. The addition of acetate reduced the recombinant ADH level by 70% in the control when compared with the absence of acetate. In contrast, recombinant ADH accumulation was not affected by acetate in TCL1. In the presence of acetate, TCL1 accumulated 6.4-fold more recombinant ADH than did the control. Furthermore, recombinant ADH produced in TCL1 showed 1.5-fold higher enzyme activity than that produced in the control in the presence or absence of acetate. Our study showed that heterologously expressed DcHsp17.7 from carrot can alleviate the negative effects of acetate on E. coli.

Ubiquitin profiling of lysophagy identifies actin stabilizer CNN2 as a target of VCP/p97 and uncovers a link to HSPB1

Lysosomal membrane permeabilization (LMP) is an underlying feature of diverse conditions including neurodegeneration. Cells respond by extensive ubiquitylation of membrane-associated proteins for clearance of the organelle through lysophagy that is facilitated by the ubiquitin-directed AAA-ATPase VCP/p97. Here, we assessed the ubiquitylated proteome upon acute LMP and uncovered a large diversity of targets and lysophagy regulators. They include calponin-2 (CNN2) that, along with the Arp2/3 complex, translocates to damaged lysosomes and regulates actin filaments to drive phagophore formation. Importantly, CNN2 needs to be ubiquitylated during the process and removed by VCP/p97 for efficient lysophagy. Moreover, we identified the small heat shock protein HSPB1 that assists VCP/p97 in the extraction of CNN2 and show that other membrane regulators including SNAREs, PICALM, AGFG1, and ARL8B are ubiquitylated during lysophagy. Our data reveal a framework of how ubiquitylation and two effectors, VCP/p97 and HSPB1, cooperate to protect cells from the deleterious effects of LMP.

The Small Heat Shock Protein, HSPB1, Interacts with and Modulates the Physical Structure of Membranes.

Small heat shock proteins (sHSPs) have been demonstrated to interact with lipids and modulate the physical state of membranes across species. Through these interactions, sHSPs contribute to the maintenance of membrane integrity. HSPB1 is a major sHSP in mammals, but its lipid interaction profile has so far been unexplored. In this study, we characterized the interaction between HSPB1 and phospholipids. HSPB1 not only associated with membranes via membrane-forming lipids, but also showed a strong affinity towards highly fluid membranes. It participated in the modulation of the physical properties of the interacting membranes by altering rotational and lateral lipid mobility. In addition, the in vivo expression of HSPB1 greatly affected the phase behavior of the plasma membrane under membrane fluidizing stress conditions. In light of our current findings, we propose a new function for HSPB1 as a membrane chaperone.

The Monomeric α-Crystallin Domain of the Small Heat-shock Proteins αB-crystallin and Hsp27 Binds Amyloid Fibril Ends

Small heat-shock proteins (sHSPs) are ubiquitously expressed molecular chaperones present in all kingdoms of life that inhibit protein misfolding and aggregation. Despite their importance in proteostasis, the structure–function relationships of sHSPs remain elusive. Human sHSPs are characterised by a central, highly conserved α-crystallin domain (ACD) and variable-length N- and C-terminal regions. The ACD forms antiparallel homodimers via an extended β-strand, creating a shared β-sheet at the dimer interface. The N- and C-terminal regions mediate formation of higher order oligomers that are thought to act as storage forms for chaperone-active dimers. We investigated the interactions of the ACD of two human sHSPs, αB-crystallin (αB-C) and Hsp27, with apolipoprotein C-II amyloid fibrils using analytical ultracentrifugation and nuclear magnetic resonance spectroscopy. The ACD was found to interact transiently with amyloid fibrils to inhibit fibril elongation and naturally occurring fibril end-to-end joining. This interaction was sensitive to the concentration of fibril ends indicating a ‘fibril-capping’ interaction. Furthermore, resonances arising from the ACD monomer were attenuated to a greater extent than those of the ACD dimer in the presence of fibrils, suggesting that the monomer may bind fibrils. This hypothesis was supported by mutagenesis studies in which disulfide cross-linked ACD dimers formed by both αB-C and Hsp27 were less effective at inhibiting amyloid fibril elongation and fibril end-to-end joining than ACD constructs lacking disulfide cross-linking. Our results indicate that sHSP monomers inhibit amyloid fibril elongation, highlighting the importance of the dynamic oligomeric nature of sHSPs for client binding.

