Comparative Analysis of Acute Kidney Injury Models and Related Fibrogenic Responses: Convergence on Methylation Patterns Regulated by Cold Shock Protein.

BACKGROUND: Fibrosis is characterized by excessive extracellular matrix formation in solid organs, disrupting tissue architecture and function. The Y-box binding protein-1 (YB-1) regulates fibrosis-related genes (e.g., Col1a1, Mmp2, and Tgfß1) and contributes significantly to disease progression. This study aims to identify fibrogenic signatures and the underlying signaling pathways modulated by YB-1. METHODS: Transcriptomic changes associated with matrix gene patterns in human chronic kidney diseases and murine acute injury models were analyzed with a focus on known YB-1 targets. Ybx1-knockout mouse strains (Ybx1ΔRosaERT+TX and Ybx1ΔLysM) were subjected to various kidney injury models. Fibrosis patterns were characterized by histopathological staining, transcriptome analysis, qRT-PCR, methylation analysis, zymography, and Western blotting. RESULTS: Integrative transcriptomic analyses revealed that YB-1 is involved in several fibrogenic signatures related to the matrisome, the WNT, YAP/TAZ, and TGFß pathways, and regulates Klotho expression. Changes in the methylation status of the Klotho promoter by specific methyltransferases (DNMT) are linked to YB-1 expression, extending to other fibrogenic genes. Notably, kidney-resident cells play a significant role in YB-1-modulated fibrogenic signaling, whereas infiltrating myeloid immune cells have a minimal impact. CONCLUSIONS: YB-1 emerges as a master regulator of fibrogenesis, guiding DNMT1 to fibrosis-related genes. This highlights YB-1 as a potential target for epigenetic therapies interfering in this process.

Y-Box Binding Protein 1: Unraveling the Multifaceted Role in Cancer Development and Therapeutic Potential.

Y-box binding protein 1 (YBX1), a member of the Cold Shock Domain protein family, is overexpressed in various human cancers and is recognized as an oncogenic gene associated with poor prognosis. YBX1's functional diversity arises from its capacity to interact with a broad range of DNA and RNA molecules, implicating its involvement in diverse cellular processes. Independent investigations have unveiled specific facets of YBX1's contribution to cancer development. This comprehensive review elucidates YBX1's multifaceted role in cancer across cancer hallmarks, both in cancer cell itself and the tumor microenvironment. Based on this, we proposed YBX1 as a potential target for cancer treatment. Notably, ongoing clinical trials addressing YBX1 as a target in breast cancer and lung cancer have showcased its promise for cancer therapy. The ramp up in in vitro research on targeting YBX1 compounds also underscores its growing appeal. Moreover, the emerging role of YBX1 as a neural input is also proposed where the high level of YBX1 was strongly associated with nerve cancer and neurodegenerative diseases. This review also summarized the up-to-date advanced research on the involvement of YBX1 in pancreatic cancer.

Cold-shock proteins accumulate in centrosomes and their expression and primary cilium morphology are regulated by hypothermia and shear stress.

Primary cilia act as cellular sensors for multiple extracellular stimuli and regulate many intracellular signaling pathways in response. Here we investigate whether the cold-shock proteins (CSPs), CIRP and RBM3, are present in the primary cilia and the physiological consequences of such a relationship. R28, an immortalized retinal precursor cell line, was stained with antibodies against CIRP, RBM3, and ciliary markers. Both CSPs were found in intimate contact with the basal body of the cilium during all stages of the cell cycle, including migrating with the centrosome during mitosis. In addition, the morphological and physiological manifestations of exposing the cells to hypothermia and shear stress were investigated. Exposure to moderately cold (32°C) temperatures, the hypothermia mimetic small molecule zr17-2, or to shear stress resulted in a significant reduction in the number and length of primary cilia. In addition, shear stress induced expression of CIRP and RBM3 in a complex pattern depending on the specific protein, flow intensity, and type of flow (laminar versus oscillatory). Flow-mediated CSP overexpression was detected by qRT-PCR and confirmed by Western blot, at least for CIRP. Furthermore, analysis of public RNA Seq databases on flow experiments confirmed an increase of CIRP and RBM3 expression following exposure to shear stress in renal cell lines. In conclusion, we found that CSPs are integral components of the centrosome and that they participate in cold and shear stress sensing.

