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The underlying mechanism of thermotolerance, which is a key virulence factor essential for pathogenic fungi such as Cryptococcus neoformans, is largely unexplored. In this study, our findings suggest that Set302, a homolog of Set3 and a subunit of histone deacetylase complex Set3C, contributes to thermotolerance in C. neoformans. Specifically, the deletion of the predicted Set3C core subunit, Set302, resulted in further reduction in the growth of C. neoformans at 39°C, and survival of transient incubation at 50°C. Transcriptomics analysis revealed that the expression levels of numerous heat stress-responsive genes altered at both 30°C and 39°C due to the lack of Set302. Notably, at 39°C, the absence of Set302 led to the downregulation of gene expression related to the ubiquitin-proteasome system (UPS). Based on the GFP-α-synuclein overexpression model to characterize misfolded proteins, we observed a pronounced accumulation of misfolded GFP-α-synuclein at 39°C, consequently inhibiting C. neoformans thermotolerance. Furthermore, the loss of Set302 exacerbated the accumulation of misfolded GFP-α-synuclein during heat stress. Interestingly, the set302∆ strain exhibited a similar phenotype under proteasome stress as it did at 39°C. Moreover, the absence of Set302 led to reduced production of capsule and melanin. set302∆ strain also displayed significantly reduced pathogenicity and colonization ability compared to the wild-type strain in the murine infection model. Collectively, our findings suggest that Set302 modulates thermotolerance by affecting the degradation of misfolded proteins and multiple virulence factors to mediate the pathogenicity of C. neoformans.IMPORTANCECryptococcus neoformans is a pathogenic fungus that poses a potential and significant threat to public health. Thermotolerance plays a crucial role in the wide distribution in natural environments and host colonization of this fungus. Herein, Set302, a critical core subunit for the integrity of histone deacetylase complex Set3C and widely distributed in various fungi and mammals, governs thermotolerance and affects survival at extreme temperatures as well as the formation of capsule and melanin in C. neoformans. Additionally, Set302 participates in regulating the expression of multiple genes associated with the ubiquitin-proteasome system (UPS). By eliminating misfolded proteins under heat stress, Set302 significantly contributes to the thermotolerance of C. neoformans. Moreover, Set302 regulates the pathogenicity and colonization ability of C. neoformans in a murine model. Overall, this study provides new insight into the mechanism of thermotolerance in C. neoformans.

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Mitochondria are the cellular energy hub and central target of metabolic regulation. Mitochondria also facilitate proteostasis through pathways such as the 'mitochondria as guardian in cytosol' (MAGIC) whereby cytosolic misfolded proteins (MPs) are imported into and degraded inside mitochondria. In this study, a genome-wide screen in Saccharomyces cerevisiae uncovered that Snf1, the yeast AMP-activated protein kinase (AMPK), inhibits the import of MPs into mitochondria while promoting mitochondrial biogenesis under glucose starvation. We show that this inhibition requires a downstream transcription factor regulating mitochondrial gene expression and is likely to be conferred through substrate competition and mitochondrial import channel selectivity. We further show that Snf1/AMPK activation protects mitochondrial fitness in yeast and human cells under stress induced by MPs such as those associated with neurodegenerative diseases.

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Mitochondria are key organelles for the optimal function of the cell. Among their many functions, they maintain protein homeostasis through their own proteostatic machinery, which involves proteases and chaperones that regulate protein import and folding inside mitochondria. In the early 2000s, the mitochondrial unfolded protein response (UPRmt) was first described in mammalian cells. This stress response is activated by the accumulation of unfolded/misfolded proteins within the mitochondrial matrix, which results in the transmission of a signal to the nucleus to increase the expression of proteases and chaperones to address the abnormal mitochondrial protein load. After its discovery, this retrograde signaling pathway has also been described in other organisms of different complexities, suggesting that it is a conserved stress response. Although there are some specific differences among organisms, the mechanism of this stress response is mostly similar and involves the transmission of a signal from mitochondria to the nucleus that induces chromatin remodeling to allow the binding of specific transcription factors to the promoters of chaperones and proteases. In the last decade, proteins and signaling pathways that could be involved in the regulation of the UPRmt, including the Wnt signaling pathway, have been described. This minireview aims to summarize what is known about the mechanism of the UPRmt and its regulation, specifically in mammals and C. elegans.