Cloning and expression of three small heat shock proteins in Cherax quadricarinatus under Vibrio alginolyticus or Aeromonas hydrophila stress

Cherax quadricarinatus has been widely introduced and cultured around the world and has been seriously affected by pathogens, including viruses and bacteria. Small heat shock proteins (sHsps) have been shown to have a positive effect on immune responses in animals. In this study, three subunits of sHsps (CqsHsps, named CqHsp24.06, CqHsp41.97 and CqHsp40.17) from C. quadricarinatus were cloned. The coding sequence lengths of the CqHsp24.06, CqHsp41.97 and CqHsp40.17 genes were 654 bp, 1128 bp and 1074 bp, with a molecular weight of 24.06, 41.97 and 40.17 kDa, respectively. In normal tissues, the expression level of CqHsp24.06 was the highest in the intestinal tract and was 12 times higher than that in the hepatopancreas, while CqHsp41.97 was mostly expressed in the gills which was 8 times higher than that in the nerves. However, CqHsp40.17 was highest in the nerves and was 12 times higher than its expression level in the gonads. In gills of C. quadricarinatus stressed by Vibrio Alginolyticus, the relative expression levels of CqHsp24.06, CqHsp41.97 and CqHsp40.17 in gill first increased and then decreased, and the maximum expression increased by 25 times. Under Aeromonas hydrophila stress, the expression of CqHsp24.06 and CqHsp41.97 reached the highest level at the 3-h time point and at the 12-h time point for CqHsp40.17, and the maximum expression increased by 3.5 times. These results indicated that CqsHsps could induce the activation of the immune system, and they were more sensitive to Vibrio Alginolyticus than to Aeromonas hydrophila.

Differential Roles of Three α-Crystallin Domain-Containing sHsps of Beauveria bassiana in Asexual Development, Multiple Stress Tolerance and Virulence.

Small heat shock proteins (sHsps) containing conserved α-crystallin domain play important roles in many cellular processes, but little is known about the functions of sHsps in filamentous entomopathogens. Here, three sHsps of Hsp20, Hsp30a, and Hsp30b were characterized in Beauveria bassiana, a filamentous fungal insect pathogen that serves as the main source of wide-spectrum fungal insecticides. The results demonstrated that these three genes are interrelated at the transcriptional level under normal and heat-shocked conditions. Meanwhile, all the deletion mutants showed significant but differential changes in cell wall integrity, antioxidant activity, hyphal tolerance to carbendazim fungicide, conidial tolerance to 45 °C wet heat and virulence. However, only Δhsp30b showed growth defects on rich and minimal media at 25 °C and Δhsp30a displayed the reduction in conidiophores and conidia. Moreover, the single deletion of hsp30a and hsp30b caused the decreases in hyphal growth at 34 °C and conidial tolerance to UV-B irradiation. Our findings provide a global insight into vital roles of hsp20, hsp30a, and hsp30b in asexual development, environmental adaptation, and fungal virulence of B. bassiana.

Abstract 865: The small heat shock protein αB-crystallin protects versus withaferin A-induced apoptosis and confers a more metastatic phenotype in cisplatin-resistant ovarian cancer cells

