Translationally Controlled Tumor Protein Gene Research Articles

Effects of Sarcoptes scabiei Translationally Controlled Tumor Protein (TCTP) on Histamine Release and Degranulation of KU812 Cells.

Scabies is a common parasitic dermatological infection worldwide that is often neglected. Scabies mites stimulate host inflammatory symptoms via secreted and excreted proteins, which induce basophil and mast cell degranulation and host histamine release. However, the mechanism of degranulation and histamine release is unclear. Moreover, the Sarcoptes scabiei translationally controlled tumor protein (TCTP) is predicted as an excreted protein, which may be involved in host inflammatory response regulation. First, we evaluated S. scabiei TCTP gene (SsTCTP) transcription in larvae, nymphs, and adults by qRT-PCR, and SsTCTP transcription was highest in larvae, followed by nymphs. Second, we found that the S. scabiei TCTP recombinant protein (rSsTCTP) promoted mice histamine release in vivo by Evans blue Miles assay. Therefore, to further explore the possible role of S. scabiei TCTP in host inflammatory response regulation, we established a degranulation model of KU812 cells. The results of the degranulation model suggested that rSsTCTP could induce enhanced degranulation of KU812 cells and increase the secretion of histamine and the expression of IL-4, IL-6, and IL-13 in vitro. In conclusion, we speculate that scabies mites could stimulate host histamine release and Th2 response by excreting S. scabiei TCTP.

Orchestration of myeloid-derived suppressor cells in the tumor microenvironment by ubiquitous cellular protein TCTP released by tumor cells.

One of most challenging issues in tumor immunology is a better understanding of the dynamics in the accumulation of myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment (TIME), as this would lead to the development of new cancer therapeutics. Here, we show that translationally controlled tumor protein (TCTP) released by dying tumor cells is an immunomodulator crucial to full-blown MDSC accumulation in the TIME. We provide evidence that extracellular TCTP mediates recruitment of the polymorphonuclear MDSC (PMN-MDSC) population in the TIME via activation of Toll-like receptor-2. As further proof of principle, we show that inhibition of TCTP suppresses PMN-MDSC accumulation and tumor growth. In human cancers, we find an elevation of TCTP and an inverse correlation of TCTP gene dosage with antitumor immune signatures and clinical prognosis. This study reveals the hitherto poorly understood mechanism of the MDSC dynamics in the TIME, offering a new rationale for cancer immunotherapy.

Comprehensive analysis of Translationally Controlled Tumor Protein (TCTP) provides insights for lineage-specific evolution and functional divergence.

BackgroundTranslationally controlled tumor protein (TCTP) is a conserved, multifunctional protein involved in numerous cellular processes in eukaryotes. Although the functions of TCTP have been investigated sporadically in animals, invertebrates, and plants, few lineage-specific activities of this molecule, have been reported. An exception is in Arabidopsis thaliana, in which TCTP (AtTCTP1) functions in stomatal closuer by regulating microtubule stability. Further, although the development of next-generation sequencing technologies has facilitated the analysis of many eukaryotic genomes in public databases, inter-kingdom comparative analyses using available genome information are comparatively scarce.MethodologyTo carry out inter-kingdom comparative analysis of TCTP, TCTP genes were identified from 377 species. Then phylogenetic analysis, prediction of protein structure, molecular docking simulation and molecular dynamics analysis were performed to investigate the evolution of TCTP genes and their binding proteins.ResultsA total of 533 TCTP genes were identified from 377 eukaryotic species, including protozoa, fungi, invertebrates, vertebrates, and plants. Phylogenetic and secondary structure analyses reveal lineage-specific evolution of TCTP, and inter-kingdom comparisons highlight the lineage-specific emergence of, or changes in, secondary structure elements in TCTP proteins from different kingdoms. Furthermore, secondary structure comparisons between TCTP proteins within each kingdom, combined with measurements of the degree of sequence conservation, suggest that TCTP genes have evolved to conserve protein secondary structures in a lineage-specific manner. Additional tertiary structure analysis of TCTP-binding proteins and their interacting partners and docking simulations between these proteins further imply that TCTP gene variation may influence the tertiary structures of TCTP-binding proteins in a lineage-specific manner.ConclusionsOur analysis suggests that TCTP has undergone lineage-specific evolution and that structural changes in TCTP proteins may correlate with the tertiary structure of TCTP-binding proteins and their binding partners in a lineage-specific manner.

