Chaperonin Protein Research Articles

Improving soluble recombinant SARS-CoV-2 papain-like protease production in Escherichia coli through chaperonin and maltose-binding protein tag: purification and kinetic characterization

Although COVID-19 is now becoming endemic, SARS-CoV-2 persists potential jeopardy to clinically vulnerable populations. Hence, further study is still necessary to discover novel antiviral agents against SARS-CoV-2 for proactive preparedness. SARS-CoV-2 papain-like protease (PL Pro) is a target enzyme for searching anti-Covid candidates. Our prior study revealed the major formation of inclusion bodies during PL Pro expression in E. coli RIPL. In this study, we tried using chaperonin in the E. coli Arctic Express system and both codon optimization and maltose-binding protein (MBP) fusion protein to make PL Pro more soluble. Recombinant PL Pro encoded on the pET21d(+) plasmid was expressed in E. coli Arctic express. However, the soluble protein yield remained low and unstable due to suboptimal codon usage in the insert gene. Whereas, fusion of the MBP protein with optimized codon of PL Pro enhanced the enzyme expression and solubility. Recombinant PL Pro cleaved the linker between MBP and PL Pro, which served as a cleavage site recognized by PL Pro (LKGG↓A). The purified enzyme from a 200-mL culture generated 1 mL of pure PL Pro enzyme at a 1.913 mg/mL concentration. It exhibited favorable activity against the Z-RLRGG-AMC substrate, with a Km value of 33.40 μM and a Vmax of 5.10 RFU/min.

Heterozygous de novo variants in HSPD1 cause hypomyelinating leukodystrophy through impaired HSP60 oligomerisation

IntroductionHypomyelinating leukodystrophies are a group of genetic disorders, characterised by severe permanent myelin deficiency. Their clinical features include developmental delay with or without neuroregression, nystagmus, central hypotonia, progressing to spasticity...

Oxysterol Induces Expression of 60 kDa Chaperone Protein on Cell Surface of Microglia.

Microglia, essential immune cells in the brain, play crucial roles in neuroinflammation by performing various functions such as neurogenesis, synaptic pruning, and pathogen defense. These cells are activated by inflammatory factors like β-amyloid (Aβ) and oxysterols, leading to morphological and functional changes, including the secretion of inflammatory cytokines and the upregulation of MHC class II molecules. This study focused on identifying specific markers for microglial activation, with a particular emphasis on the roles of oxysterols in this process. We used the HMC3 human microglial cell line to investigate the induction of heat shock protein 60 (HSP60), a chaperonin protein by oxysterols, specifically in the presence of 25-hydroxycholesterol (25OHChol) and 27-hydroxycholesterol (27OHChol). Our findings obtained by the proteomics approach revealed that these oxysterols significantly increased HSP60 expression on microglial cells. This induction was further confirmed using Western blot analysis and immunofluorescence microscopy. Additionally, Aβ1-42 also promoted HSP60 expression, indicating its role as a microglial activator. HSP60 involved in protein folding and immune modulation was identified as a potential marker for microglial activation. This study underscores the importance of HSP60 in the inflammatory response of microglia, suggesting its utility as a target for new therapeutic approaches in neuroinflammatory diseases such as Alzheimer's disease (AD).

The Influence of Chitosan Derivatives in Combination with Bacillus subtilis Bacteria on the Development of Systemic Resistance in Potato Plants with Viral Infection and Drought.