Structural and biochemical insights into the bacteriophage PlyGRCS endolysin targeting methicillin-resistant Staphylococcus aureus (MRSA) and serendipitous discovery of its interaction with a cold shock protein C (CspC).

Methicillin-resistant Staphylococcus aureus (MRSA) causes life-threatening human infections. Bacteriophage-encoded endolysins degrade the cell walls of Gram-positive bacteria by selectively hydrolyzing the peptidoglycan layer and thus are promising candidates to combat bacterial infections. PlyGRCS, the S. aureus-specific bacteriophage endolysin, contains a catalytic CHAP domain and a cell-wall binding SH3_5 domain connected by a linker. Here, we show the crystal structure of full-length PlyGRCS refined to 2.1 Å resolution. In addition, a serendipitous finding revealed that PlyGRCS binds to cold-shock protein C (CspC) by interacting with its CHAP and SH3_5 domains. CspC is an RNA chaperone that plays regulatory roles by conferring bacterial adaptability to various stress conditions. PlyGRCS has substantial lytic activity against S. aureus and showed only minimal change in its lytic activity in the presence of CspC. Whereas the PlyGRCS-CspC complex greatly reduced CspC-nucleic acid binding, the aforesaid complex may downregulate the CspC function during bacterial infection. Overall, the crystal structure and biochemical results of PlyGRCS provide a molecular basis for the bacteriolytic activity of PlyGRCS against S. aureus.

Two cold shock domain containing proteins trigger the development of infectious Trypanosoma brucei.

Cold shock proteins are members of a family of DNA- and RNA-binding proteins with one or more evolutionarily conserved cold shock domain (CSD). These proteins have a wide variety of biological functions, including DNA-damage repair, mRNA stability, and regulation of transcription, splicing and translation. We previously identified two CSD containing proteins, CSD1 and CSD2, in the protozoan parasite Trypanosoma brucei to be required for RBP6-driven metacyclic production, albeit at different steps of the developmental program. During metacyclogenesis T. brucei undergoes major morphological and metabolic changes that culminate in the establishment of quiescent metacyclic parasites and the acquisition of mammalian infectivity. To investigate the specific role of CSD1 and CSD2 in this process, we ectopically expressed CSD1 or CSD2 in non-infectious procyclic parasites and discovered that each protein is sufficient to produce infectious metacyclic parasites in 24 hours. Domain truncation assays determined that the N-terminal domain, but not the C-terminal domain, of CSD1 and CSD2 was required for metacyclic development. Furthermore, conserved amino acid residues in the CSD of CSD1 and CSD2, known to be important for binding nucleic acids, were found to be necessary for metacyclic production. Using single-end enhanced crosslinking and immunoprecipitation (seCLIP) we identified the specific binding motif of CSD1 and CSD2 as "ANACAU" and the bound mRNAs were enriched for biological processes, including lipid metabolism, microtubule-based movement and nucleocytoplasmic transport that are likely involved in the transition to bloodstream form-like cells.

FGF21 modulates hippocampal cold-shock proteins and CA2-subregion proteins in neonatal mice with hypoxia-ischemia.