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BACKGROUND: Preeclampsia (PE) is one of the most common hypertensive diseases, affecting 2%-8% of all pregnancies. The high maternal and fetal mortality rates of PE are due to a lack of early identification of affected pregnant women that would have led to closer monitoring and care. Recent data suggest that misfolded proteins might be a promising biomarker for PE prediction, which can be detected in urine samples of pregnant women according to their congophilia (aggregated) characteristic. OBJECTIVE: The main purpose of this trial is to evaluate the value of the urine congophilia-based detection of misfolded proteins for the imminent prediction of PE in women presenting with suspected PE. The secondary objectives are to demonstrate that the presence of urine misfolded proteins correlates with PE-related maternal or neonatal adverse outcomes, and to establish an accurate PE prediction model by combining misfolded proteins with multiple indicators. METHODS: At least 300 pregnant women with clinical suspicion of PE will be enrolled in this prospective cohort study. Participants should meet the following inclusion criteria in addition to a suspicion of PE: ≥18 years old, gestational week between 20+0 and 33+6, and single pregnancy. Consecutive urine samples will be collected, blinded, and tested for misfolded proteins and other PE-related biomarkers at enrollment and at 4 follow-up visits. Clinical assessments of PE status and related complications for all participants will be performed at regular intervals using strict diagnostic criteria. Investigators and participants will remain blinded to the results. Follow-up will be performed until 42 days postpartum. Data from medical records, including maternal and fetal outcomes, will be collected. The performance of urine misfolded proteins alone and combined with other biomarkers or clinical variables for the prediction of PE will be statistically analyzed. RESULTS: Enrollment started in July 2023 and was still open upon manuscript submission. As of March 2024, a total of 251 eligible women have been enrolled in the study and enrollment is expected to continue until August 2024. Results analysis is scheduled to start after all participants reach the follow-up endpoint and complete clinical data are collected. CONCLUSIONS: Upon completion of the study, we expect to derive an accurate PE prediction model, which will allow for proactive management of pregnant women with clinical suspicion of PE and possibly reduce the associated adverse pregnancy outcomes. The additional prognostic value of misfolded proteins is also expected to be confirmed. TRIAL REGISTRATION: Chinese Clinical Trials Registry ChiCTR2300074878; https://www.chictr.org.cn/showproj.html?proj=202096. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): PRR1-10.2196/54026.

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Major histocompatibility complex (MHC) class II molecules play a crucial role in immunity by presenting peptide antigens to helper T cells. Immune cells are generally tolerant to self-antigens. However, when self-tolerance is broken, immune cells attack normal tissues or cells, leading to the development of autoimmune diseases. Genome-wide association studies have shown that MHC class II is the gene most strongly associated with the risk of most autoimmune diseases. When misfolded self-antigens, called neoself antigens, are associated with MHC class II molecules in the endoplasmic reticulum, they are transported by the MHC class II molecules to the cell surface without being processed into peptides. Moreover, neoself antigens that are complexed with MHC class II molecules of autoimmune disease risk alleles exhibit distinct antigenicities compared to normal self-antigens, making them the primary targets of autoantibodies in various autoimmune diseases. Elucidation of the immunological functions of neoself antigens presented on MHC class II molecules is crucial for understanding the mechanism of autoimmune diseases.

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OBJECTIVES: Preeclampsia (PE) poses a serious threat to the health of the pregnant woman and her developing fetus due to the difficulty in diagnosing the condition. The disease can develop and worsen suddenly without noticeable signs and symptoms. Thus, there is an urgent need for a simple Point of Care Test (POCT) that improves accessibility to testing and can be used as an aid in the diagnosis of PE. CercaTest Red is a noninvasive detection kit for impending preeclampsia using urine from pregnant women. This is especially pertinent for women who have limited access to secondary/tertiary healthcare as those in remote settings, low-income countries or simply lack of out of hours laboratory services. METHODS: The kit employs an absorptive column that separates Congo red dye bound to urinary misfolded protein from pregnant women and unbound dye. When a solution of Congo red dye pre-mixed with urine is loaded onto the absorptive matrix in a detection cuvette, the presence (positive) or absence (negative) of misfolded proteins can be determined based on the color of eluate collected in the lower section of the cuvette. 190 and 937 pregnant women who were >18 years old at the gestational age of ≥20 weeks were enrolled for the feasibility and validation cohort, respectively. The consistency between CercaTest Red and clinical diagnosis of PE according to the American College of Obstetricians and Gynecologist (ACOG) Guidelines was analyzed using the kappa statistic. RESULTS: The POCT has a limit of detection (LoD) of human urinary misfolded proteins equivalent to 0.45 µM of denatured human serum albumin, with high reproducibility and stability. An accuracy of 96.84% for diagnosis of preeclampsia with a Kappa statistic of 0.746 (p < 0.001) was validated in a cohort of 937 subjects. CONCLUSION: This test is easy to use, cost-effective and portable with short turnaround time and no laboratory instrument requirement. In the future, the test may have the potential to become quantitative using spectroscopy (Chinese Clinical Trial Registry No. ChiCTR1800017692).

