Nascent Polypeptide-associated Complex Research Articles

Improving dermal fibroblast-to-epidermis communications and aging wound repair through extracellular vesicle-mediated delivery of Gstm2 mRNA

Skin aging is characterized by the disruption of skin homeostasis and impaired skin injury repair. Treatment of aging skin has long been limited by the unclear intervention targets and delivery techniques. Engineering extracellular vesicles (EVs) as an upgraded version of natural EVs holds great potential in regenerative medicine. In this study, we found that the expression of the critical antioxidant and detoxification gene Gstm2 was significantly reduced in aging skin. Thus, we constructed the skin primary fibroblasts-derived EVs encapsulating Gstm2 mRNA (EVsGstm2), and found that EVsGstm2 could significantly improve skin homeostasis and accelerate wound healing in aged mice. Mechanistically, we found that EVsGstm2 alleviated oxidative stress damage of aging dermal fibroblasts by modulating mitochondrial oxidative phosphorylation, and promoted dermal fibroblasts to regulate skin epidermal cell function by paracrine secretion of Nascent Polypeptide-Associated Complex Alpha subunit (NACA). Furthermore, we confirmed that NACA is a novel skin epidermal cell protective molecule that regulates skin epidermal cell turnover through the ROS-ERK-ETS-Cyclin D pathway. Our findings demonstrate the feasibility and efficacy of EVs-mediated delivery of Gstm2 for aged skin treatment and unveil novel roles of GSTM2 and NACA for improving aging skin.Graphical

Chp1 is a dedicated chaperone at the ribosome that safeguards eEF1A biogenesis

Cotranslational protein folding depends on general chaperones that engage highly diverse nascent chains at the ribosomes. Here we discover a dedicated ribosome-associated chaperone, Chp1, that rewires the cotranslational folding machinery to assist in the challenging biogenesis of abundantly expressed eukaryotic translation elongation factor 1A (eEF1A). Our results indicate that during eEF1A synthesis, Chp1 is recruited to the ribosome with the help of the nascent polypeptide-associated complex (NAC), where it safeguards eEF1A biogenesis. Aberrant eEF1A production in the absence of Chp1 triggers instant proteolysis, widespread protein aggregation, activation of Hsf1 stress transcription and compromises cellular fitness. The expression of pathogenic eEF1A2 variants linked to epileptic-dyskinetic encephalopathy is protected by Chp1. Thus, eEF1A is a difficult-to-fold protein that necessitates a biogenesis pathway starting with dedicated folding factor Chp1 at the ribosome to protect the eukaryotic cell from proteostasis collapse.

The Nascent Polypeptide-Associated Complex Subunit Alpha-Like Protein 1 (PmNACA1) Enhances Drought Tolerance by Scavenging ROS in Prunus mira Koehne

The Nascent Polypeptide-Associated Complex Subunit Alpha-Like Protein 1 (PmNACA1) Enhances Drought Tolerance by Scavenging ROS in Prunus mira Koehne

The nascent polypeptide-associated complex subunit Egd1 is required for efficient selective mitochondrial degradation in budding yeast

Selective degradation of dysfunctional or excess mitochondria is a fundamental process crucial for cell homeostasis in almost all eukaryotes. This process relies on autophagy, an intracellular self-eating system conserved from yeast to humans and is thus called mitophagy. Detailed mechanisms of mitophagy remain to be fully understood. Here we show that mitochondrial degradation in budding yeast, which requires the pro-mitophagic protein Atg32, is strongly reduced in cells lacking Egd1, a beta subunit of the nascent polypeptide-associated complex acting in cytosolic ribosome attachment and protein targeting to mitochondria. By contrast, loss of the sole alpha subunit Egd2 or the beta subunit paralogue Btt1 led to only a partial or slight reduction in mitophagy. We also found that phosphorylation of Atg32, a crucial step for priming mitophagy, is decreased in the absence of Egd1. Forced Atg32 hyperphosphorylation almost completely restored mitophagy in egd1-null cells. Together, we propose that Egd1 acts in Atg32 phosphorylation to facilitate mitophagy.

