Tetratricopeptide Research Articles

Tah1, A Key Component of R2TP Complex that Regulates Assembly of snoRNP, is Involved in De Novo Generation and Maintenance of Yeast Prion [URE3

The cellular chaperone machinery plays key role in the de novo formation and propagation of yeast prions (infectious protein). Though the role of Hsp70s in the prion maintenance is well studied, how Hsp90 chaperone machinery affects yeast prions remains unclear. In the current study, we examined the role of Hsp90 and its co-chaperones on yeast prions [PSI+] and [URE3]. We show that the overproduction of Hsp90 co-chaperone Tah1, cures [URE3] which is a prion form of native protein Ure2 in yeast. The Hsp90 co-chaperone Tah1 is involved in the assembly of small nucleolar ribonucleoproteins (snoRNP) and chromatin remodelling complexes. We found that Tah1 deletion improves the frequency of de novo appearance of [URE3]. The Tah1 was found to interact with Hsp70. The lack of Tah1 not only represses antagonizing effect of Ssa1 Hsp70 on [URE3] but also improves the prion strength suggesting role of Tah1 in both fibril growth and replication. We show that the N-terminal tetratricopeptide repeat domain of Tah1 is indispensable for [URE3] curing. Tah1 interacts with Ure2, improves its solubility in [URE3] strains, and affects the kinetics of Ure2 fibrillation in vitro. Its inhibitory role on Ure2 fibrillation is proposed to influence [URE3] propagation. The present study shows a novel role of Tah1 in yeast prion propagation, and that Hsp90 not only promotes its role in ribosomal RNA processing but also in the prion maintenance. SummaryPrions are self-perpetuating infectious proteins. What initiates the misfolding of a protein into its prion form is still not clear. The understanding of cellular factors that facilitate or antagonize prions is crucial to gain insight into the mechanism of prion formation and propagation. In the current study, we reveal that Tah1 is a novel modulator of yeast prion [URE3]. The Hsp90 co-chaperone Tah1, is required for the formation of small nucleolar ribonucleoprotein complex. We show that the absence of Tah1 improves the induction of [URE3] prion. The overexpressed Tah1 cures [URE3], and this function is promoted by Hsp90 chaperones. The current study thus provides a novel cellular factor and the underlying mechanism, involved in the prion formation and propagation

Dermatopontin as a potential pathogenic factor in endometrial cancer

The present study aimed to determine the differential expression profiles of proteins in endometrial carcinoma and to screen the proteins associated with the occurrence and development of endometrial cancer (EC). In total, 15 samples of human EC and paracancerous tissues were selected for proteomic analysis using a label-free quantification method based on liquid chromatography-tandem mass spectrometry. The differential proteins were analysed using bioinformatics and verified using reverse transcription-quantitative PCR (RT-qPCR) and western blotting. Finally, the expression of differential proteins in 75 endometrial carcinoma samples and 30 normal endometrial tissue samples were detected using immunohistochemical staining, and the associations between differential protein expression and clinicopathological features were analysed. In total, 579 up-regulated proteins and 346 down-regulated proteins were identified between the two groups and seven proteins with the most significant differences were selected; these proteins included interferon-induced protein with tetratricopeptide repeats 3, poly(ADP-ribose) polymerase family member 9, solute carrier family 34 member 2, cytochrome b5 reductase 1, protein tyrosine phosphatase non-receptor type 1, dermatopontin (DPT) and secretory leukocyte peptidase inhibitor. RT-qPCR and western blotting showed that DPT expression was down-regulated (P<0.001), which was consistent with the mass spectrometry results. The immunohistochemical staining results showed that the positive expression of DPT in EC and normal endometrial tissues was statistically significant (P<0.001). The positive expression of DPT was significantly decreased in poorly differentiated, late stage, lymph node metastasis and myometrial invasion depth ≥1/2 samples (P<0.05). DPT expression was significantly lower in EC, which might play role in the pathogenesis of EC.