Abstract A majority of ovarian tumors eventually recur in a drug-resistant form leaving patients few treatment options. The goal of this study was to explore the phenotypic and molecular characteristics of a cisplatin-resistant ovarian cancer cell line (OVCAR8R) as compared to its cisplatin-sensitive syngeneic counterpart (OVCAR8) and to explore the effectiveness of a novel therapeutic, Withaferin A (WA). In addition to unique morphological characteristics and increased expression of vimentin, the small heat shock proteins (Hsps) αB-Crystallin and Hsp27 are constitutively expressed in OVCAR8R cells while OVCAR8 cells do not express these Hsps, supporting that Hsp overexpression may confer resistance to chemotherapy and promote a more aggressive tumor type. WA treatment causes apoptosis in a dose-dependent manner in OVCAR8 cells, while the OVCAR8R cells remain viable at comparable doses of WA. αB-Crystallin was upregulated in a dose dependent manner in the WA-treated OVCAR8R cells. To determine the significance of αB-crystallin expression in conferring a more aggressive phenotype, αB-crystallin was silenced by CRISPR-Cas9 in OVCAR8R cells. Strikingly, the morphology of the OVCAR8R-αB-crystallin-knockout clones revert to the morphology of the cisplatin-sensitive OVCAR8 cell line. Molecular characterization reveals concurrent downregulation of Hsp27, and decreased expression of vimentin supporting that loss of αB-Crystallin is coincident with a less aggressive molecular phenotype. Upon challenge with WA, vimentin is downregulated in the αB-crystallin-null clones and in the cisplatin-sensitive OVCAR8 cells, but not in the OVCAR8R cells. Transient transfection of wildtype αB-Crystallin, but not a chaperone-defective mutant, alters the morphology of the OVCAR8 cells where the αB-Crystallin-knockout clones closely resemble the cisplatin-resistant OVCAR8R cells. Further, transient overexpression of wildtype αB-crystallin, but not a chaperone-defective-mutant, protects cells from WA-induced apoptosis as compared to GFP-transfected cells. Migration assays reveal that OVCAR8R cells have increased migration capability as compared to their cisplatin-sensitive counterparts and silencing of αB-Crystallin decreases migration of cisplatin-resistant cells. This research supports the potential effectiveness of WA as a therapy for ovarian cancer cells that acquire resistance to platinum-based therapies and underscores that αB-crystallin confers a more aggressive cellular phenotype and therefore, may be a promising molecular target for a better clinical outcome. Citation Format: Merideth K. Krevosky, Melissa M. Carmichael. The small heat shock protein αB-crystallin protects versus withaferin A-induced apoptosis and confers a more metastatic phenotype in cisplatin-resistant ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 865.

Heat shock factor HSFA2 fine-tunes resetting of thermomemory via plastidic metalloprotease FtsH6.

Plants 'memorize' stressful events and protect themselves from future, often more severe, stresses. To maximize growth after stress, plants 'reset' or 'forget' memories of stressful situations, which requires an intricate balance between stress memory formation and the degree of forgetfulness. HEAT SHOCK PROTEIN 21 (HSP21) encodes a small heat shock protein in plastids of Arabidopsis thaliana. HSP21 functions as a key component of thermomemory, which requires a sustained elevated level of HSP21 during recovery from heat stress. A heat-induced metalloprotease, filamentation temperature-sensitive H6 (FtsH6), degrades HSP21 to its pre-stress abundance, thereby resetting memory during the recovery phase. The transcription factor heat shock factor A2 (HSFA2) activates downstream genes essential for mounting thermomemory, acting as a positive regulator in the process. Here, using a yeast one-hybrid screen, we identify HSFA2 as an upstream transactivator of the resetting element FtsH6. Constitutive and inducible overexpression of HSFA2 increases expression of FtsH6, whereas it is drastically reduced in the hsfa2 knockout mutant. Chromatin immunoprecipitation reveals in planta binding of HSFA2 to the FtsH6 promoter. Importantly, overexpression of HSFA2 improves thermomemory more profoundly in ftsh6 than wild-type plants. Thus, by activating both memory-supporting and memory-resetting genes, HSFA2 acts as a cellular homeostasis factor during thermomemory.

Evolution of the DEHYDRATION-RESPONSIVE ELEMENT-BINDING PROTEIN subfamily in green plants.

Adapting to unfavorable environments is a necessary step in plant terrestrialization and radiation. The dehydration-responsive element-binding (DREB) protein subfamily plays a pivotal role in plant abiotic stress regulation. However, relationships between the origin and expansion of the DREB subfamily and adaptive evolution of land plants are still being elucidated. Here, we constructed the evolutionary history of the DREB subfamily by compiling APETALA2/ethylene-responsive element-binding protein superfamily genes from 169 representative species of green plants. Through extensive phylogenetic analyses and comparative genomic analysis, our results revealed that the DREB subfamily diverged from the ethylene-responsive factor (ERF) subfamily in the common ancestor of Zygnemophyceae and Embryophyta during the colonization of land by plants, followed by expansions to form three different ancient archetypal genes in Zygnemophyceae species, designated as groups archetype-I, archetype-II/III, and archetype-IV. Four large-scale expansions paralleling the evolution of land plants led to the nine-subgroup divergence of group archetype-II/III in angiosperms, and five whole-genome duplications during Brassicaceae and Poaceae radiation shaped the diversity of subgroup IIb-1. We identified a Poaceae-specific gene in subgroup IIb-1, ERF014, remaining in a Poaceae-specific microsynteny block and co-evolving with a small heat shock protein cluster. Expression analyses demonstrated that heat acclimation may have driven the neofunctionalization of ERF014s in Pooideae by engaging in the conserved heat-responsive module in Poaceae. This study provides insights into lineage-specific expansion and neofunctionalization in the DREB subfamily, together with evolutionary information valuable for future functional studies of plant stress biology.