TbNACα negatively regulates Trichoderma breve T069 synthesis of ethyl caffeate and enhances antagonism of Sclerotium rolfsii1

The nascent polypeptide-associated complex (NAC) is involved in various biological functions in eukaryotes which have been extensively studied in animals and plants, but its role in the biocontrol action of microorganisms needs to be better understood. In this study, we investigated the function of TbNACα, one of the subunits of NAC, in the biocontrol activity of Trichoderma breve T069 against Sclerotium rolfsii. The TbNACα gene was deleted from T. breve T069, and the ΔTbNACα mutant showed significantly reduced mycelial growth, spore production, and spore germination. Additionally, volatile substances from ΔTbNACα had no significant effect on S. rolfsii, while non-volatile substances significantly inhibited the growth of S. rolfsii. Transcriptome sequencing results showed that compared to wild-type T069, the ΔTbNACα mutant had 3,398 differentially expressed genes, mainly regulating the expression of genes related to secondary metabolite biosynthetic enzymes, hydrolases, and membrane transport proteins. Untargeted metabolomics identified 50 upregulated metabolites (27 in positive ion mode and 23 in negative ion mode) in crude extracts from ΔTbNACα mutant metabolite broth. Moreover, the metabolic substances of ethyl caffeate had the strongest activity against S. rolfsii, with an EC50 of 107.15 μg·mL-1. Quantitative Real-time PCR (qPCR) analysis revealed that genes involved in the ethyl caffeate synthesis pathway were significantly upregulated in ΔTbNACα strains. This study provides a basis for the negative regulation of ethyl caffeate synthesis and elucidates the antagonistic inhibition mechanism of TbNACα in T. breve T069.

Nacα protects the larval fat body from cell death by maintaining cellular proteostasis in Drosophila

Protein homeostasis (proteostasis) is crucial for the maintenance of cellular homeostasis. Impairment of proteostasis activates proteotoxic and unfolded protein response pathways to resolve cellular stress or induce apoptosis in damaged cells. However, the responses of individual tissues to proteotoxic stress and evoking cell death program have not been extensively explored in vivo. Here, we show that a reduction in Nascent polypeptide-associated complex protein alpha subunit (Nacα) specifically and progressively induces cell death in Drosophila fat body cells. Nacα mutants disrupt both ER integrity and the proteasomal degradation system, resulting in caspase activation through JNK and p53. Although forced activation of the JNK and p53 pathways was insufficient to induce cell death in the fat body, the reduction of Nacα sensitized fat body cells to intrinsic and environmental stresses. Reducing overall protein synthesis by mTor inhibition or Minute mutants alleviated the cell death phenotype in Nacα mutant fat body cells. Our work revealed that Nacα is crucial for protecting the fat body from cell death by maintaining cellular proteostasis, thus demonstrating the coexistence of a unique vulnerability and cell death resistance in the fat body.

NAC controls cotranslational N-terminal methionine excision in eukaryotes.

N-terminal methionine excision from newly synthesized proteins, catalyzed cotranslationally by methionine aminopeptidases (METAPs), is an essential and universally conserved process that plays a key role in cell homeostasis and protein biogenesis. However, how METAPs interact with ribosomes and how their cleavage specificity is ensured is unknown. We discovered that in eukaryotes the nascent polypeptide-associated complex (NAC) controls ribosome binding of METAP1. NAC recruits METAP1 using a long, flexible tail and provides a platform for the formation of an active methionine excision complex at the ribosomal tunnel exit. This mode of interaction ensures the efficient excision of methionine from cytosolic proteins, whereas proteins targeted to the endoplasmic reticulum are spared. Our results suggest a broader mechanism for how access of protein biogenesis factors to translating ribosomes is controlled.