Optimizing the First TPR Domain of the Human SPAG1 Protein Provides Insight into the HSP70 and HSP90 Binding Properties.

Tetratricopeptide repeat domains, or TPR domains, are protein domains that mediate protein:protein interaction. As they allow contacts between proteins, they are of particular interest in transient steps of the assembly process of macromolecular complexes, such as the ribosome or the dynein arms. In this study, we focused on the first TPR domain of the human SPAG1 protein. SPAG1 is a multidomain protein that is important for ciliogenesis whose known mutations are linked to primary ciliary dyskinesia syndrome. It can interact with the chaperones RUVBL1/2, HSP70, and HSP90. Using protein sequence optimization in combination with structural and biophysical approaches, we analyzed, with atomistic precision, how the C-terminal tails of HSPs bind a variant form of SPAG1-TPR1 that mimics the wild-type domain. We discuss our results with regard to other complex three-dimensional structures with the aim of highlighting the motifs in the TPR sequences that could drive the positioning of the HSP peptides. These data could be important for the druggability of TPR regulators.

LncRNA HOTAIR modulates chondrocyte apoptosis and inflammation in osteoarthritis via regulating miR-1277-5p/SGTB axis.

Osteoarthritis (OA) is a common degenerative joint disease in the elderly. This study aimed to investigate the role and mechanism of lncRNA HOX transcript antisense RNA (HOTAIR) in lipopolysaccharide (LPS)-treated chondrocytes in OA. CHON-001 chondrocytes treated with LPS were used as a cell model of OA. The levels of HOTAIR, miR-1277-5p and small glutamine rich tetratricopeptide repeat containing beta (SGTB) were measured via quantitative real-time polymerase chain reaction or western blot. Cell viability and apoptosis were assessed using Cell Counting Kit-8 and flow cytometry. The levels of inflammation-related factors were examined via enzyme-linked immunosorbent assay (ELISA). Aggrecan and Collagen II protein levels were detected using western blot. The interaction among HOTAIR, miR-1277-5p and SGTB were validated by dual-luciferase reporter analysis. HOTAIR and SGTB were up-regulated, while miR-1277-5p was down-regulated in OA cartilages and LPS-stimulated CHON-001 chondrocytes. HOTAIR depletion inhibited LPS-induced apoptosis and inflammation in chondrocytes. Moreover, down-regulation of HOTAIR attenuated LPS-triggered chondrocyte apoptosis and inflammation via sponging miR-1277-5p. Also, miR-1277-5p repressed LPS-induced chondrocyte apoptosis and inflammation by targeting SGTB. Furthermore, HOTAIR enhanced SGTB expression by sponging miR-1277-5p. HOTAIR aggravated chondrocyte apoptosis and inflammation in OA via regulating miR-1277-5p/SGTB pathway.

Protein Substrates Engage the Lumen of O-GlcNAc Transferase's Tetratricopeptide Repeat Domain in Different Ways.

Glycosylation of nuclear and cytoplasmic proteins is an essential post-translational modification in mammals. O-GlcNAc transferase (OGT), the sole enzyme responsible for this modification, glycosylates more than 1000 unique nuclear and cytoplasmic substrates. How OGT selects its substrates is a fundamental question that must be answered to understand OGT's unusual biology. OGT contains a long tetratricopeptide repeat (TPR) domain that has been implicated in substrate selection, but there is almost no information about how changes to this domain affect glycosylation of individual substrates. By profiling O-GlcNAc in cell extracts and probing glycosylation of purified substrates, we show here that ladders of asparagines and aspartates that extend the full length of OGT's TPR lumen control substrate glycosylation. Different substrates are sensitive to changes in different regions of OGT's TPR lumen. We also found that substrates with glycosylation sites close to the C-terminus bypass lumenal binding. Our findings demonstrate that substrates can engage OGT in a variety of different ways for glycosylation.