BACKGROUND: Fibroblast growth factor 21 (FGF21) is a neuroprotectant with cognitive enhancing effects but with poorly characterized mechanism(s) of action, particularly in females. Prior studies suggest that FGF21 may regulate cold-shock proteins (CSPs) and CA2-marker proteins in the hippocampus but empirical evidence is lacking. METHODS: We assessed in normothermic postnatal day (PND) 10 female mice, if hypoxic-ischemic (HI) brain injury (25 min 8% O2/92% N2) altered endogenous levels of FGF21 in serum or in the hippocampus, or its receptor ß-klotho. We also tested if systemic administration of FGF21 (1.5 mg/kg) modulated hippocampal CSPs or CA2 proteins. Finally, we measured if FGF21 therapy altered markers of acute hippocampal injury. RESULTS: HI increased endogenous serum FGF21 (24 h), hippocampal tissue FGF21 (4d), and decreased hippocampal ß-klotho levels (4d). Exogenous FGF21 therapy modulated hippocampal CSP levels, and dynamically altered hippocampal CA2 marker expression (24 h and 4d). Finally, FGF21 ameliorated neuronal damage markers at 24 h but did not affect GFAP (astrogliosis) or Iba1 (microgliosis) levels at 4d. CONCLUSIONS: FGF21 therapy modulates CSP and CA2 protein levels in the injured hippocampus. These proteins serve different biological functions, but our findings suggest that FGF21 administration modulates them in a homeostatic manner after HI. IMPACT: Hypoxic-ischemic (HI) injury in female post-natal day (PND) 10 mice decreases hippocampal RNA binding motif 3 (RBM3) levels in the normothermic newborn brain. HI injury in normothermic newborn female mice alters serum and hippocampal fibroblast growth factor 21 (FGF21) levels 24 h post-injury. HI injury in normothermic newborn female mice alters hippocampal levels of N-terminal EF-hand calcium binding protein 2 (NECAB2) in a time-dependent manner. Exogenous FGF21 therapy ameliorates the HI-mediated loss of hippocampal cold-induced RNA-binding protein (CIRBP). Exogenous FGF21 therapy modulates hippocampal levels of CA2-marker proteins after HI.

HNRNPH1 regulates the neuroprotective cold-shock protein RBM3 expression through poison exon exclusion.

Enhanced expression of the cold-shock protein RNA binding motif 3 (RBM3) is highly neuroprotective both in vitro and in vivo. Whilst upstream signalling pathways leading to RBM3 expression have been described, the precise molecular mechanism of RBM3 cold induction remains elusive. To identify temperature-dependent modulators of RBM3, we performed a genome-wide CRISPR-Cas9 knockout screen using RBM3-reporter human iPSC-derived neurons. We found that RBM3 mRNA and protein levels are robustly regulated by several splicing factors, with heterogeneous nuclear ribonucleoprotein H1 (HNRNPH1) being the strongest positive regulator. Splicing analysis revealed that moderate hypothermia significantly represses the inclusion of a poison exon, which, when retained, targets the mRNA for nonsense-mediated decay. Importantly, we show that HNRNPH1 mediates this cold-dependent exon skipping via its thermosensitive interaction with a G-rich motif within the poison exon. Our study provides novel mechanistic insights into the regulation of RBM3 and provides further targets for neuroprotective therapeutic strategies.

Effect of low temperature on the resistance of Listeria monocytogenes and Escherichia coli O157:H7 to acid electrolyzed water.

Low temperature can affect the resistance of pathogenic bacteria to other external stress. The present study was envisaged to assess the tolerance of L. monocytogenes and E. coli O157:H7 to acidic electrolyzed water (AEW) under low temperature stress. AEW treatment caused a damage to cell membrane of the pathogenic bacteria, which led to protein leakage and DNA damage. Compared with the pathogenic bacteria cultured at 37 °C (pure culture), the L. monocytogenes and E. coli O157:H7 cells cultivated at low temperature presented a less damage and had a higher survival rate when exposed to AEW. Therefore, 4 °C or 10 °C grown bacteria were less susceptible to AEW than those cultured at 37 °C. And this phenomenon was verified when AEW was used to treat the pathogenic bacteria inoculated in salmon. In addition, transcriptomic sequencing technology (RNA-seq) was used to reveal the mechanism of AEW tolerance of L. monocytogenes under low temperature stress. Transcriptomic analysis showed the expression of the cold shock protein, regulation of DNA-templated transcription, ribosome pathway, phosphotransferase system (PTS), bacteria chemotaxis, SOS response and DNA repair were involved in the resistance of L. monocytogenes to AEW. We speculated that the direct modulation of the expression of cold shock protein CspD, the indirect effect on the expression of cspD by inhibiting the expression of Crp/Fnr family transcriptional regulator or enhancing the level of cAMP by regulating PTS could reduce the resistance of L. monocytogenes cultivated at 4 °C to AEW. Our study contributes to solving the problem of the reduced bacteriostatic effect in cold storage environment.

Development of a novel ex vivo organ culture system to improve preservation methods of regenerative tissues.