Lysine Methyltransferase SMYD1 Regulates Myogenesis via skNAC Methylation.

The SMYD family is a unique class of lysine methyltransferases (KMTases) whose catalytic SET domain is split by a MYND domain. Among these, Smyd1 was identified as a heart- and skeletal muscle-specific KMTase and is essential for cardiogenesis and skeletal muscle development. SMYD1 has been characterized as a histone methyltransferase (HMTase). Here we demonstrated that SMYD1 methylates is the Skeletal muscle-specific splice variant of the Nascent polypeptide-Associated Complex (skNAC) transcription factor. SMYD1-mediated methylation of skNAC targets K1975 within the carboxy-terminus region of skNAC. Catalysis requires physical interaction of SMYD1 and skNAC via the conserved MYND domain of SMYD1 and the PXLXP motif of skNAC. Our data indicated that skNAC methylation is required for the direct transcriptional activation of myoglobin (Mb), a heart- and skeletal muscle-specific hemoprotein that facilitates oxygen transport. Our study revealed that the skNAC, as a methylation target of SMYD1, illuminates the molecular mechanism by which SMYD1 cooperates with skNAC to regulate transcriptional activation of genes crucial for muscle functions and implicates the MYND domain of the SMYD-family KMTases as an adaptor to target substrates for methylation.

Transcriptional analysis reveals induction of systemic resistance in tomato treated with Chlorella microalgae

The application of algae-based products has been reported to promote plant growth and yield of tomato plants, especially by enhancing flowering. However, how microalgae (MA) affect plants at the molecular level remains elusive. The aim of this study was to elucidate the effects of live microalgae application on plant photosynthesis and the transcriptome of the unopened flower buds of tomato plants. Microalgae increased leaf temperature differential in tomato leaves but hardly affected photosynthesis. Contrary to our expectations, RNA-seq data revealed remarkable differential expression of several genes participating in responses to abiotic stresses but only a few genes involved in flowering or pollen/ spore development. Late Embryogenesis Abundant (LEA) proteins (mostly dehydrins), oleosins, ethylene, and abscisic-related genes, and Nascent Polypeptide-Associated Complex (NAC)-domain-containing proteins were upregulated. Genes involved in carbohydrate metabolism were also differentially expressed; glycolysis-related genes were upregulated, while those involved in sugar transport were downregulated. The only upregulated gene implicated in the induction of flowering was Solyc07g006500.3.1 (encoding trehalose-6-phosphate synthase enzyme TPS1). Overall, microalgae treatment led to an empirical upregulation of genes involved in jasmonic acid, abscisic acid, and ethylene pathways which are all essential for abiotic stress response. This study shows that microalgae treatment primed plants by inducing the expression of genes involved in response to abiotic stress, especially cold and water stress, as well as pathogen attack.

Differential plasma protein expression after ingestion of essential amino acid-based dietary supplement verses whey protein in low physical functioning older adults

In a recent randomized, double-blind, placebo-controlled trial, we were able to demonstrate the superiority of a dietary supplement composed of essential amino acids (EAAs) over whey protein, in older adults with low physical function. In this paper, we describe the comparative plasma protein expression in the same subject groups of EAAs vs whey. The plasma proteomics data was generated using SOMA scan assay. A total of twenty proteins were found to be differentially expressed in both groups with a 1.5-fold change. Notably, five proteins showed a significantly higher fold change expression in the EAA group which included adenylate kinase isoenzyme 1, casein kinase II 2-alpha, Nascent polypeptide-associated complex subunit alpha, peroxiredoxin-1, and peroxiredoxin-6. These five proteins might have played a significant role in providing energy for the improved cardiac and muscle strength of older adults with LPF. On the other hand, fifteen proteins showed slightly lower fold change expression in the EAA group. Some of these 15 proteins regulate metabolism and were found to be associated with inflammation or other comorbidities. Gene Ontology (GO) enrichment analysis showed the association of these proteins with several biological processes. Furthermore, protein–protein interaction network analysis also showed distinct networks between upregulated and downregulated proteins. In conclusion, the important biological roles of the upregulated proteins plus better physical function of participants in the EAAs vs whey group demonstrated that EAAs have the potential to improve muscle strength and physical function in older adults. This study was registered with ClinicalTrials.gov: NCT03424265 “Nutritional interventions in heart failure.”