Molecular Characterization and Expression Analysis of SKIV Infection of Interferon-Induced Protein with Tetratricopeptide Repeats 1 (IFIT1) in Epinephelus lanceolatus

Interferon-induced protein with tetratricopeptide repeats 1 (IFIT1), also known as interferon-induced protein 56 (IFI56) or Interferon-stimulated protein 56 (ISG56), was originally identified as a protein induced upon treatment with interferon and inhibited by viral replication and translational initiation. In this study, Epinephelus lanceolatus IFIT1 (ELIFIT1) gene was cloned for the first time. The complete cDNA of ElIFIT1 gene includes 2921 nucleotides, and encodes a 437-amino acid (AA) protein. The putative ELIFIT1 protein has 9 TRP domains and is highly similar with IFIT1 proteins in other teleosts. In healthy fish, ELIFIT1 gene was highly expressed in the blood, which indicate its specific function in the peripheral immune system. Its expression was also observed in various immunity-related tissues including spleen, intestine, and kidney, Inducted with spotted knifejaw iridovirus (SKIV), ELIFIT1 gene expression was upregulated in the spleen, kidney, and liver 24 h after induction and reached its peak at 72 h, indicating that ELIFIT1 may play an important role in antivirus. These findings contribute to the understanding of the antiviral regulation of ELIFIT1 gene in teleost.

Differential regulation of the glucocorticoid receptor nucleocytoplasmic shuttling by TPR-domain proteins

A dimer of the heat-shock protein of 90-kDa (Hsp90) represents the critical core of the chaperone complex associated to the glucocorticoid receptor (GR) oligomer. The C-terminal end of the Hsp90 dimer shapes a functional acceptor site for co-chaperones carrying tetratricopeptide repeat (TPR) domains, where they bind in a mutually exclusive and competitive manner. They impact on the biological properties of the GR•Hsp90 complex and are major players of the GR transport machinery. Recently, we showed that the overexpression of a chimeric TPR peptide influences the subcellular distribution of GR. In this study, the functional role of endogenous proteins carrying TPR or TPR-like sequences on GR subcellular distribution was characterized. It is demonstrated that, contrarily to the positive influence of FKBP52 on GR nuclear accumulation, FKBP51 and 14-3-3 impaired this property. While SGT1α showed no significant effect, the overexpression of the Ser/Thr phosphatase PP5 resulted in a nearly equal nuclear-cytoplasmic redistribution of GR rather than its typical cytoplasmic localization in the absence of steroid. This observation led to analyse the influence of the phosphorylation status of GR, which resulted not linked to its nucleo-cytoplasmic shuttling mechanism. Nonetheless, it was evidenced that both PP5 and FKBP52 are related to the anchorage of the GR to nucleoskeleton structures. The influence of these TPR domain proteins on the steroid-dependent transcriptional activity of GR was also characterized. It is postulated that the pleiotropic actions of the GR in different cell types may be the consequence of the relative abundance of different TPR-domain interacting co-chaperones.

Versatile allosteric properties in Pex5-like tetratricopeptide repeat proteins to induce diverse downstream function.

Proteins composed of tetratricopeptide repeat (TPR) arrays belong to the α-solenoid tandem-repeat family that have unique properties in terms of their overall conformational flexibility and ability to bind to multiple protein ligands. The peroxisomal matrix protein import receptor Pex5 comprises two TPR triplets that recognize protein cargos with a specific C-terminal Peroxisomal Targeting Signal (PTS) 1 motif. Import of PTS1-containing protein cargos into peroxisomes through a transient pore is mainly driven by allosteric binding, coupling and release mechanisms, without a need for external energy. A very similar TPR architecture is found in the functionally unrelated TRIP8b, a regulator of the hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channel. TRIP8b binds to the HCN ion channel via a C-terminal sequence motif that is nearly identical to the PTS1 motif of Pex5 receptor cargos. Pex5, Pex5-related Pex9, and TRIP8b also share a less conserved N-terminal domain. This domain provides a second protein cargo-binding site and plays a distinct role in allosteric coupling of initial cargo loading by PTS1 motif-mediated interactions and different downstream functional readouts. The data reviewed here highlight the overarching role of molecular allostery in driving the diverse functions of TPR array proteins, which could form a model for other α-solenoid tandem-repeat proteins involved in translocation processes across membranes.