Viral diseases of potatoes are among the main problems causing deterioration in the quality of tubers and loss of yield. The growth and development of potato plants largely depend on soil moisture. Prevention strategies require comprehensive protection against pathogens and abiotic stresses, including modeling the beneficial microbiome of agroecosystems combining microorganisms and immunostimulants. Chitosan and its derivatives have great potential for use in agricultural engineering due to their ability to induce plant immune responses. The effect of chitosan conjugate with caffeic acid (ChCA) in combination with Bacillus subtilis 47 on the transcriptional activity of PR protein genes and changes in the proteome of potato plants during potato virus Y (PVY) infection and drought was studied. The mechanisms of increasing the resistance of potato plants to PVY and lack of moisture are associated with the activation of transcription of genes encoding PR proteins: the main protective protein (PR-1), chitinase (PR-3), thaumatin-like protein (PR-5), protease inhibitor (PR-6), peroxidase (PR-9), and ribonuclease (PR-10), as well as qualitative and quantitative changes in the plant proteome. The revealed activation of the expression of marker genes of systemic acquired resistance and induced systemic resistance under the influence of combined treatment with B. subtilis and chitosan conjugate indicate that, in potato plants, the formation of resistance to viral infection in drought conditions proceeds synergistically. By two-dimensional electrophoresis of S. tuberosum leaf proteins followed by MALDI-TOF analysis, 10 proteins were identified, the content and composition of which differed depending on the experiment variant. In infected plants treated with ChCA, the synthesis of proteinaceous RNase P 1 and oxygen-evolving enhancer protein 2 was enhanced in conditions of normal humidity, and 20 kDa chaperonin and TMV resistance protein N-like was enhanced in conditions of lack of moisture. The virus coat proteins were detected, which intensively accumulated in the leaves of plants infected with potato Y-virus. ChCA treatment reduced the content of these proteins in the leaves, and in plants treated with ChCA in combination with Bacillus subtilis, viral proteins were not detected at all, both in conditions of normal humidity and lack of moisture, which suggests the promising use of chitosan derivatives in combination with B. subtilis bacteria in the regulation of plant resistance.

Identification of moonlighting adhesins of highly-adhesive Lactobacillus plantarum PO23 isolated from the intestine of Paralichthys olivaceus

At present, feeding probiotics has been an effective method to prevent and control enterogenous diseases of cultured fish. It is very important and urgent to study the molecular mechanism of the adhesion and colonization of probiotics to improve the use efficiency accurately. In our previous research, highly-adhesive Lactobacillus plantarum PO23 (LP-PO23) was isolated from the intestine of olive flounder (Paralichthys olivaceus). To clarify the specific adhesion of LP-PO23 to intestinal epithelial cells (IECs), six adhesins, d-lactate dehydrogenase (LDH), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), elongation factor Tu (EF-Tu), enolase (ENO), 60 kDa chaperonin (Cpn60) and chaperone protein DnaK (DnaK) were identified as moonlighting proteins by far-Western blot and liquid chromatography-mass spectrometry in this research. The recombinant proteins rLDH, rGAPDH, rEF-Tu, rENO, rCpn60 and rDnak containing FLAG tag were expressed and employed as antigens to produce mouse polyclonal antibodies (pAbs). Using pAbs, Western blot, immunofluorescence (IF) and immunoelectron microscopy (IEM) were carried out to confirm the surface distribution of six adhesins on LP-PO23 cells respectively. Finally, the specific binding of rLDH, rGAPDH, rEF-Tu, rENO, rCpn60 and rDnak to IECs was verified by IF. In binding inhibition assay, the mouse anti-GAPDH pAb displayed the strongest inhibition activity among six pAbs. These results confirmed that six moonlighting adhesins participated in the specific adhesion of LP-PO23 to IECs and would facilitate understanding the adhesion mechanism of lactobacilli.