Nascent polypeptide-Associated Complex and Signal Recognition Particle have cardiac-specific roles in heart development and remodeling.

Establishing a catalog of Congenital Heart Disease (CHD) genes and identifying functional networks would improve our understanding of its oligogenic underpinnings. Our studies identified protein biogenesis cofactors Nascent polypeptide-Associated Complex (NAC) and Signal-Recognition-Particle (SRP) as disease candidates and novel regulators of cardiac differentiation and morphogenesis. Knockdown (KD) of the alpha- (Nacα) or beta-subunit (bicaudal, bic) of NAC in the developing Drosophila heart disrupted cardiac developmental remodeling resulting in a fly with no heart. Heart loss was rescued by combined KD of Nacα with the posterior patterning Hox gene Abd-B. Consistent with a central role for this interaction in cardiogenesis, KD of Nacα in cardiac progenitors derived from human iPSCs impaired cardiac differentiation while co-KD with human HOXC12 and HOXD12 rescued this phenotype. Our data suggest that Nacα KD preprograms cardioblasts in the embryo for abortive remodeling later during metamorphosis, as Nacα KD during translation-intensive larval growth or pupal remodeling only causes moderate heart defects. KD of SRP subunits in the developing fly heart produced phenotypes that targeted specific segments and cell types, again suggesting cardiac-specific and spatially regulated activities. Together, we demonstrated directed function for NAC and SRP in heart development, and that regulation of NAC function depends on Hox genes.

The nascent polypeptide-associated complex (NAC) controls translation initiation in cis by recruiting nucleolin to the encoding mRNA.

Protein aggregates and abnormal proteins are toxic and associated with neurodegenerative diseases. There are several mechanisms to help cells get rid of aggregates but little is known on how cells prevent aggregate-prone proteins from being synthesised. The EBNA1 of the Epstein-Barr virus (EBV) evades the immune system by suppressing its own mRNA translation initiation in order to minimize the production of antigenic peptides for the major histocompatibility (MHC) class I pathway. Here we show that the emerging peptide of the disordered glycine–alanine repeat (GAr) within EBNA1 dislodges the nascent polypeptide-associated complex (NAC) from the ribosome. This results in the recruitment of nucleolin to the GAr-encoding mRNA and suppression of mRNA translation initiation in cis. Suppressing NAC alpha (NACA) expression prevents nucleolin from binding to the GAr mRNA and overcomes GAr-mediated translation inhibition. Taken together, these observations suggest that EBNA1 exploits a nascent protein quality control pathway to regulate its own rate of synthesis that is based on sensing the nascent GAr peptide by NAC followed by the recruitment of nucleolin to the GAr-encoding RNA sequence.

Extended disordered regions of ribosome-associated NAC proteins paralogs belong only to the germline in Drosophila melanogaster