Recent advances in regenerative technology have made the regeneration of various organs using pluripotent stem cells possible. However, a simpler screening method for evaluating regenerated organs is required to apply this technology to clinical regenerative medicine in the future. We have developed a simple evaluation method using a mouse tooth germ culture model of organs formed by epithelial-mesenchymal interactions. In this study, we successfully established a simple method that controls tissue development in a temperature-dependent manner using a mouse tooth germ ex vivo culture model. We observed that the development of the cultured tooth germ could be delayed by low-temperature culture and resumed by the subsequent culture at 37 °C. Furthermore, the optimal temperature for the long-term preservation of tooth germ was 25 °C, a subnormothermic temperature that maintains the expression of stem cell markers. We also found that subnormothermic temperature induces the expression of cold shock proteins, such as cold-inducible RNA-binding protein, RNA-binding motif protein 3, and serine and arginine rich splicing factor 5. This study provides a simple screening method to help establish the development of regenerative tissue technology using a tooth organ culture model. Our findings may be potentially useful for making advances in the field of regenerative medicine.

Regulation of CIRP by genetic factors of SP1 related to cold sensitivity.

Cold-inducible RNA-binding-protein (CIRP) is a cold shock protein that plays a protective role in genotoxic stress response. CIRP modulates inflammation in human diseases, inhibits cell proliferation, and protects cells from genotoxic damage during cellular stress. The mild cold responsive element and specificity protein 1 (SP1) play a role in Cirp expression at low temperatures. Although previous studies have provided insights into the immune functions of SP1 or CIRP, the mechanisms by which CIRP and SP1 me diate inflammatory responses remain largely unknown. Therefore, in the current study, we examined whether Cirp expression is affected by genetic factors related to temperature sensitivity as well as under low temperature. We performed a genome-wide association study on cold sensitivity in 2,000 participants. Fifty-six genome-wide significant trait-locus pairs were identified (p<1×10-5, false discovery rate < 0.05). Among these variants, rs1117050 and rs11170510 had a strong linkage disequilibrium (r2 > 0.8) relationship and expression quantitative trait locus-associated signals with the nearest Sp1 gene. We confirmed that the minor alleles of rs11170510 and rs58123204 were associated with increased Sp1 expression. Additionally, Sp1 overexpression led to CIRP translocation from the nucleus to the cytoplasm. CIRP protein levels increased in serum samples that had minor alleles of rs11170510 and rs58123204. Levels of various pro-inflammatory cytokines were also significantly increased in human peripheral blood mononuclear cells with minor alleles of rs11170510 and rs58123204. These results suggest that genetic factors related to cold sensitivity regulate CIRP expression and function and provide valuable insights into prediction of potential diseases through analysis of inherent genetic factors in humans.

Functional genomics analysis of a phyllospheric Pseudomonas spp with potential for biological control against coffee rust.

BACKGROUND: Pseudomonas spp. promotes plant growth and colonizes a wide range of environments. During the annotation of a Coffea arabica ESTs database, we detected a considerable number of contaminant Pseudomonas sequences, specially associated with leaves. The genome of a Pseudomonas isolated from coffee leaves was sequenced to investigate in silico information that could offer insights about bacterial adaptation to coffee phyllosphere. In parallel, several experiments were performed to confirm certain physiological characteristics that could be associated with phyllospheric behavior. Finally, in vivo and in vitro experiments were carried out to verify whether this isolate could serve as a biocontrol agent against coffee rust and how the isolate could act against the infection.  RESULTS: The isolate showed several genes that are associated with resistance to environmental stresses, such as genes encoding heat/cold shock proteins, antioxidant enzymes, carbon starvation proteins, proteins that control osmotic balance and biofilm formation. There was an increase of exopolysaccharides synthesis in response to osmotic stress, which may protect cells from dessication on phyllosphere. Metabolic pathways for degradation and incorporation into citrate cycle of phenolic compounds present in coffee were found, and experimentally confirmed. In addition, MN1F was found to be highly tolerant to caffeine. The experiments of biocontrol against coffee leaf rust showed that the isolate can control the progress of the disease, most likely through competition for resources. CONCLUSION: Genomic analysis and experimental data suggest that there are adaptations of this Pseudomonas to live in association with coffee leaves and to act as a biocontrol agent.