Metatranscriptome and Resistome of the Endodontic Microbiome

IntroductionIn this study, we used metatranscriptomics for the first time to investigate microbial composition, functional signatures, and antimicrobial resistance gene expression in endodontic infections. MethodsRoot canal samples were collected from ten teeth, including five primary and five persistent/secondary endodontic infections. RNA from endodontic samples was extracted, and RNA sequencing was performed on a NovaSeq6000 system (Illumina). Taxonomic analysis was performed using the Kraken2 bacterial database. Then, sequences with a taxonomic classification were annotated against the Universal Protein Knowledgebase for functional annotation and the Comprehensive Antibiotic Resistance Database for AR-like gene identification. ResultsProteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria represented the dominant phyla, whereas Fusobacteria, Spirochetes, and Synergistetes were among the nondominant phyla. The top ten species were mainly represented by obligate (or quasiobligate) anaerobes, including Gram-negative (eg, Capnocytophaga sp. oral taxon 323, Fusobacterium nucleatum, Prevotella intermedia, Prevotella oris, Tannerella forsythia, and Tannerella sp. oral taxon HOT−286) and Gram-positive species (eg, Olsenella uli and Parvimonas micra). Transcripts encoding moonlighting proteins (eg, glycolytic proteins, translational elongation factors, chaperonin, and heat shock proteins) were highly expressed, potentially affecting bacterial adhesion, biofilm formation, host defense evasion, and inflammation induction. Endodontic bacteria expressed genes conferring resistance to antibiotic classes commonly used in dentistry, with a high prevalence and expression of tetracycline and lincosamide resistance genes. Antibiotic efflux and antibiotic target alteration/protection were the main resistance mechanisms. ConclusionsMetatranscriptomics revealed the activity of potential endodontic pathogens, which expressed putative virulence factors and a wide diversity of genes potentially involved in AR.

Chaperonin TRiC/CCT subunit CCT7 is involved in the replication of canine parvovirus in F81 cells.

Canine parvovirus (CPV) is one of the most common lethal viruses in canines. The virus disease is prevalent throughout the year, with high morbidity and mortality rate, causing serious harm to dogs and the dog industry. Previously, yeast two hybrid method was used to screen the protein chaperonin containing TCP-1 (CCT7) that interacts with VP2. However, the mechanism of interactions between CCT7 and VP2 on CPV replication remains unclear. In this study, we first verified the interaction between CCT7 and viral VP2 proteins using yeast one-to-one experiment and co-immunoprecipitation (CoIP) experiment. Laser confocal microscopy observation showed that CCT7 and VP2 were able to co-localize and were mostly localized in the cytoplasm. In addition, the study of VP2 truncated mutant found that the interaction region of VP2 with CCT7 was located between amino acids 231 and 320. Cycloheximide (CHX) chase experiments showed that CCT7 can improve the stability of VP2 protein. After further regulation of CCT7 expression in F81 cells, it was found that the expression level of VP2 protein was significantly reduced after knocking down CCT7 expression by RNA interference (RNAi) or HSF1A inhibitor, and increased after overexpressing host CCT7. The study reveals the role of VP2 interacting protein CCT7 in the replication process of CPV, which could provide a potential target for the prevention and control of CPV.

Rapid and accurate taxonomic classification of cpn60 amplicon sequence variants

The “universal target” region of the gene encoding the 60 kDa chaperonin protein (cpn60, also known as groEL or hsp60) is a proven sequence barcode for bacteria and a useful target for marker gene amplicon-based studies of complex microbial communities. To date, identification of cpn60 sequence variants from microbiome studies has been accomplished by alignment of queries to a reference database. Naïve Bayesian classifiers offer an alternative identification method that provides variable rank classification and shorter analysis times. We curated a set of cpn60 barcode sequences to train the RDP classifier and tested its performance on data from previous human microbiome studies. Results showed that sequences accounting for 79%, 86% and 92% of the observations (read counts) in saliva, vagina and infant stool microbiome data sets were classified to the species rank. We also trained the QIIME 2 q2-feature-classifier on cpn60 sequence data and demonstrated that it gives results consistent with the standalone RDP classifier. Successful implementation of a naïve Bayesian classifier for cpn60 sequences will facilitate future microbiome studies and open opportunities to integrate cpn60 amplicon sequence identification into existing analysis pipelines.

Mycobacterial chaperonins in cellular proteostasis: Evidence for chaperone function of Cpn60.1 and Cpn60.2-mediated protein folding.