Promoter of shrimp translationally controlled tumor protein (TCTP) gene has higher activity than cytomegavirus (CMV) promoter in driving foreign genes to express in primary shrimp cells

Abstract Active promoters are urgently needed for shrimp cells which are hard to be immortalized. Translationally controlled tumor protein (TCTP) is a widely and abundantly expressed, growth-related protein. In this study we successfully isolated the promoter (Ptctp) and ORF (= open reading frame) of the TCTP gene from Litopenaeus vannamei and analysed the promoter activity of Ptctp and the growth-promoting effects of over-expressed TCTP protein in the primary Oka organ (lymphoid tissue) cells. It was found, that cytomegavirus (CMV) promoter, highly active in mammalian and fish cells, had a driving activity too low to be detected in shrimp cells. However, shrimp Ptctp had a higher driving activity than Pcmv in the shrimp cells and an obvious fluorescent signal was observed in the shrimp cells transfected by a Pcmv-Ptctp-driven plasmid. However, no obvious growth-promoting effects were observed in the eGFP/TCTP [eGFP = enhanced green fluorescent protein] or TCTP-transfected shrimp cells possibly due to the relatively low expression efficiency.

Characterization, homology modeling and expression of the putative translationally controlled tumor protein gene from giant river prawn Macrobrachium rosenbergii (de Man)

The translationally controlled tumor protein (TCTP) is a multifunctional protein that promotes cell growth and viability. The TCTP has been associated with shrimp immune response by high levels being detected during the onset of white spot syndrome virus (WSSV) infection. The present study characterized the putative TCTP gene from the giant river prawn M. rosenbergii and investigated a putative function of this protein in response to Aeromonas hydrophila injection. A full-length cDNA sequence was obtained by assembly of short reads of RNA sequences from a 454-transcriptome dataset of river prawns and was designated as putative MrTCTP. The three-dimensional (3D) structure of MrTCTP was constructed using a homology modeling approach to predict the functional properties of the protein. Expression of MrTCTP was assessed in hepatopancreas tissues 96 h after injection using quantitative real-time polymerase chain reaction. The 704 bp full length cDNA of MrTCTP contained an open reading frame of 507 bp encoding a polypeptide of 168 amino acid residues. The conserved signature sequences of TCTP, I45-D55 and F123Y145 shared 61–92.3% similarity with the previously identified TCTP. The quality assessment and parameter validation indicated that the putative MrTCTP modeled structure was reliable. Expression levels of MrTCTP were significantly different (p < 0.05) between the control and infected prawns during 16–48 h. Transcript levels significantly increased 1.21-fold at 16 h to 2.26-fold at 48 h. The results suggested that the putative MrTCTP may be involved in the anti-apoptotic mechanism of river prawn.

CRISPR disruption of TCTP gene impaired normal development in the silkworm Bombyx mori.

The translationally controlled tumor protein (TCTP) is a highly conserved and multifunctional protein with activities ranging from cytoskeletal regulation to transcription regulation in numerous organisms. In insects, TCTP is essential for cell growth and proliferation. Recently, TCTP has been reported to affect the innate intestinal immune pathway in the Bombyx mori silkworm, a lepidopteran model insect. However, the comprehensive physiological roles of TCTP in the silkworm remain poorly understood. Here, we performed functional analysis of BmTCTP by using a binary transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeat/RNA‐guided CRISPER‐associated protein 9 nucleases) system. Disruption of BmTCTP led to developmental arrestment and subsequent lethality in third instar larvae. Histological analysis revealed that growth impairment originated from decreased cell size, and the proliferation and differentiation of intestinal epithelial cells were also affected. RNA‐seq analysis revealed that genes involved in carbohydrate metabolism, lipid metabolism and digestive system pathways were significantly affected by BmTCTP depletion. Together, the results demonstrated that BmTCTP plays a key role in controlling larval growth and development.

Structural divergence of plant TCTPs.