Human HspB1, HspB3, HspB5 and HspB8: Shaping these disease factors during vertebrate evolution

Small heat shock proteins (sHSPs) emerged early in evolution and occur in all domains of life and nearly in all species, including humans. Mutations in four sHSPs (HspB1, HspB3, HspB5, HspB8) are associated with neuromuscular disorders. The aim of this study is to investigate the evolutionary forces shaping these sHSPs during vertebrate evolution. We performed comparative evolutionary analyses on a set of orthologous sHSP sequences, based on the ratio of non-synonymous: synonymous substitution rates for each codon. We found that these sHSPs had been historically exposed to different degrees of purifying selection, decreasing in this order: HspB8 > HspB1, HspB5 > HspB3. Within each sHSP, regions with different degrees of purifying selection can be discerned, resulting in characteristic selective pressure profiles. The conserved α-crystallin domains were exposed to the most stringent purifying selection compared to the flanking regions, supporting a 'dimorphic pattern' of evolution. Thus, during vertebrate evolution the different sequence partitions were exposed to different and measurable degrees of selective pressures. Among the disease-associated mutations, most are missense mutations primarily in HspB1 and to a lesser extent in the other sHSPs. Our data provide an explanation for this disparate incidence. Contrary to the expectation, most missense mutations cause dominant disease phenotypes. Theoretical considerations support a connection between the historic exposure of these sHSP genes to a high degree of purifying selection and the unusual prevalence of genetic dominance of the associated disease phenotypes. Our study puts the genetics of inheritable sHSP-borne diseases into the context of vertebrate evolution.

Bacteria binding and antibacterial activities of three small heat shock protein from the swimming crab, Portunus trituberculatus

Small heat shock proteins (sHSPs), a group of molecular chaperones with small molecular mass, are involved in immune responses in many organisms. However, the actual roles of sHSPs in immune defence against bacteria remain largely unclear. In this study, three sHSPs, named PtHSP10, PtHSP20 and PtHSP40 were identified from Portunus trituberculatus, and their immune function was investigated. Sequence analysis showed that the deduced amino acid sequences of PtHSP10, PtHSP20 and PtHSP40 exhibited conserved structures and the functional features. The tissue distributions and transcriptional profiles of these sHSPs in response to Vibrio alginolyticus challenge were analysed by quantitative real-time PCR (qRT-PCR). The results showed that the genes of the three sHSPs were ubiquitously expressed in various tissues, and mainly expressed in hepatopancreas or muscle. The transcriptions of these sHSPs in hepatopancreas of P. trituberculatus injected with V. alginolyticus were upregulated rapidly at 3 h post injection. Moreover, bacterial binding assay showed that the three sHSPs displayed distinct binding activities towards Gram-negative bacteria (V. alginolyticus and V. parahaemolyticus) and Gram-positive bacteria (Staphylococcus aureus). PtHSP10 could bind to V. parahaemolyticus and S. aureus, while PtHSP40 exhibited binding activity towards V. alginolyticus and V. parahaemolyticus. No binding was observed between PtHSP20 and the three tested bacteria. Moreover, PtHSP40 showed antibacterial activity against V. alginolyticus. Our findings indicated that these sHSPs displayed different immune function in immune defence.