Mycobacterium tuberculosis encodes two chaperonin proteins, MtbCpn60.1 and MtbCpn60.2, that share substantial sequence similarity with the Escherichia coli chaperonin, GroEL. However, unlike GroEL, MtbCpn60.1 and MtbCpn60.2 purify as lower-order oligomers. Previous studies have shown that MtbCpn60.2 can functionally replace GroEL in E. coli, while the function of MtbCpn60.1 remained an enigma. Here, we demonstrate the molecular chaperone function of MtbCpn60.1 and MtbCpn60.2, by probing their ability to assist the folding of obligate chaperonin clients, DapA, FtsE and MetK, in an E. coli strain depleted of endogenous GroEL. We show that both MtbCpn60.1 and MtbCpn60.2 support cell survival and cell division by assisting the folding of DapA and FtsE, but only MtbCpn60.2 completely rescues GroEL-depleted E. coli cells. We also show that, unlike MtbCpn60.2, MtbCpn60.1 has limited ability to support cell growth and proliferation and assist the folding of MetK. Our findings suggest that the client pools of GroEL and MtbCpn60.2 overlap substantially, while MtbCpn60.1 folds only a small subset of GroEL clients. We conclude that the differences between MtbCpn60.1 and MtbCpn60.2 may be a consequence of their intrinsic sequence features, which affect their thermostability, efficiency, clientomes and modes of action.

ATP-Responsive Nanoparticles Covered with Biomolecular Machine "Chaperonin GroEL".

Herein, we report an ATP-responsive nanoparticle (GroELNP) whose surface is fully covered with the biomolecular machine "chaperonin protein GroEL". GroELNP was synthesized by DNA hybridization between a gold NP with DNA strands on its surface and GroEL carrying complementary DNA strands at its apical domains. The unique structure of GroELNPs was visualized by transmission electron microscopy including under cryogenic conditions. The immobilized GroEL units retain their machine-like function and enable GroELNPs to capture denatured green fluorescent protein and release it in response to ATP. Interestingly, the ATPase activity of GroELNPs per GroEL was 4.8 and 4.0 times greater than those of precursor cysGroEL and its DNA-functionalized analogue, respectively. Finally, we confirmed that GroELNPs could be iteratively extended to double-layered (GroEL)2NPs.

ATP‐Responsive Nanoparticles Covered with Biomolecular Machine “Chaperonin GroEL”

AbstractHerein, we report an ATP‐responsive nanoparticle (GroELNP) whose surface is fully covered with the biomolecular machine “chaperonin protein GroEL”. GroELNP was synthesized by DNA hybridization between a gold NP with DNA strands on its surface and GroEL carrying complementary DNA strands at its apical domains. The unique structure of GroELNP was visualized by transmission electron microscopy including under cryogenic conditions. The immobilized GroEL units retain their machine‐like function and enable GroELNP to capture denatured green fluorescent protein and release it in response to ATP. Interestingly, the ATPase activity of GroELNP per GroEL was 4.8 and 4.0 times greater than those of precursor cysGroEL and its DNA‐functionalized analogue, respectively. Finally, we confirmed that GroELNP could be iteratively extended to double‐layered NP.

Anchovy’s protein as a potential precursor of Angiotensin-I Converting Enzyme (ACE) inhibitory peptide and Dipeptidyl Peptidase-IV (DPP-IV) inhibitory peptide by an in silico approach

Protein from fish is known as the precursor of biologically active peptides that exert various health benefits such as antihypertensive, antitumor, and immunomodulatory properties. Hence, this study aimed to perform an extraction of anchovy, LC-MS/MS analysis and in silico evaluation of the major proteins in anchovies as potential precursors of biologically active peptides in addition to determining whether such peptides can be released by selected proteolytic enzymes. Anchovy was subjected to protein extraction followed by total soluble protein concentration determination using Bradford assay. The sample was subjected to in-solution trypsin digestion, which was then analysed by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). A bioinformatic approach by PEAKS Studio was used to identify the protein. A total of four anchovy’s proteins which are myosin light chain 1, V(D)J recombination-activating protein, 60 kDa chaperonin and heat shock protein 90AA1 were identified. Then, the identified proteins were subjected to in silico approach using the BIOPEP database. The biological potential of the theoretically released angiotensin-I converting enzyme (ACE) inhibitory peptides and dipeptidyl peptidase (DPP)-IV inhibitory peptides were predicted by determining the frequency of occurrence of fragments with a given activity. 60 kDa chaperonin and heat shock protein 90AA1 predicted the highest number of biological activities for ACE inhibitory peptides (284 and 264 fragments) and DPP-IV inhibitory peptides (395 and 409 fragments). The most promising enzyme for the generation of bioactive peptides from anchovy protein was anticipated to be pepsin (pH > 2), which theoretically released a high number of DPP-IV inhibitory peptides and ACE inhibitory peptides through the action of in silico proteolysis. Overall, this work highlighted that anchovy protein could be a promising precursor of bioactive peptides that have ACE and DPP-IV inhibitory activities for developing functional food or nutraceutical products.