The nascent polypeptide-associated complex (NAC) consisting of α- and β-subunits is an essential ribosome-associated protein conserved in eukaryotes. NAC is a ubiquitously expressed co-translational regulator of nascent protein folding and sorting providing for homeostasis of cellular proteins. Here we report on discovering the germline-specific NACαβ paralogs (gNACs), whose β-subunits, non-distinguishable by ordinary immunodetection, are encoded by five highly homologous gene copies, while the α-subunit is encoded by a single αNAC gene. The gNAC expression is detected in the primordial embryonic and adult gonads via immunostaining. The germline-specific α and β subunits differ from the ubiquitously expressed paralogs by the extended intrinsically disordered regions (IDRs) acquired at the N- and C-termini of the coding regions, predicted to be phosphorylated. The presence of distinct phosphorylated isoforms of gNAC-β subunits is confirmed by comparing of their profiles by 2D-isoeletrofocusing resolution before and after phosphatase treatment of testis ribosomes. We revealed that the predicted S/T sites of phosphorylation in the individual orthologous IDRs of gNAC-β sequences of Drosophila species are positionally conserved despite these disordered regions are drastically different. We propose the IDR-dependent molecular crowding and specific coordination of NAC and other proteostasis regulatory factors at the ribosomes of germinal cells. Our findings imply that there may be a functional crosstalk between the germinal and ubiquitous α- and β-subunits based on assessing their depletion effects on the fly viability and gonad development.

Proteomic Profile Analysis of Pulmonary Artery in a Rat Model Under Hypoxic Pulmonary Hypertension

Aim: Proteomic profile analysis of pulmonary artery in a rat model under hypoxic pulmonary hypertension Background: Background: Hypoxic pulmonary hypertension (HPH) is a pathological condition exemplified by a constant rise in pulmonary artery pressure in high-altitudes. Objective: Objective: To investigated the proteome profile and response mechanisms of SD rats under hypoxia over a period of four-weeks. Method: Method: Proteomic profile analysis of pulmonary artery in a rat model under hypoxic pulmonary hypertension. Results: Results: With 3,204 proteins identified, 49 were up-regulated while 46 were down-regulated. Upregulated genes included Prolargin, Protein S100-A6 and Transgelin-2, whereas Nascent polypeptide-associated complex and Elongator complex protein 1 were down-regulated. KEGG enriched pathways had purine metabolism, cancer and lipolysis regulation as significantly enriched in hypoxic group. Conclusion: Conclusion: In conclusion, our findings submit basis for downstream studies on tissue hypoxia mechanisms alongside the associated physiological conditions. Hypoxic pulmonary hypertension (HPH) is a pathological condition exemplified by a constant rise in pulmonary artery pressure in high altitudes. Herein, we investigated the proteome profile and response mechanisms of Sprague-Dawley (SD) rats under hypoxia over a period of four weeks. Unbiased iTRAQ-based quantitative proteomics was utilized in proteome profile analysis of a rat model exposed to HPH. With 3,204 proteins identified, 49 were upregulated while 46 were downregulated. Upregulated genes included Prolargin, Protein, S100-A6 and Transgelin-2, whereas Nascent polypeptide-associated complex and Elongator complex protein 1 were downregulated. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enriched pathways had purine metabolism, cancer, and lipolysis regulation as significantly enriched in hypoxic group. In conclusion, the findings from this study submit a basis for downstream studies on tissue hypoxia mechanisms alongside the associated physiological conditions.

Investigating the Roles of NAC and RAC in Suppressing Prion Formation in Saccharomyces cerevisiae

The correct three‐dimensional folding of a polypeptide is essential for its correct structure and function. The appearance of misfolded proteins can be found in relation to many disease processes. Prions are a class of infectious misfolded proteins, which are characterized by their ability to self‐propagate. In order to help ensure correct protein folding for the survival of the cell, several chaperone quality control complexes interact with nascent polypeptides as they are being synthesized by the ribosome. In yeast, two important ribosome‐tethered chaperone complexes exist, the Ribosome‐Associated Complex (RAC), composed of two subunits, Hsp70 chaperone Ssz1 and Hsp40 chaperone Zuo1, and the Nascent Polypeptide Associated Complex (NAC), a heterodimeric complex composed of an α subunit, Egd2, and one of the two β subunits, Btt1 or Egd1. Previous research by our lab and others has shown that cells lacking Zuo1 and thus, lacking functional RAC, show an increase in prion formation of the [PSI+] prion, a misfolded form of the Sup35 translation termination factor protein. In addition to its involvement in protein folding in the RAC, Zuo1 serves as a transcriptional co‐activator for genes in the pleiotropic drug resistance (PDR) response, which aids yeast survival in response to environmental stressors such as drugs or toxins. Environmental stress can also trigger prion formation in yeast. An overexpression screen for factors that increase yeast survival in the presence of the antibiotic cycloheximide, a compound known to result in PDR activation, revealed Btt1 to increase cell survival in a Zuo1 dependent manner. To investigate the interactions between RAC and NAC, as well as the role these complexes play in modulating prion formation during periods of cellular stress, we conducted prion formation assays and growth assays to assess how the complexes work together and separately in relation to the formation of the [PSI+] prion.