Soybean F-Box-Like Protein GmFBL144 Interacts With Small Heat Shock Protein and Negatively Regulates Plant Drought Stress Tolerance

The F-box gene family is one of the largest gene families in plants. These genes regulate plant growth and development, as well as biotic and abiotic stress responses, and they have been extensively researched. Drought stress is one of the major factors limiting the yield and quality of soybean. In this study, bioinformatics analysis of the soybean F-box gene family was performed, and the role of soybean F-box-like gene GmFBL144 in drought stress adaptation was characterized. We identified 507 F-box genes in the soybean genome database, which were classified into 11 subfamilies. The expression profiles showed that GmFBL144 was highly expressed in plant roots. Overexpression of GmFBL144 increased the sensitivity of transgenic Arabidopsis to drought stress. Under drought stress, the hydrogen peroxide (H2O2) and malonaldehyde (MDA) contents of transgenic Arabidopsis were higher than those of the wild type (WT) and empty vector control, and the chlorophyll content was lower than that of the control. Y2H and bimolecular fluorescence complementation (BiFC) assays showed that GmFBL144 can interact with GmsHSP. Furthermore, our results showed that GmFBL144 can form SCFFBL144 (E3 ubiquitin ligase) with GmSkp1 and GmCullin1. Altogether, these results indicate that the soybean F-box-like protein GmFBL144 may negatively regulate plant drought stress tolerance by interacting with sHSP. These findings provide a basis for molecular genetics and breeding of soybean.

Light Induces Carotenoid Biosynthesis-Related Gene Expression, Accumulation of Pigment Content, and Expression of the Small Heat Shock Protein in Apple Fruit.

The coloration of the apple fruit (Malus × domestica Borkh.) depends on pigment content. Light stimulus activates a broad range of photosynthesis-related genes, including carotenoids. The effect of light on two red commercial apple cultivars, ‘Summer Prince’ and ‘Arisoo’ at the juvenile stage were examined. Apple fruits were either bagged to reduce light irradiation or were exposed to direct, enhanced sunlight (reflected). The pigment content and the expression of carotenoid metabolism genes in the peel and flesh of apple fruits were significantly different between the shaded and the reflected parts. These parameters were also different in the two cultivars, highlighting the contribution of the genetic background. Further, a combination of light and transient overexpression of carotenogenic genes increased fruit coloration and pigment content in the variety ‘RubyS’. Western blot analysis showed the expression of small heat shock proteins (smHSP) in lysates extracted from the reflected part of the fruits but not in the bagged fruits, indicating the activation of smHSP in response to heat generated by the reflected light. Therefore, the synergy between the genes and the environment dictates the color of apple fruits.

The mechanism for thermal-enhanced chaperone-like activity of α-crystallin against UV irradiation-induced aggregation of γD-crystallin

Exposure to solar UV irradiation damages γ-crystallin, leading to cataract formation via aggregation. α-Crystallin, as a small heat shock protein, efficiently suppresses this irreversible aggregation by selectively binding the denatured γ-crystallin monomer. In this study, liquid chromatography tandem mass spectrometry was used to evaluate UV-325 nm irradiation-induced photodamage of human γD-crystallin in the presence of bovine α-crystallin, atomic force microscope (AFM) and dynamic light scattering (DLS) techniques were used to detect the quaternary structure changes of the α-crystallin oligomer, and Fourier transform infrared spectroscopy and temperature-jump nanosecond time-resolved IR absorbance difference spectroscopy were used to probe the secondary structure changes of bovine α-crystallin. We find that the thermal-induced subunit dissociation of the α-crystallin oligomer involves the breaking of hydrogen bonds at the dimeric interface, leading to three different spectral components at varied temperature regions as resolved from temperature-dependent IR spectra. Under UV-325 nm irradiation, unfolded γD-crystallin binds to the dissociated α-crystallin subunit to form an αγ-complex, then follows the reassociation of the αγ-complex to the partially dissociated α-crystallin oligomer. This prevents the aggregation of denatured γD-crystallin. The formation of the γD-bound α-crystallin oligomer is further confirmed by AFM and DLS analysis, which reveals an obvious size expansion in the reassociated αγ-oligomers. In addition, UV-325 nm irradiation causes a peptide bond cleavage of γD-crystallin at Ala158 in the presence of α-crystallin. Our results suggest a very effective protection mechanism for subunits dissociated from α-crystallin oligomers against UV irradiation-induced aggregation of γD-crystallin, at the expense of a loss of a short C-terminal peptide in γD-crystallin.

Deciphering the Molecular Mechanisms of Insecticide Resistance From the Transcriptome Data of Field Evolved Spinosad Resistant and Susceptible Populations of Plutella xylostella (Lepidoptera: Plutellidae).