Chaperonin: Co-chaperonin Interactions.

Co-chaperonins function together with chaperonins to mediate ATP-dependent protein folding in a variety of cellular compartments. Chaperonins are evolutionarily conserved and form two distinct classes, namely, group I and group II chaperonins. GroEL and its co-chaperonin GroES form part of group I and are the archetypal members of this family of protein folding machines. The unique mechanism used by GroEL and GroES to drive protein folding is embedded in the complex architecture of double-ringed complexes, forming two central chambers that undergo conformational rearrangements that enable protein folding to occur. GroES forms a lid over the chamber and in doing so dislodges bound substrate into the chamber, thereby allowing non-native proteins to fold in isolation. GroES also modulates allosteric transitions of GroEL. Group II chaperonins are functionally similar to group I chaperonins but differ in structure and do not require a co-chaperonin. A significant number of bacteria and eukaryotes house multiple chaperonin and co-chaperonin proteins, many of which have acquired additional intracellular and extracellular biological functions. In some instances, co-chaperonins display contrasting functions to those of chaperonins. Human HSP60 (HSPD) continues to play a key role in the pathogenesis of many human diseases, in particular autoimmune diseases and cancer. A greater understanding of the fascinating roles of both intracellular and extracellular Hsp10 on cellular processes will accelerate the development of techniques to treat diseases associated with the chaperonin family.

Subretinal gene therapy delays vision loss in a Bardet-Biedl Syndrome type 10 mouse model

Blindness in Bardet-Biedl syndrome (BBS) is caused by dysfunction and loss of photoreceptor cells in the retina. BBS10, mutations of which account for approximately 21% of all BBS cases, encodes a chaperonin protein indispensable for the assembly of the BBSome, a cargo adaptor important for ciliary trafficking. The loss of BBSome function in the eye causes a reduced light sensitivity of photoreceptor cells, photoreceptor ciliary malformation, dysfunctional ciliary trafficking, and photoreceptor cell death. Cone photoreceptors lacking BBS10 have congenitally low electrical function in electroretinography. In this study, we performed gene augmentation therapy by injecting a viral construct subretinally to deliver the coding sequence of the mouse Bbs10 gene to treat retinal degeneration in a BBS10 mouse model. Long-term efficacy was assessed by measuring the electrical functions of the retina over time, imaging of the treated regions to visualize cell survival, conducting visually guided swim assays to measure functional vision, and performing retinal histology. We show that subretinal gene therapy slowed photoreceptor cell death and preserved retinal function in treated eyes. Notably, cone photoreceptors regained their electrical function after gene augmentation. Measurement of functional vision showed that subretinal gene therapy provided a significant benefit in delaying vision loss.

Tandem mass tag labeling to assess proteome differences between intermediate and very tender beef steaks.