Assessing the Role of Cold-Shock Protein C: a Novel Regulator of Acinetobacter baumannii Biofilm Formation and Virulence.

Acinetobacter baumannii is a formidable opportunistic pathogen that is notoriously difficult to eradicate from hospital settings. This resilience is often attributed to a proclivity for biofilm formation, which facilitates a higher tolerance toward external stress, desiccation, and antimicrobials. Despite this, little is known regarding the mechanisms orchestrating A. baumannii biofilm formation. Here, we performed RNA sequencing (RNA-seq) on biofilm and planktonic populations for the multidrug-resistant isolate AB5075 and identified 438 genes with altered expression. To assess the potential role of genes upregulated within biofilms, we tested the biofilm-forming capacity of their respective mutants from an A. baumannii transposon library. In so doing, we uncovered 24 genes whose disruption led to reduced biofilm formation. One such element, cold shock protein C (cspC), had a highly mucoid colony phenotype, enhanced tolerance to polysaccharide degradation, altered antibiotic tolerance, and diminished adherence to abiotic surfaces. RNA-seq of the cspC mutant revealed 201 genes with altered expression, including the downregulation of pili and fimbria genes and the upregulation of multidrug efflux pumps. Using transcriptional arrest assays, it appears that CspC mediates its effects, at least in part, through RNA chaperone activity, influencing the half-life of several important transcripts. Finally, we show that CspC is required for survival during challenge by the human immune system and is key for A. baumannii dissemination and/or colonization during systemic infection. Collectively, our work identifies a cadre of new biofilm-associated genes within A. baumannii and provides unique insight into the global regulatory network of this emerging human pathogen.

Heme binding to cold shock protein D, CspD, from Vibrio cholerae.

Cold shock protein D (CspD) is one of the homologous proteins of cold shock protein A (CspA), inhibiting DNA replication by binding to single-stranded DNA. We found that CspD from Vibrio cholerae (VcCspD) possesses one heme regulatory motif (HRM) sequence and specifically binds heme with a stoichiometry of 1:1. The binding of a synthetic single-stranded DNA oligomer (ssDNA) was followed by fluorescence quenching of Trp. The fluorescence quenching associated with the addition of ssDNA was suppressed in the presence of heme, indicating that heme binding to VcCspD inhibited the formation of the VcCspD-ssDNA complex. Such heme-induced inhibition was not observed for the VcCspD mutant with replacement of Cys22 in the HRM with alanine (C22A). Heme binding at Cys22 is, therefore, essential for the inhibition of ssDNA binding for VcCspD. The growth of Escherichia coli at 37 °C was slowed when VcCspD was overexpressed, indicating that VcCspD hampers the growth of E. coli. When the production of heme in cells was promoted by the addition of a heme precursor, δ-aminolevulinic acid, the growth of E. coli expressing VcCspD was decelerated, but the growth of E. coli expressing the C22A mutant was not decelerated. These observations allow us to conclude that heme specifically binds to the HRM region in VcCspD and inhibits the binding of target ssDNA, which suggests that heme functions as a regulatory molecule for DNA replication.

Correlating multi-functional role of cold shock domain proteins with intrinsically disordered regions.

Cold shock proteins (CSPs) are an ancient and conserved family of proteins. They are renowned for their role in response to low-temperature stress in bacteria and nucleic acid binding activities. In prokaryotes, cold and non-cold inducible CSPs are involved in various cellular and metabolic processes such as growth and development, osmotic oxidation, starvation, stress tolerance, and host cell invasion. In prokaryotes, cold shock condition reduces cell transcription and translation efficiency. Eukaryotic cold shock domain (CSD) proteins are evolved form of prokaryotic CSPs where CSD is flanked by N- and C-terminal domains. Eukaryotic CSPs are multi-functional proteins. CSPs also act as nucleic acid chaperons by preventing the formation of secondary structures in mRNA at low temperatures. In human, CSD proteins play a crucial role in the progression of breast cancer, colon cancer, lung cancer, and Alzheimer's disease. A well-defined three-dimensional structure of intrinsically disordered regions of CSPs family members is still undetermined. In this article, intrinsic disorder regions of CSPs have been explored systematically to understand the pleiotropic role of the cold shock family of proteins.