Diamondback moth, Plutella xylostella is a serious pest of cruciferous vegetables and causes substantial economic loss all over the world. This study was undertaken to decipher the molecular mechanisms involved in the field evolved insecticide resistance in P. xylostella upon exposure to spinosad. To do so, spinosad-resistant and susceptible larval populations were subjected to transcriptome analysis using Illumina paired-end sequencing. De novo assembly was generated from raw reads of both the samples which resulted in the identification of 41,205 unigenes. Functional annotation and digital gene expression analysis were carried out to determine the differentially expressed genes. 1,348 unigenes were found to have a significant differential expression in the resistant population. Several genes involved in insecticide resistance like CYP P450, GSTs, small heat shock protein, and UDP glycosyltransferase were found to be up-regulated while genes related to mitochondrial energy metabolism and cuticular processes were down-regulated. Further, gene mining and phylogenetic analysis of two important gene families namely, CYP and GSTs were performed and the results revealed that these genes could play a major role in the development of field evolved spinosad resistance in P. xylostella by gene duplication and differential gene expression.

Minimal Yet Powerful: The Role of Archaeal Small Heat Shock Proteins in Maintaining Protein Homeostasis.

Small heat shock proteins (sHsp) are a ubiquitous group of ATP-independent chaperones found in all three domains of life. Although sHsps in bacteria and eukaryotes have been studied extensively, little information was available on their archaeal homologs until recently. Interestingly, archaeal heat shock machinery is strikingly simplified, offering a minimal repertoire of heat shock proteins to mitigate heat stress. sHsps play a crucial role in preventing protein aggregation and holding unfolded protein substrates in a folding-competent form. Besides protein aggregation protection, archaeal sHsps have been shown recently to stabilize membranes and contribute to transferring captured substrate proteins to chaperonin for refolding. Furthermore, recent studies on archaeal sHsps have shown that environment-induced oligomeric plasticity plays a crucial role in maintaining their functional form. Despite being prokaryotes, the archaeal heat shock protein repository shares several features with its highly sophisticated eukaryotic counterpart. The minimal nature of the archaeal heat shock protein repository offers ample scope to explore the function and regulation of heat shock protein(s) to shed light on their evolution. Moreover, similar structural dynamics of archaeal and human sHsps have made the former an excellent system to study different chaperonopathies since archaeal sHsps are more stable under in vitro experiments.

Identification, expression analysis and functional verification of two genes encoding small heat shock proteins in the western flower thrips, Frankliniella occidentalis (Pergande)

Small heat shock proteins (sHSPs) help prevent the irreversible aggregation of denatured proteins that occurs in response to organismal stress. In this study, we identified two intron-free genes encoding sHSPs from Frankliniella occidentalis; these were designated FoHSP11.6 and FoHSP28.0 and belonged to an atypical and typical sHSP family, respectively. Both FoHSPs were transcribed in all developmental stages of F. occidentalis with the highest expression levels in pupae and adults and greater expression in males than females. Although the FoHSPs had different temperature-induced expression profiles, they were generally induced by both low and high temperatures and reached maximal expression levels after 0.5–1 h of temperature stress. The FoHSPs expression levels in pupae were induced by drought and high humidity, and higher expression levels were correlated with lower survival rates. The thermotolerance of F. occidentalis decreased when theFoHSPs were silenced by RNA interference. Our results show that FoHSP11.6 and FoHSP28.0 are involved in the response to temperature and drought and may also function in growth and development of F. occidentalis.

Spatiotemporal bio-shielding of bacteria through consolidated geometrical structuring

The probiotic bacterium Lactobacillus plantarum is often reckoned as a ‘generalist’ for its ability to adapt and survive in diverse ecological niches. The genomic signatures of L. plantarum have shown its intricate evolutionary ancestry and dynamic lifestyles. Here, we report on a unique geometrical arrangement of the multicellular population of L. plantarum cells. Prominently, a phenomenon of the cone-shaped colony formation and V-shaped cell chaining are discovered in response to the acidic-pH environment. Moreover, subsequent cold stress response triggers an unusual cellular arrangement of consolidated bundles, which appeared to be independently governed by a small heat shock protein (HSP 1). We further report that the V-shaped L. plantarum chaining demonstrates potent antagonistic activity against Candida albicans, a pathogenic yeast, both in vitro and in a Caenorhabditis elegans co-infection model. Finally, we deduce that the multifaceted traits manifested by this probiotic bacterium is an outcome of its dynamic flexibility and cellular heterogeneity.