Tenderness is considered as one of the most important quality attributes dictating consumers' overall satisfaction and future purchasing decisions of fresh beef. However, the ability to predict and manage tenderness has proven very challenging due to the numerous factors that contribute to variation in end-product tenderness. Proteomic profiling allows for global examination of differentially abundant proteins in the meat and can provide new insight into biological mechanisms related to meat tenderness. Hence, the objective of this study was to examine proteomic profiles of beef longissimus lumborum (LL) steaks varying in tenderness, with the intention to identify potential biomarkers related to tenderness. For this purpose, beef LL muscle samples were collected from 99 carcasses at 0 and 384 h postmortem. Based on Warner-Bratzler shear force values at 384 h, 16 samples with the highest (intermediate tender, IT) and lowest (very tender, VT) values were selected to be used for proteomic analysis in this study (n = 8 per category). Using tandem mass tag-based proteomics, a total of 876 proteins were identified, of which 51 proteins were differentially abundant (P < 0.05) between the tenderness categories and aging periods. The differentially identified proteins encompassed a wide array of biological processes related to muscle contraction, calcium signaling, metabolism, extracellular matrix organization, chaperone, and apoptosis. A greater (P < 0.05) relative abundance of proteins associated with carbohydrate metabolism and apoptosis, and a lower (P < 0.05) relative abundance of proteins involved in muscle contraction was observed in the VT steaks after aging compared with the IT steaks, suggesting that more proteolysis occurred in the VT steaks. This may be explained by the greater (P < 0.05) abundance of chaperonin and calcium-binding proteins in the IT steaks, which could have limited the extent of postmortem proteolysis in these steaks. In addition, a greater (P < 0.05) abundance of connective tissue proteins was also observed in the IT steaks, which likely contributed to the difference in tenderness due to added background toughness. The established proteomic database obtained in this study may provide a reference for future research regarding potential protein biomarkers that are associated with meat tenderness.

P-266 Expression of HSP60 in colorectal cancer and implication in chemotherapeutic responses

Heat shock protein 60 (HSP60), also known as chaperonin and HSPD1, belongs to the heat shock protein family. The HSP60 protein is an intramitochondrial chaperonin protein that takes part in post-translational modification for the client proteins with other family members. Recently, HSP60 has been reported to have a possible role as an oncogenic protein in various cancer types, both in their development and progression and also in the metabolism of certain drugs. The aim of the current study was to explore HSP60 expression at transcript and protein levels in clinical cohort and tissue microarray of colorectal cancers and its role in chemotherapeutic responses in colorectal cancer cell lines.

Identification of AHSA1 as a Potential Therapeutic Target for Breast Cancer: Bioinformatics Analysis and in vitro Studies.

Shenling Baizhu Powder (SBP), a famous Traditional Chinese Medicine (TCM) formulation, has been widely used in the adjuvant treatment of cancers, including breast cancer. This study aims to identify potential new targets for breast cancer treatment based on the network pharmacology of SBP. By analyzing the relationship between herbs and target proteins, potential targets of multiple herbs in SBP were identified by network pharmacology analysis. Besides, by comparing the data of breast cancer tissue with normal tissue, upregulated genes in two breast cancer expression profiles were found. Thereafter, the expression level and prognosis of activator of heat shock protein 90 (HSP90) ATPase activity 1 (AHSA1) were further analyzed in breast cancer by bioinformatics analysis, and the network module of AHSA1 binding protein was constructed. Furthermore, the effect of knocking down AHSA1 on the proliferation, migration, and invasion of breast cancer cells was verified by MTT, clone formation assay, and transwell assay. Vascular endothelial growth factor A (VEGFA), intercellular adhesion molecule 1 (ICAM1), chemokine (C-X-C motif) ligand 8 (CXCL8), AHSA1, and serpin family E member 1 (SERPINE1) were associated with multiple herbs in SBP. AHSA1 was remarkably upregulated in breast cancer tissues and positively correlated with poor overall survival and disease metastasis- free survival. Furthermore, knockdown of AHSA1 significantly inhibited the migration and invasion in MCF-7 and MDA-MB-231 breast cancer cells but had no obvious effect on proliferation. In addition, among the proteins that bind to AHSAl, the network composed of proteasome, chaperonin, and heat shock proteins is closely connected, and these proteins are associated with poor prognosis in a variety of cancers. AHSA1 is positively correlated with breast cancer progression and might act as a novel therapeutic target for breast cancer.