RBM3 is an outstanding cold shock protein with multiple physiological functions beyond hypothermia.

RNA-binding motif protein 3 (RBM3), an outstanding cold shock protein, is rapidly upregulated to ensure homeostasis and survival in a cold environment, which is an important physiological mechanism in response to cold stress. Meanwhile, RBM3 has multiple physiological functions and participates in the regulation of various cellular physiological processes, such as antiapoptosis, circadian rhythm, cell cycle, reproduction, and tumogenesis. The structure, conservation, and tissue distribution of RBM3 in human are demonstrated in this review. Herein, the multiple physiological functions of RBM3 were summarized based on recent research advances. Meanwhile, the cytoprotective mechanism of RBM3 during stress under various adverse conditions and its regulation of transcription were discussed. In addition, the neuroprotection of RBM3 and its oncogenic role and controversy in various cancers were investigated in our review.

New Insights into Cold Shock Proteins Effects in Human Cancer: Correlation with Susceptibility, Prognosis and Therapeutical Perspectives.

The microenvironment of the tumor cells is central to its phenotypic modification. One of the essential elements of this milieu is thermal regulation. An augment in local temperature has been reported to augment the tumor cell's responsiveness to chemoand radiation treatment. Cold shock proteins are RNA/DNA binding proteins identified by the existence of one or more cold shock domains. In humans, the best studied components of this group of proteins are called Y-box binding proteins, such as Y-box binding protein-1 (YB-1), but several other proteins have been recognized. Biological functions of these proteins extend from the control of transcription, translation and splicing to the regulation of exosomal RNA content. Several findings correlate an altered cold shock protein expression profile with tumor diseases. In this review we summarize the data for a causative participation of cold shock proteins in cancer onset and diffusion. Furthermore, the possible use of cold shock proteins for diagnostics, prognosis, and as targets for cancer treatment is exposed.

Cellular RNA Targets of Cold Shock Proteins CspC and CspE and Their Importance for Serum Resistance in Septicemic Escherichia coli.

The RNA chaperones, cold shock proteins CspC and CspE, are important in stress response and adaptation. We studied their role in the pathogenesis of a virulent Escherichia coli, representative of extraintestinal pathogenic E. coli (ExPEC) which are serum resistant and septicemic. We performed a global analysis to identify transcripts that interact with these cold shock proteins (CSPs), focusing on virulence-related genes. We used CLIP-seq, which combines UV cross-linking, immunoprecipitation and RNA sequencing. A large number of transcripts bound to the CSPs were identified, and many bind both CspC and CspE. Many transcripts were of genes involved in protein synthesis, transcription and energy metabolism. In addition, there were virulence-related genes, (i.e., fur and ryhB), essential for iron homeostasis. The CLIP-seq results were validated on two transcripts, clpX and tdcA, reported as virulence-associated. Deletion of either CspC or CspE significantly decreased their transcript levels and in a double deletion mutant cspC/cspE, the transcript stability of tdcA and clpX was reduced by 32-fold and 10-fold, respectively. We showed that these two genes are important for virulence, as deleting either of them resulted in loss of serum resistance, a requirement for sepsis. As several virulence-related transcripts interact with CspC or CspE, we determined the importance of these proteins for growth in serum and showed that deletion of either gene significantly reduced serum survival. This phenotype could be partially complemented by cspE and fully complemented by cspC. These results indicate that the two RNA chaperones are essential for virulence, and that CspC particularly critical. IMPORTANCE Virulent Escherichia coli strains that cause infections outside the intestinal tract-extraintestinal pathogenic E. coli (ExPEC)-constitute a major clinical problem worldwide. They are involved in several distinct conditions, including urinary tract infections, newborn meningitis, and sepsis. Due to increasing antibiotic resistance, these strains are a main factor in hospital and community-acquired infections. Because many strains, which do not cross-react immunologically are involved, developing a simple vaccine is not possible. Therefore, it is essential to understand the pathogenesis of these bacteria to identify potential targets for developing drugs or vaccines. One of the least investigated systems involves RNA binding proteins, important for stability of transcripts and global gene regulation. Two such proteins are CspC and CspE ("cold shock proteins"), RNA chaperones involved in stress adaptation. Here we performed a global analysis to identify the transcripts which are affected by these two chaperones, with focus on virulence-associated transcripts.