Derivation of stem cell line UMi028-A-2 containing a CRISPR/Cas9 induced Alzheimer's disease risk variant p.S1038C in the TTC3 gene.

The UMi028-A-2 human induced pluripotent stem cell line carries a homozygous mutation (rs377155188, C>G, p.S1038C) in the tetratricopeptide repeat domain 3 (TTC3) gene that was introduced via CRISPR/Cas9 genome editing. The line was originally derived from a neurologically normal male and has been thoroughly characterized following editing. The p.S1038C variant has been shown to potentially contribute to the risk of late onset Alzheimer's disease and is a resource to further investigate the consequences of TTC3 and this alteration in disease pathology.

Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1.

The process of displaying functional peptides by ‘grafting’ them onto loops of a stable protein scaffold can be used to impart binding affinity for a target, but it can be difficult to predict the affinity of the grafted peptide and the effect of grafting on scaffold stability. In this study, we show that a series of peptides that bind to the E3 ubiquitin ligase Keap1 can be grafted into the inter-repeat loop of a consensus-designed tetratricopeptide repeat (CTPR) protein resulting in proteins with high stability. We found that these CTPR-grafted peptides had similar affinities to their free peptide counterparts and achieved a low nanomolar range. This result is likely due to a good structural match between the inter-repeat loop of the CTPR and the Keap1-binding peptide. The grafting process led to the discovery of a new Keap1-binding peptide, Ac-LDPETGELL-NH2, with low nanomolar affinity for Keap1, highlighting the potential of the repeat-protein class for application in peptide display.

Characteristics of Clinical and Electrophysiological Pattern in a Large Cohort of Chinese Patients With Charcot-Marie-Tooth 4C.

The “Src homology 3 (SH3) domain and tetratricopeptide repeats 2” (SH3TC2) gene is mutated in individuals with Charcot-Marie-Tooth disease (CMT) and considered relevant to a demyelinating or intermediate subtype of CMT disease, CMT4C. In this study, we screened a cohort of 465 unrelated Chinese CMT patients alongside 650 controls. We used Sanger, next-generation, or whole-exome sequencing to analyze SH3TC2 and other CMT-related genes and identified 12 SH3TC2 variants (eight novel) in seven families. Of the eight novel variants, seven were likely pathogenic (c.280–2 A > G, c.732–1 G > A, c.1177+6 T > C, c.3328–1 G > A, G299S, R548W, L1048P), and 1 had uncertain significance (S221P). The CMT4C frequency was calculated to be 4.24% in demyelinating or intermediate CMT patients without PMP22 duplication. Additionally, we detected variant R954* in the Chinese cohort in our study, indicating that this variant may be present among Asians, albeit with a relatively low frequency. The onset age varied among the eight patients, three of whom presented scoliosis. We summarized phenotypes in the Chinese CMT cohort and concluded that the absence of scoliosis, cranial nerve involvement, or late-onset symptoms does not necessarily preclude SH3TC2 involvement in a given case.

Dynamic Changes in the Expression of Interferon-Stimulated Genes in Joints of SPF Chickens Infected With Avian Reovirus.