Characterization of Zymomonas mobilis promoters that are functional in Escherichia coli

Zymomonas mobilis ZM4 is a gram-negative, facultative anaerobic, natural ethanologenic bacterium used in industrial production of bio-products. For expression of genes, promoters are required. However, most of the promoters reported from Z.mobilis poorly function in Escherichia coli. This makes the process of expression and screening labor-intensive. In the present study, we compared the strengths of two Z.mobilis promoters, Pchap and Ppap, which drive the expression of chaperonin and phosphatase PAP2 family protein, respectively, with Ptac promoter. In E.coli, the Ptac promoter was found to be the strongest followed by Ppap and Ppdc, while in Z.mobilis, Ppdc was found to be the strongest and Pchap the weakest promoter. Further characterization of the promoters was done by cloning the gfpuv gene which expresses the green fluorescent protein, under their control and measuring the fluorescence of the E.coli transformants. The activity of these promoters was also studied at different pH (pH 5, 7 and 9) and different temperatures (30°C, 37°C and 42°C) in exponential and stationary phases. Both Pchap and Ppap promoters showed higher activity in stationary phase than in exponential phase. Since the promoters were active at all temperatures and pH studied, they can be used for gene expression in E.coli under desired environmental conditions.

CCT8 recovers WTp53-suppressed cell cycle evolution and EMT to promote colorectal cancer progression

LIM and SH3 protein 1 (LASP1) is a metastasis-related protein reported to enhance tumor progression in colorectal cancer (CRC). However, the underlying mechanism is still elusive. The chaperonin protein containing TCP1 (CCT) is a cellular molecular chaperone complex, which is necessary for the correct folding of many proteins. It contains eight subunits, CCT1-8. CCT8 is overexpressed in many cancers, however, studies on CCT8 are limited and its role on CRC development and progression remains elusive. In this study, we confirmed that CCT8 and LASP1 can interact with each other and express positively in CRC cells. CCT8 could recover the ability of LASP1 to promote the invasion of CRC; CCT8 could significantly promote the proliferation, invasion, and metastasis of colorectal cells in vivo and in vitro. Mechanically, CCT8 inhibited the entry of WTp53 into the nucleus, and there was a negative correlation between the expression of CCT8 and the nuclear expression of WTp53 in clinical colorectal tissues. CCT8 promoted the cell cycle evolution and EMT progression of CRC by inhibiting the entry of WTp53 into the nucleus. Clinically, CCT8 was highly expressed in CRC. More importantly, the overall survival of CRC patients with high expression of CCT8 was worse than that of patients with low expression of CCT8. These findings indicate that as LASP1-modulated proteins, CCT8 plays a key role in promoting the progression of colorectal cancer, which provides a potential target for clinical intervention in patients with colorectal cancer.

Recurring exposure to low humidity induces transcriptional and protein level changes in the vocal folds of rabbits

Voice disorders are an important human health condition. Hydration is a commonly recommended preventive measure for voice disorders though it is unclear how vocal fold dehydration is harmful at the cellular level. Airway surface dehydration can result from exposure to low humidity air. Here we have induced airway surface dehydration in New Zealand White rabbits exposed to a recurring 8-h low humidity environment over 15 days. This model mimics an occupational exposure to a low humidity environment. Exposure to moderate humidity was the control condition. Full thickness soft-tissue samples, including the vocal folds and surrounding laryngeal tissue, were collected for molecular analysis. RT-qPCR demonstrated a significant upregulation of MUC4 (mucin 4) and SCL26A9 (chloride channel) and a large fold-change though statistically non-significant upregulation of SCNNA1 (epithelial sodium channel). Proteomic analysis demonstrated differential regulation of proteins clustering into prospective functional groups of muscle structure and function, oxidative stress response, and protein chaperonin stress response. Together, the data demonstrate that recurring exposure to low humidity is sufficient to induce both transcriptional and translational level changes in laryngeal tissue and suggest that low humidity exposure induces cellular stress at the level of the vocal folds.