The Translationally Controlled Tumor Protein (TCTP) is a highly conserved protein at the level of sequence, considered to play an essential role in the regulation of growth and development in eukaryotes. However, this function has been inferred from studies in a few model systems, such as mice and mammalian cell lines, Drosophila and Arabidopsis. Thus, the knowledge regarding this protein is far from complete. In the present study bioinformatic analysis showed the presence of one or more TCTP genes per genome in plants with highly conserved signatures and subtle variations at the level of primary structure but with more noticeable differences at the level of predicted three-dimensional structures. These structures show differences in the “pocket” region close to the center of the protein and in its flexible loop domain. In fact, all predictive TCTP structures can be divided into two groups: (1) AtTCTP1-like and (2) CmTCTP-like, based on the predicted structures of an Arabidopsis TCTP and a Cucurbita maxima TCTP; according to this classification we propose that their probable function in plants may be inferred in principle. Thus, different TCTP genes in a single organism may have different functions; additionally, in those species harboring a single TCTP gene this could carry multiple functions. On the other hand, in silico analysis of AtTCTP1-like and CmTCTP-like promoters suggest that these share common motifs but with different abundance, which may underscore differences in their gene expression patterns. Finally, the absence of TCTP genes in most chlorophytes with the exception of Coccomyxa subellipsoidea, indicates that other proteins perform the roles played by TCTP or the pathways regulated by TCTP occur through alternative routes. These findings provide insight into the evolution of this gene family in plants.

Molecular cloning, expression profiles and characterization of a novel translationally controlled tumor protein in rubber tree (Hevea brasiliensis)

The translationally controlled tumor protein (TCTP) is a multi-functioning protein that carries out vital roles in various life processes. In this study, a new TCTP gene, designated as HbTCTP1, was isolated in Hevea brasiliensis. The full-length complementary DNA (cDNA) of HbTCTP1 contained a maximum open reading frame (ORF) of 507base pair (bp) encoding 168 amino acids. The sequence comparison showed that the deduced HbTCTP1 indicated high identities to plant TCTP proteins, and clustered in the dicot cluster of plant TCTPs. Although HbTCTP1 and human TCTP proteins did not parallel in overall sequence similarity, they indicated highly similar 3D structures with a nearly identical spatial organization of α-helices, β-sheets, and coil regions. Real time reverse-transcription PCR (RT-PCR) analyses showed that HbTCTP1 was expressed throughout different tissues and developmental stages of leaves. Besides being related to tapping panel dryness (TPD), the HbTCTP1 transcripts were regulated by various treatments, including drought, low temperature, high salt, ethrel (ET), wounding, H2O2, and methyl jasmonate (Me-JA) treatments. The recombinant HbTCTP1 fusion protein was shown to protect supercoiled plasmid DNA from damages induced by metal-catalyzed generation of reactive oxygen species. The 45Ca2+-overlay assay showed that HbTCTP1 was a calcium-binding protein. Our results are greatly helpful in understanding the molecular characterization and expression profiles of HbTCTP1, and lay the foundation for further analyzing the function of HbTCTP1 in rubber tree.

Overexpression of Arabidopsis translationally controlled tumor protein gene AtTCTP enhances drought tolerance with rapid ABA-induced stomatal closure.

Translationally controlled tumor protein (TCTP), also termed P23 in human, belongs to a family of calcium- and tubulin-binding proteins, and it is generally regarded as a growth-regulating protein. Recently, Arabidopsis TCTP (AtTCTP) has been reported to function as an important growth regulator in plants. On the other hand, plant TCTP has been suggested to be involved in abiotic stress signaling such as aluminum, salt, and water deficit by a number of microarray or proteomic analyses. In this study, the biological functions of AtTCTP were investigated by using transgenic Arabidopsis plants overexpressing AtTCTP. Interestingly, AtTCTP overexpression enhanced drought tolerance in plants. The expression analysis showed that AtTCTP was expressed in guard cells as well as in actively growing tissues. Physiological studies of the overexpression lines showed increased ABA- and calcium-induced stomatal closure ratios and faster stomatal closing responses to ABA. Furthermore, in vitro protein-protein interaction analysis confirmed the interaction between AtTCTP and microtubules, and microtubule cosedimentation assays revealed that the microtubule binding of AtTCTP increased after calcium treatment. These results demonstrate that the overexpression of AtTCTP confers drought tolerance to plants by rapid ABA-mediated stomatal closure via the interaction with microtubules in which calcium binding enhances the interaction. Collectively, the present results suggest that the plant TCTP has molecular properties similar to animal TCTPs, such as tubulin- and calcium-binding, and that it functions in ABA-mediated stomatal movement, in addition to regulating the growth of plants.