Avian reovirus (ARV) can induce many diseases as well as immunosuppression in chickens, severely endangering the poultry industry. Interferons (IFNs) play an antiviral role by inducing the expression of interferon-stimulated genes (ISGs). The effect of ARV infection on the expression of host ISGs is unclear. Specific-pathogen-free (SPF) chickens were infected with ARV strain S1133 in this study, and real time quantitative PCR was used to detect changes in the dynamic expression of IFNs and common ISGs in joints of SPF chickens. The results showed that the transcription levels of IFNA, IFNB, and several ISGs, including myxovirus resistance (MX), interferon-induced transmembrane protein 3 (IFITM3), protein kinase R (PKR), oligoadenylate synthase (OAS), interferon-induced protein with tetratricopeptide repeats 5 (IFIT5), interferon-stimulated gene 12 (ISG12), virus inhibitory protein (VIPERIN), interferon-alpha-inducible protein 6 (IFI6), and integrin-associated protein (CD47), were upregulated in joints on days 1–7 of infection (the levels of increase of MX, IFIT5, OAS, VIPERIN, ISG12, and IFI6 were the most significant, at hundreds-fold). In addition, the expression levels of the ISGs encoding zinc finger protein 313 (ZFP313), and DNA damage–inducible transcript 4 (DDIT4) increased suddenly on the 1st or 2nd day, then decreased to control levels. The ARV viral load in chicken joints rapidly increased after 1 day of viral challenge, and the viral load remained high within 6 days of viral challenge. The ARV viral load sharply decreased starting on day 7. These results indicate that in SPF chicken joints, many ISGs have mRNA expression patterns that are basically consistent with the viral load in joints. IFNA, IFNB, and the ISGs MX, IFITM3, PKR, OAS, IFIT5, ISG12, VIPERIN, IFI6, and CD47 play important roles in defending against ARV invasion, inhibiting ARV replication and proliferation, and promoting virus clearance. These results enrich our understanding of the innate immune response mechanisms of hosts against ARV infection and provide a theoretical basis for prevention and control of ARV infection.

Structural Basis for Recognition of the Hsp90 Closed, ATP State by the TPR-Containing Co-Chaperone FKBP51

Molecular chaperones Hsp90 and Hsp70 are critical in protein homeostasis (proteostasis), interacting directly with many “client” proteins to promote ATP-driven remodeling and activation. Co-chaperones bind Hsp90 and Hsp70 forming dynamic chaperone complexes which specify the client folding pathway. Many interact via their tetratricopeptide repeat (TPR) domain, which binds the EEVD motif at the flexible carboxyl-terminus of Hsp90 and Hsp70. However, for many TPR cochaperones, the protein:protein interactions and mechanisms which enable Hsp70/Hsp90 specificity and regulation in the folding pathway remain unclear. The peptidyl-prolyl isomerase FK506-binding protein 51 (FKBP51) interacts specifically with Hsp90 (and not Hsp70) to promote the folding of key client proteins including kinases, steroid hormone receptors, and Tau. Here we have determined a cryo-EM structure of the human Hsp90:FKBP51:p23 complex to 3.3 Å resolution, revealing a distinct TPR interaction mechanism which enables recognition of the Hsp90 closed, ATP state via binding to the C-terminal Hsp90 dimer. Moreover, this interaction positions the FK1 domain of FKBP51 close to the middle domain of Hsp90 revealing a potential mechanism for isomerase activity targeted to distinct sites of Hsp90-bound clients. Together with biochemical analysis to probe interactions, these results reveal how the immunophilin class of TPR co-chaperones may act on specific client regulation and folding steps of the Hsp90-driven chaperone pathway.

Identification and Functional Analysis of Tomato TPR Gene Family.

Tomato (Solanum lycopersicum) as an important vegetable grown around the world is threatened by many diseases, which seriously affects its yield. Therefore, studying the interaction between tomato and pathogenic bacteria is biologically and economically important. The TPR (Tetratricopeptide repeat) gene family is a class of genes containing TPR conserved motifs, which are widely involved in cell cycle regulation, gene expression, protein degradation and other biological processes. The functions of TPR gene in Arabidopsis and wheat plants have been well studied, but the research on TPR genes in tomato is not well studied. In this study, 26 TPR gene families were identified using bioinformatics based on tomato genome data, and they were analyzed for subcellular localization, phylogenetic evolution, conserved motifs, tissue expression, and GO (Gene Ontology) analysis. The qRT-PCR was used to detect the expression levels of each member of the tomato TPR gene family (SlTPRs) under biological stress (Botrytis cinerea) and abiotic stress such as drought and abscisic acid (ABA). The results showed that members of the tomato TPR family responded to various abiotic stresses and Botrytis cinerea stress, and the SlTPR2 and SlTPR4 genes changed significantly under different stresses. Using VIGS (Virus-induced gene silencing) technology to silence these two genes, the silenced plants showed reduced disease resistance. It was also shown that TPR4 can interact with atpA which encodes a chloroplast ATP synthase CF1 α subunit. The above results provide a theoretical basis for further exploring the molecular mechanism of TPR-mediated resistance in disease defense, and also provide a foundation for tomato disease resistance breeding.