Oligodendroglia-derived extracellular vesicles activate autophagy via LC3B/BAG3 to protect against oxidative stress with an enhanced effect for HSPB8 enriched vesicles.

BackgroundThe contribution of native or modified oligodendroglia-derived extracellular vesicles (OL-EVs) in controlling chronic inflammation is poorly understood. In activated microglia, OL-EVs contribute to the removal of cytotoxic proteins following a proteotoxic stress. Intracellular small heat shock protein B8 (HSPB8) sustain this function by facilitating autophagy and protecting cells against oxidative stress mediated cell death. Therefore, secretion of HSPB8 in OL-EVs could be beneficial for neurons during chronic inflammation. However, how secreted HSPB8 contribute to cellular proteostasis remains to be elucidated.MethodsWe produced oligodendroglia-derived EVs, either native (OL-EVs) or HSPB8 modified (OL-HSPB8-EVs), to investigate their effects in controlling chronic inflammation and cellular homeostasis. We analyzed the impact of both EV subsets on either a resting or activated microglial cell line and on primary mixed neural cell culture cells. Cells were activated by stimulating with either tumor necrosis factor-alpha and interleukin 1-beta or with phorbol-12-myristate-13-acetate.ResultsWe show that OL-EVs and modified OL-HSPB8-EVs are internalized by C20 microglia and by primary mixed neural cells. The cellular uptake of OL-HSPB8-EVs increases the endogenous HSPB8 mRNA expression. Consistently, our results revealed that both EV subsets maintained cellular homeostasis during chronic inflammation with an increase in the formation of autophagic vesicles. Both EV subsets conveyed LC3B-II and BAG3 autophagy markers with an enhanced effect observed for OL-HSPB8-EVs. Moreover, stimulation with either native or modified OL-HSPB8-EVs showed a significant reduction in ubiquitinated protein, reactive oxygen species and mitochondrial depolarization, with OL-HSPB8-EVs exhibiting a more protective effect. Both EV subsets did not induce cell death in the C20 microglia cell line or the primary mixed neural cultures.ConclusionWe demonstrate that the functions of oligodendroglia secreted EVs enriched with HSPB8 have a supportive role, comparable to the native OL-EVs. Further development of engineered oligodendroglia derived EVs could be a novel therapeutic strategy in countering chronic inflammation. Video Supplementary InformationThe online version contains supplementary material available at 10.1186/s12964-022-00863-x.

Fungal communities represent the majority of root-specific transcripts in the transcriptomes of Agave plants grown in semiarid regions.

Agave plants present drought resistance mechanisms, commercial applications, and potential for bioenergy production. Currently, Agave species are used to produce alcoholic beverages and sisal fibers in semi-arid regions, mainly in Mexico and Brazil. Because of their high productivities, low lignin content, and high shoot-to-root ratio, agaves show potential as biomass feedstock to bioenergy production in marginal areas. Plants host many microorganisms and understanding their metabolism can inform biotechnological purposes. Here, we identify and characterize fungal transcripts found in three fiber-producing agave cultivars (Agave fourcroydes, A. sisalana, and hybrid 11648). We used leaf, stem, and root samples collected from the agave germplasm bank located in the state of Paraiba, in the Brazilian semiarid region, which has faced irregular precipitation periods. We used data from a de novo assembled transcriptome assembly (all tissues together). Regardless of the cultivar, around 10% of the transcripts mapped to fungi. Surprisingly, most root-specific transcripts were fungal (58%); of these around 64% were identified as Ascomycota and 28% as Basidiomycota in the three communities. Transcripts that code for heat shock proteins (HSPs) and enzymes involved in transport across the membrane in Ascomycota and Basidiomycota, abounded in libraries generated from the three cultivars. Indeed, among the most expressed transcripts, many were annotated as HSPs, which appear involved in abiotic stress resistance. Most HSPs expressed by Ascomycota are small HSPs, highly related to dealing with temperature stresses. Also, some KEGG pathways suggest interaction with the roots, related to transport to outside the cell, such as exosome (present in the three Ascomycota communities) and membrane trafficking, which were further investigated. We also found chitinases among secreted CAZymes, that can be related to pathogen control. We anticipate that our results can provide a starting point to the study of the potential uses of agaves’ fungi as biotechnological tools.