Cloning and prokaryotic expression of translationally controlled tumor protein (TaTCTP) gene from wheat and preparation of antiserum

The translationally controlled tumor protein (TCTP) is a multi-functioning protein that performs vital roles, particularly in various complicated life processes. To further understand the biological function of translationally controlled tumor protein (TaTCTP) gene from wheat, total RNA was isolated from wheat leaves and then TaTCTP gene was amplified by PCR after reverse transcription by using the anchored primers oligo(dT)18. And then TaTCTP gene was connected into PMD19-T vector for sequence. The recombinant expression vector (pET30a-GST-TaTCTP-His) was constructed and transformed into E. coli strain Rosetta (DE3) subsequently, then a proximate 20 kDa protein in Rosetta (DE3) was expressed and characterized by SDS-PAGE. Moreover, TaTCTP fusion protein was purified by Ni-NTA affinity chromatography from recombinant bacterial lysate and was used to immunize rabbit to produce polyclonal antibody. The titer and specificity of the anti-TaTCTP antibody were successfully detected by indirect ELISA and Western blot analysis.

A Novel TCTP Gene From the Crustacean Eriocheir sinensis: Possible Role Involving Metallic Cu2+ Stress

The translationally controlled tumor protein (TCTP), which was initially described as a growth-related protein, primarily expresses proteins for numerous biological processes in eukaryotes. In the present study, the TCTP gene in the Chinese mitten crab Eriocheir sinensis (Es-TCTP) was identified from tissues of the hepatopancreas, by the construction of a cDNA library and the rapid amplification of cDNA ends. The results showed that the full length of Es-TCTP cDNA comprises 727 nucleotides, with an open reading frame of 507 bp that encodes 168 amino acid residues. The deduced amino acid sequence of Es-TCTP has a 51%-92% similarity to TCTPs of other eukaryotic species. The mRNA transcripts of Es-TCTP were detected by using RT-PCR for all tissues, with the highest level occurring in the hepatopancreas. In addition, temporal expression of the Es-TCTP was measured to analyze the role of Es-TCTP in an acute stress condition after CuSO(4) treatment. The expression of the Es-TCTP transcripts in hepatopancreas had no significant difference at the first 4 h, but increased significantly after 8 h, peaked at 24 h (4.3-fold higher than the control), and was still high after 72 h. Our results indicate that Es-TCTP is an acute-phase protein that is involved in Cu(2+) stress, with a possible anti-stress function in invertebrates.

Gene silencing of translationally controlled tumor protein (TCTP) by siRNA inhibits cell growth and induces apoptosis of human prostate cancer cells

Translationally controlled tumor protein (TCTP) is a novel anti apoptotic protein which is highly expressed in several cancer cell types including prostate cancer. However, studies investigating the role of TCTP in prostate cancer are scarce. Therefore, in this study we evaluated the effect of small interference RNA (siRNA) based knocking down of TCTP gene in prostate cancer cells. Cell proliferation and apoptosis were evaluated. Our results showed that TCTP is highly expressed in LNCaP cells compared to normal prostate epithelial cells. Transfection with TCTP siRNA specifically and drastically reduced the expression of both mRNA and protein levels of TCTP in LNCaP cells. The decreased expression of TCTP was associated with decreased viability of LNCaP cells. Further analysis of the transfected LNCaP cells showed that they undergo apoptosis via caspase-8 and caspase-3 dependent pathways. Results presented herein suggest a potential therapeutic application for prostate cancer by targeting TCTP gene using an siRNA approach.