Chromosome-scale genome assembly provides insights into the evolution and flavor synthesis of passion fruit (Passiflora edulis Sims)

Passion fruit (Passiflora edulis Sims) is an economically valuable fruit that is cultivated in tropical and subtropical regions of the world. Here, we report an ~1341.7 Mb chromosome-scale genome assembly of passion fruit, with 98.91% (~1327.18 Mb) of the assembly assigned to nine pseudochromosomes. The genome includes 23,171 protein-coding genes, and most of the assembled sequences are repetitive sequences, with long-terminal repeats (LTRs) being the most abundant. Phylogenetic analysis revealed that passion fruit diverged after Brassicaceae and before Euphorbiaceae. Ks analysis showed that two whole-genome duplication events occurred in passion fruit at 65 MYA and 12 MYA, which may have contributed to its large genome size. An integrated analysis of genomic, transcriptomic, and metabolomic data showed that ‘alpha-linolenic acid metabolism’, ‘metabolic pathways’, and ‘secondary metabolic pathways’ were the main pathways involved in the synthesis of important volatile organic compounds (VOCs) in passion fruit, and this analysis identified some candidate genes, including GDP-fucose Transporter 1-like, Tetratricopeptide repeat protein 33, protein NETWORKED 4B isoform X1, and Golgin Subfamily A member 6-like protein 22. In addition, we identified 13 important gene families in fatty acid pathways and eight important gene families in terpene pathways. Gene family analysis showed that the ACX, ADH, ALDH, and HPL gene families, especially ACX13/14/15/20, ADH13/26/33, ALDH1/4/21, and HPL4/6, were the key genes for ester synthesis, while the TPS gene family, especially PeTPS2/3/4/24, was the key gene family for terpene synthesis. This work provides insights into genome evolution and flavor trait biology and offers valuable resources for the improved cultivation of passion fruit.

TRK-Fused Gene (TFG), a protein involved in protein secretion pathways, is an essential component of the antiviral innate immune response.

Antiviral innate immune response to RNA virus infection is supported by Pattern-Recognition Receptors (PRR) including RIG-I-Like Receptors (RLR), which lead to type I interferons (IFNs) and IFN-stimulated genes (ISG) production. Upon sensing of viral RNA, the E3 ubiquitin ligase TNF Receptor-Associated Factor-3 (TRAF3) is recruited along with its substrate TANK-Binding Kinase (TBK1), to MAVS-containing subcellular compartments, including mitochondria, peroxisomes, and the mitochondria-associated endoplasmic reticulum membrane (MAM). However, the regulation of such events remains largely unresolved. Here, we identify TRK-Fused Gene (TFG), a protein involved in the transport of newly synthesized proteins to the endomembrane system via the Coat Protein complex II (COPII) transport vesicles, as a new TRAF3-interacting protein allowing the efficient recruitment of TRAF3 to MAVS and TBK1 following Sendai virus (SeV) infection. Using siRNA and shRNA approaches, we show that TFG is required for virus-induced TBK1 activation resulting in C-terminal IRF3 phosphorylation and dimerization. We further show that the ability of the TRAF3-TFG complex to engage mTOR following SeV infection allows TBK1 to phosphorylate mTOR on serine 2159, a post-translational modification shown to promote mTORC1 signaling. We demonstrate that the activation of mTORC1 signaling during SeV infection plays a positive role in the expression of Viperin, IRF7 and IFN-induced proteins with tetratricopeptide repeats (IFITs) proteins, and that depleting TFG resulted in a compromised antiviral state. Our study, therefore, identifies TFG as an essential component of the RLR-dependent type I IFN antiviral response.