Positively charged amino acids at the N terminus of select mitochondrial proteins mediate early recognition by import proteins αβ′-NAC and Sam37

A major challenge in eukaryotic cells is the proper distribution of nuclear-encoded proteins to the correct organelles. For a subset of mitochondrial proteins, a signal sequence at the N terminus (matrix-targeting sequence [MTS]) is recognized by protein complexes to ensure their proper translocation into the organelle. However, the early steps of mitochondrial protein targeting remain undeciphered. The cytosolic chaperone nascent polypeptide–associated complex (NAC), which in yeast is represented as the two different heterodimers αβ-NAC and αβ′-NAC, has been proposed to be involved during the early steps of mitochondrial protein targeting. We have previously described that the mitochondrial outer membrane protein Sam37 interacts with αβ′-NAC and together promote the import of specific mitochondrial precursor proteins. In this work, we aimed to detect the region in the MTS of mitochondrial precursors relevant for their recognition by αβ′-NAC during their sorting to the mitochondria. We used targeting signals of different mitochondrial proteins (αβ′-NAC-dependent Oxa1 and αβ′-NAC-independent Mdm38) and fused them to GFP to study their intracellular localization by biochemical and microscopy methods, and in addition followed their import kinetics in vivo. Our results reveal the presence of a positively charged amino acid cluster in the MTS of select mitochondrial precursors, such as Oxa1 and Fum1, which are crucial for their recognition by αβ′-NAC. Furthermore, we explored the presence of this cluster at the N terminus of the mitochondrial proteome and propose a set of precursors whose proper localization depends on both αβ′-NAC and Sam37.

Mechanism of signal sequence handover from NAC to SRP on ribosomes during ER-protein targeting.

The nascent polypeptide-associated complex (NAC) interacts with newly synthesized proteins at the ribosomal tunnel exit and competes with the signal recognition particle (SRP) to prevent mistargeting of cytosolic and mitochondrial polypeptides to the endoplasmic reticulum (ER). How NAC antagonizes SRP and how this is overcome by ER targeting signals are unknown. Here, we found that NAC uses two domains with opposing effects to control SRP access. The core globular domain prevented SRP from binding to signal-less ribosomes, whereas a flexibly attached domain transiently captured SRP to permit scanning of nascent chains. The emergence of an ER-targeting signal destabilized NAC's globular domain and facilitated SRP access to the nascent chain. These findings elucidate how NAC hands over the signal sequence to SRP and imparts specificity of protein localization.

Osj10gBTF3-Mediated Import of Chloroplast Protein Is Essential for Pollen Development in Rice.