Novel translationally controlled tumor protein homologue in the buccal gland secretion of Lampetra japonica

We have cloned a homologue of the translationally controlled tumor protein (TCTP) from the buccal gland of Lampetra japonica according to information from a cDNA library and primary analysis of expressed sequence tags. Sequence analysis of L. japonica TCTP showed that it had two signature regions of high sequence homology termed TCTP-1 and TCTP-2, respectively. TCTP is highly conserved in evolution. It showed more than 40% identification similarities with parasite TCTPs that had effect on immune responses of host. Phylogeny of 31 TCTP sequences showed that lamprey was closer to jawed vertebrates than to Amphioxus and was a sister group of gnathostomes. TCTP gene from L. japonica was expressed in a pET23b vector and purified by using His Bind affinity chromatography. Polyclonal antibody to recombinant protein was generated in New Zealand Rabbit. Immunoblot analysis to localize the recombinant protein in buccal gland secretion proves that recombinant TCTP is a secretion protein, which may be secreted through a non-classical secretion pathway. A characterization study shows that recombinant TCTP has histamine-releasing function in vitro. It mediated histamine release from rat basophilic leukemia (RBL-2H3) cells. TCTP links both the innate and the adaptive immune responses by modulating the secretion of cytokines from mast cells, basophils, eosinophils, and T and B lymphocytes. These may indicate a potential role of TCTP in the inflammatory process and immune regulation between L. japonica and host.

Genomic organization of the translationally controlled tumor protein (TCTP) gene from shrimp Marsupenaeus japonicus

The shrimp translationally controlled tumor protein (TCTP) gene was previously found up-regulated in response to viral infection. In order to further understand the transcriptional regulation of TCTP gene expression under viral challenge, the 5'-flanking promoter region of the TCTP gene in shrimp Marsupenaeus japonicus (Mj-TCTP) was cloned by using genomic walking method. The sequence shows that this region contains many important potential binding sites for transcription factors, such as activator protein1 (AP1), cAMP response element binding protein (CREB), Ets like transcription factor-1 (Elk-1), suggesting its expression is highly regulated. The promoter activity of this 5'-flanking region was confirmed by fusing to the enhanced green fluorescence protein (EGFP) gene and testing in Trichoplusia ni High Five cell line. Along with the introns and exons cloned in this study, the Mj-TCTP DNA sequence was reported here as the first TCTP gene with complete genomic organization in crustaceans.

Expression of translationally controlled tumor protein mRNA in human colon cancer

The translationally controlled tumor protein (TCTP) was first identified as a tumor-associated protein. Here we described the expression profile of TCTP mRNA using a panel of normal human tissues. We also investigated the cell-cycle-dependent expression of the TCTP mRNA using synchronized cultures of Saccahromyces cerevisiae. The TCTP gene was constantly expressed through the yeast cell cycle. The expression of TCTP mRNA from three human colon carcinoma cell lines and their tumors xenografted in a nude mouse was studied. Our data imply that the TCTP is a housekeeping gene and that its deregulation may be involved in the tumor development of colon carcinoma.

Structure of the promoter and complete sequence of the gene coding for the rabbit translationally controlled tumor protein (TCTP) P23.

We have isolated genomic recombinants containing the complete gene coding for the rabbit translationally controlled tumor protein (TCTP), also known as histamine-releasing factor (HRF) P23. The gene is organized into five introns and six exons and its total length amounts to 3819 nucleotides. All intron/exon boundaries are in accordance with the GT/AG rule. Transcription of the gene generates two mRNAs of 843 and 1163 nucleotides differing in the length of their 3'-untranslated regions. They are formed by alternative polyadenylation. The transcription initiation site has been determined by comparison of sequences of the gene and several processed TCTP pseudogenes. The full-length 5'-untranslated region comprises 116 nucleotides and starts with an oligopyrimidine tract important for translational regulation. Additionally 1.2 kb of the 5'-flanking promoter region has been sequenced. The promoter contains a TATA box at -30 and potential binding sites for transcription factors such as stimulating protein 1 (Sp1), nuclear factor 1 (NF1), activator protein 1 (AP1), c-Ets1, cAMP-response element (CP2), myeloid-specific zinc finger protein 1 (MZF1) and others. For functional analysis 5'-flanking sequences up to -918 were fused to the chloramphenicol acetyltransferase (CAT) gene and tested using a rabbit aortic smooth-muscle cell line by cell transfection and CAT assays. The results confirm that the analyzed gene is the actively transcribed TCTP gene.