Purification and characterization of a novel and conserved TPR-domain protein that binds both Hsp90 and Hsp70 and is expressed in all developmental stages of Leishmania major

Heat shock proteins (Hsps) are involved in several important aspects of the cell proteostasis. Hsp90 interacts with at least a tenth of the cell proteome helping a large number of proteins to fold correctly. Hsp90 function is modulated by several co-chaperones having TPR (tetratricopeptide repeat) domains that allow for interaction with the C-terminal MEEVD motif of the chaperone. Another important chaperone, Hsp70, has a C-terminal EEVD motif that binds to TPR. Leishmania is a protozoan that causes leishmaniasis, a neglected disease in humans and other animals. There is still no effective treatment for leishmaniasis, however the study of structure and function of the proteins of the parasite may generate potential targets for future therapeutic intervention studies. In this work, the genome of Leishmania major was searched for a novel TPR-domain gene, which is conserved in Leishmania. The recombinant protein, LmTPR, was produced in pure and folded state and was characterized by biophysical tools as a monomer with an elongated conformation. Studies in Leishmania major were also preformed to complement these in vitro experiments. Splice Leader RNA-seq analysis and Western blot indicated that the protein was expressed in all developmental stages of the parasite. Binding assays confirmed that both Hsp90 and Hsp70 bind specifically to LmTPR. Finally, sequence and structural predictions indicated a C-terminal region as a RPAP3 domain. Altogether, this study identified a novel TPR-domain co-chaperone of Hsp90 that is conserved and expressed in all developmental stages of Leishmania major.

Identification of a PLCE1‑regulated competing endogenous RNA regulatory network for esophageal squamous cell carcinoma.

Phospholipase C epsilon 1 (PLCE1) and the competing endogenous RNA (ceRNA) network are crucial for tumorigenesis and the progression of esophageal squamous cell carcinoma (ESCC). However, whether PLCE1 can regulate the ceRNA network in ESCC has not been clarified. In the present study, we aimed to identify the PLCE1-regulated ceRNA network and further elucidate the regulatory mechanisms by which ESCC is promoted. Microarray analysis was used to identify differentially expressed lncRNAs (DELs) and differentially expressed genes (DEGs) from three pairs of samples of PLCE-silenced Eca109 and control Eca109 cells. Next, the ceRNA regulatory network was established and visualized in Cytoscape, and functional enrichment analysis was performed to analyze DEGs from ceRNAs. Protein-protein interaction (PPI) networks among the DEGs were established by the STRING database to screen hub genes. Kaplan-Meier survival analysis was used to validate hub genes. Finally, PLCE1-related hub gene/lncRNA/miRNA axes were also constructed based on the ceRNA network. A total of 105 DELs and 346 DEGs were found to be dysregulated in the microarray data (|log2FC| >1.5, adjusted P<0.05). We constructed a PLCE1-regulated ceRNA network that incorporated 12 lncRNAs, 43 miRNAs, and 169 mRNAs. Functional enrichment analysis indicated that the DEGs might be associated with ESCC onset and development. A PPI network was established, and 9 hub genes [WD and tetratricopeptide repeats 1 (WDTC1), heat shock protein family A (Hsp70) member 5 (HSPA5), N-ethylmaleimide sensitive factor, vesicle fusing ATPase (NSF), fibroblast growth factor 2 (FGF2), cyclin dependent kinase inhibitor 1A (CDKN1A or P21), bone morphogenetic protein 2 (BMP2), complement C3 (C3), GM2 ganglioside activator (GM2A) and discs large MAGUK scaffold protein 4 (DLG4)] were determined from the network. Kaplan-Meier survival analysis validated four hub genes (BMP2, CDKN1A, GM2A, and DLG4) that were treated as prognostic factors. Ultimately, hub gene/lncRNA/miRNA subnetworks were obtained based on the 4 hub genes, 13 DEmiRNAs, and 10 DELs. In conclusion, the PLCE1-regulated ceRNA contributes to the onset and progression of ESCC and the underlying molecular mechanisms may provide insights into personalized prognosis and new therapies for ESCC patients.