Annotation survey and life cycle transcriptomics of transcription factors in rust fungi (Pucciniales) identify a possible role for cold shock proteins in dormancy exit.

Fungi of the order Pucciniales are obligate plant biotrophs causing rust diseases. They exhibit a complex life cycle with the production of up to five spore types, infection of two unrelated hosts and an overwintering stage. Transcription factors (TFs) are key regulators of gene expression in eukaryote cells. In order to better understand genetic programs expressed during major transitions of the rust life cycle, we surveyed the complement of TFs in fungal genomes with an emphasis on Pucciniales. We found that despite their large gene numbers, rust genomes have a reduced repertoire of TFs compared to other fungi. The proportions of C2H2 and Zinc cluster - two of the most represented TF families in fungi - indicate differences in their evolutionary relationships in Pucciniales and other fungal taxa. The regulatory gene family encoding cold shock protein (CSP) showed a striking expansion in Pucciniomycotina with specific duplications in the order Pucciniales. The survey of expression profiles collected by transcriptomics along the life cycle of the poplar rust fungus revealed TF genes related to major biological transitions, e.g. response to environmental cues and host infection. Particularly, poplar rust CSPs were strongly expressed in basidia produced after the overwintering stage suggesting a possible role in dormancy exit. Expression during transition from dormant telia to basidia confirmed the specific expression of the three poplar rust CSP genes. Their heterologous expression in yeast improved cell growth after cold stress exposure, suggesting a probable regulatory function when the poplar rust fungus exits dormancy. This study addresses for the first time TF and regulatory genes involved in developmental transition in the rust life cycle opening perspectives to further explore molecular regulation in the biology of the Pucciniales.

A switch from α-helical to ß-strand conformation during co-translational protein folding.

Cellular proteins begin to fold as they emerge from the ribosome. The folding landscape of nascent chains is not only shaped by their amino acid sequence but also by the interactions with the ribosome. Here, we combine biophysical methods with cryo-EM structure determination to show that folding of a ß-barrel protein begins with formation of a dynamic α-helix inside the ribosome. As the growing peptide reaches the end of the tunnel, the N-terminal part of the nascent chain refolds to a ß-hairpin structure that remains dynamic until its release from the ribosome. Contacts with the ribosome and structure of the peptidyl transferase center depend on nascent chain conformation. These results indicate that proteins may start out as α-helices inside the tunnel and switch into their native folds only as they emerge from the ribosome. Moreover, the correlation of nascent chain conformations with reorientation of key residues of the ribosomal peptidyl-transferase center suggest that protein folding could modulate ribosome activity.

piRNAs Interact with Cold-Shock Domain-Containing RNA Binding Proteins and Regulate Neuronal Gene Expression During Differentiation.

piRNAs (PIWI-interacting RNAs) are a class of small non-coding RNAs (ncRNAs) abundantly expressed in germline cells and involved in suppressing the transposon activity. Interestingly, recent studies have found piRNA expression in the central nervous system (CNS), yet the underlying biological significance remains largely unknown. In this study, we investigated the expression and function of piRNAs during the retinoic acid (RA)-mediated neuronal differentiation in NT2 cells, a human embryonal carcinoma cell line. We identified a cohort of differentially expressed piRNAs by microarray. Two piRNAs, DQ582359 and DQ596268, were increasingly upregulated during the RA-induced differentiation and involved in regulating the expression of neuronal markers, MAP2 and TUBB3. Furthermore, these piRNAs were found to associate with cold-shock domain (CSD)-containing RNA binding proteins, DIS3, DIS3L2, and YB-1. Markedly, overexpression of these piRNAs further enhanced the protein levels of MAP2 and TUBB3, potentially by downregulating DIS3, DIS3L2, and YB-1. Hence, our study has identified a novel somatic function of piRNAs in regulating neuronal gene expression. The interaction of piRNA with some CSD-containing proteins can be further explored to enhance neuronal differentiation to treat neurodegenerative diseases.