Chloroplasts are crucial organelles for the generation of fatty acids and starch required for plant development. Nascent polypeptide-associated complex (NAC) proteins have been implicated in development as transcription factors. However, their chaperone roles in chloroplasts and their relationship with pollen development in plants remain to be elucidated. Here, we demonstrated that Osj10gBTF3, a NAC protein, regulates pollen and chloroplast development in rice by coordinating with a Hsp90 family chaperone OsHSP82 to mediate chloroplast import. Knockout of Osj10gBTF3 affects pollen and chloroplast development and significantly reduces the accumulation of fertility-related chloroplast protein OsPPR676. Both Osj10gBTF3 and OsHSP82 interact with OsPPR676. Interestingly, the interaction between OsHSP82 and OsPPR676 is only found in the cytoplasm, while the interaction between Osj10gBTF3 and OsPPR676 also occurs inside the chloroplast. The chloroplast stroma chaperone OsCpn60 can also be co-precipitated with Osj10gBTF3, but not with OsHSP82. Further investigation indicates that Osj10gBTF3 enters the chloroplast stroma possibly through the inner chloroplast membrane channel protein Tic110 and then recruits OsCpn60 for the folding or assembly of OsPPR676. Our results reveal a chaperone role of Osj10gBTF3 in chloroplast import different from Hsp90 and provide a link between chloroplast transport and pollen development in rice.

Identification of Binding Partners of CsaA - An Archaeal Chaperonic Protein of Picrophilus torridus.

CsaA is among the few chaperones which are present in both bacteria and archaea, but absent in eukaryotes. There are no reports on interactome analysis of CsaA from archaea, till date. Identification of binding partners of CsaA might be helpful in understanding CsaA-associated processes in Picrophilus torridus an extreme thermoacidophilic euryarchaeon. The present study was conducted to identify the binding partners of CsaA of P. torridus (PtCsaA). The binding partners of PtCsaA were isolated and identified using a pull down assay and liquid chromatography-mass spectrometry (LC-MS). The results revealed twelve potential binding partners of CsaA. These were thermosome subunits (Q6KZS2 and Q6L132), nascent polypeptide-associated complex protein (Q6L1N3), elongation factor 1-alpha (Q6L202), uncharacterized protein (Q6L0Y6), citrate synthase (Q6L0M8), asparaginyl- tRNA synthetase (Q6L0M5), succinyl-CoA synthetase beta chain (Q6L0B4), pyruvate ferredoxin oxidoreductase alpha and beta chain proteins (Q6KZA7 and Q6KZA6, respectively), malate dehydrogenase (Q6L0C3) and reversed fumarylacetoacetase (Q6KZ97). Functional categorization revealed that of these, six proteins were involved in energy metabolic pathways, three were archaeal chaperones, two were involved in translation and one might be a transcription regulator. STRING-based analysis of the protein-protein interactions of the experimental interactome revealed strong interactions among them. PtCsaA might be a multifaceted protein which besides translation might also play important role in metabolic processes of P. torridus. However, further experiments investigating the binding partners of CsaA in other archaea are required for a better understanding of CsaA-associated processes in archaea.

Molecular Characterization of the Oncogene BTF3 and Its Targets in Colorectal Cancer.

Colorectal cancer (CRC) is one of the most commonly diagnosed and leading causes of cancer mortality worldwide, and the prognosis of patients with CRC remains unsatisfactory. Basic transcription factor 3 (BTF3) is an oncogene and hazardous prognosticator in CRC. Although two distinct functional mechanisms of BTF3 in different cancer types have been reported, its role in CRC is still unclear. In this study, we aimed to molecularly characterize the oncogene BTF3 and its targets in CRC. Here, we first identified the transcriptional targets of BTF3 by applying combined RNA-Seq and ChIP-Seq analysis, identifying CHD1L as a transcriptional target of BTF3. Thereafter, we conducted immunoprecipitation (IP)-MS and E3 ubiquitin ligase analysis to identify potential interacting targets of BTF3 as a subunit of the nascent-polypeptide-associated complex (NAC). The analysis revealed that BTF3 might also inhibit E3 ubiquitin ligase HERC2-mediated p53 degradation. Finally, miRNAs targeting BTF3 were predicted and validated. Decreased miR-497-5p expression is responsible for higher levels of BTF3 post-transcriptionally. Collectively, we concluded that BTF3 is an oncogene, and there may exist a transcription factor and NAC-related proteolysis mechanism in CRC. This study provides a comprehensive basis for understanding the oncogenic mechanisms of BTF3 in CRC.