Downregulation of interferon-induced protein with tetratricopeptide repeats 3 relieves the inflammatory response and myocardial fibrosis of mice with myocardial infarction and improves their cardiac function.

Myocardial infarction (MI) is a common cardiovascular disease with high morbidity and mortality. In this study, we explored the role of interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) in MI. MI was induced by ligation of the left anterior descending coronary artery. Lentivirus-mediated RNA interference of IFIT3 expression was performed by tail vein injection 72 h before MI modeling. Cardiac injury indexes and inflammatory response were examined 3 days after MI. Cardiac function indexes, infarct size, and cardiac fibrosis were assessed 4 weeks after MI. IFIT3 expression was upregulated in myocardial tissues at both 3 days and 4 weeks after MI. Knockdown of IFIT3 significantly relieved the myocardial injury, as evidenced by the decrease in serum levels of cTnI and CK-MB. In addition, IFIT3 knockdown significantly reduced the number of CD68+ macrophages and the levels of interleukin-1β, interleukin-6, and tumor necrosis factor-α, indicating that the inflammatory response was relieved. Moreover, IFIT3 silencing also significantly improved cardiac function and reduced infarct size, myocardial fibrosis, and collagen content in mice with MI. Mechanically, the present study showed that the activation of the mitogen-activated protein kinase (MAPK) pathway was observed in myocardial tissues of MI mice, which was blocked by IFIT3 knockdown, as indicated by the decreased phosphorylation of JNK, p-38, and ERK. Collectively, our results revealed the role of IFIT3 in the inflammatory response and myocardial fibrosis after MI, indicating that IFIT3 might be a potential target for MI treatment.

TTC36 inactivation induce malignant properties via Wnt-β-catenin pathway in gastric carcinoma.

Objective: Tetratricopeptide repeat (TRP)-mediated cofactor proteins are involved in a wide range of cancers. TTC36 is little studied member of TRP subfamily. This study aimed to investigate the role of TTC36 in human gastric carcinoma (GC) and explore the potential underlying mechanisms.Methods: The analysis of TTC36 differential expression in GC was conducted using data from TCGA and a human tissue microarray. And effects of TTC36 expression on the prognosis of patients with gastric carcinoma were analyzed using the data from the GEO database. Lentivirus was transfected into the cell lines of AGS and BGC823 to construct overexpression and knocked down TTC36 cell model respectively. The effect of TTC36 expression on the growth, apoptosis and cell cycle of cells was explored in vitro. Downstream molecules were detected by western blotting. GSEA predicted signal pathway and related proteins were then detected.Results: TTC36 expression in human GC tissues was found significantly lower than that in adjacent normal tissues and closely related to clinical prognosis. The overexpression of TTC36 notably inhibited tumor progression, cell cycle G1/S transfer and increased apoptosis in AGS cells. Conversely, the opposite effects were observed when TTC36 was suppressed in BGC823 cells. The expression of cleaved caspase3, Survivin, cyclin D1 and c-Myc were consistent with the phenotype in TTC36 operated GC cell lines. Intriguingly, GSEA analysis predicted Wnt-β-catenin pathway involved in TTC36 induced effects in GC cells, expression of β-catenin and downstream molecules such as TCF4, c-jun and pAKT were found negative related to TTC36 expression in GC cells. Notably, treatment with the Wnt/β-catenin inhibitor XAV939 dramatically attenuated the effects of TTC36 in GC cells.Conclusion: These results signify a critical role for TTC36 as a tumor suppressor in gastric carcinoma via regulating Wnt-β-catenin pathway.