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Background: Huntington's disease is an inherited progressive neurodegenerative disorder caused by an expansion of the polyglutamine tract leading to malformation and aggregation of the mutant huntingtin protein in the cell cytoplasm and nucleus of affected brain regions. The development of neuroprotective agents from plants has received considerable research attention. Objective: Our study aims to investigate the neuroprotective effects of luteolin and the mechanisms that underline its potential mediated protection in the mutant htt neuroblastoma cells. Methods: The mutant htt neuroblastoma cells were transfected with 160Q, and the control wild-type neuroblastoma cells were transfected with 20Q htt for 24 h and later treated with luteolin. Cell viability was determined by MTT and PI staining in both groups, while western blotting was used to evaluate caspase 3 protein expression. Aggregation formation was assessed via immunofluorescence microscopy. Also, western blotting was utilized to measure the protein expression of mutant htt aggregated and soluble protein, Nrf2 and HO-1. The impact of Nrf2 on luteolin-treated neuroblastoma cells was assessed using small interfering RNAs. Results: Our study reports that luteolin can protect cultured cells from mutant huntingtin cytotoxicity, evidenced by increased viability and decreased apoptosis. Also, luteolin reduced the accumulation of soluble and insoluble mutant huntingtin aggregates in mutant htt neuroblastoma cells transfected with 160Q compared to the control wild-type. The mutant htt aggregate reduction mediated by luteolin appeared to be independent of the Nrf2 -HO-1 antioxidant pathway. Conclusion: Luteolin presents a new potential therapeutic and protective agent for the treatment and decreasing the cytotoxicity in neurodegenerative diseases such as Huntington's disease.

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Amyloid is a systemic disease characterized by extracellular deposition of misfolded protein. Gastrointestinal and peritoneal deposition of light chain (AL) amyloid is an under-recognized manifestation of this systemic disease, usually as a late sequela. Here we present a case of recently diagnosed AL peritoneal amyloid that presented in the context of recurrent, acute onset abdominal discomfort and was found to have bowel obstruction complicated by perforation in the setting of AL-mediated gastrointestinal tract infiltration and dysmotility.

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The most common factor that contributes to aging is the loss of proteostasis, resulting in an excess amount of non-functional/damaged proteins. These proteins lead to various age-associated phenotypes such as cellular senescence and dysfunction in the nutrient-sensing pathways. Despite the various factors that can contribute to aging, it is still a process that can be changed. According to recent advances in the field of biology, the ability to alter the pathways that are involved in aging can improve the lifespan of a person. Autophagy is a process that helps in preserving survival during stressful situations, such as starvation. It is a common component of various anti-aging interventions, including those that target the insulin/IGF-1 and rapamycin signaling pathways. It has been shown that altered autophagy is a common feature of old age and its impaired regulation could have significant effects on the aging process. This review aims to look into the role of autophagy in aging and how it can be used to improve one's health.

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Amyotrophic lateral sclerosis (ALS) causes progressive degeneration of the motor neurons. In this study, we delivered the genetic construct including the whole locus of human mutant superoxide dismutase 1 (SOD1) with the promoter region of human SOD1 into porcine zygotes using intracytoplasmic sperm injection-mediated gene transfer, and we thereby generated a pig model of human mutant SOD1-mediated familial ALS. The established ALS pig model exhibited an initial abnormality of motor neurons with accumulated misfolded SOD1. The ALS pig model, with a body size similar to that of human beings, will provide opportunities for cell and gene therapy platforms in preclinical translational research.

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Soluble oligomers arising from the aggregation of the amyloid beta peptide (Aß) have been identified as the main pathogenic agents in Alzheimer's disease (AD). Prefibrillar oligomers of the 42-residue form of Aß (Aß42 O) show membrane-binding capacity and trigger the disruption of Ca2+ homeostasis, a causative event in neuron degeneration. Since bioactive lipids have been recently proposed as potent protective agents against Aß toxicity, we investigated the involvement of sphingosine 1-phosphate (S1P) signalling pathway in Ca2+ homeostasis in living neurons exposed to Aß42 O. We show that both exogenous and endogenous S1P rescued neuronal Ca2+ dyshomeostasis induced by toxic Aß42 O in primary rat cortical neurons and human neuroblastoma SH-SY5Y cells. Further analysis revealed a strong neuroprotective effect of S1P1 and S1P4 receptors, and to a lower extent of S1P3 and S1P5 receptors, which activate the Gi -dependent signalling pathways, thus resulting in the endocytic internalization of the extrasynaptic GluN2B-containing N-methyl-D-aspartate receptors (NMDARs). Notably, the S1P beneficial effect can be sustained over time by sphingosine kinase-1 overexpression, thus counteracting the down-regulation of the S1P signalling induced by Aß42 O. Our findings disclose underlying mechanisms of S1P neuronal protection against harmful Aß42 O, suggesting that S1P and its signalling axis can be considered promising targets for therapeutic approaches for AD.

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Neurodegenerative diseases (NDs) are a cluster of diseases marked by progressive neuronal loss, axonal transport blockage, mitochondrial dysfunction, oxidative stress, neuroinflammation, and aggregation of misfolded proteins. NDs are more prevalent beyond the age of 50, and their symptoms often include motor and cognitive impairment. Even though various proteins are involved in different NDs, the mechanisms of protein misfolding and aggregation are very similar. Recently, several studies have discovered that, like prions, these misfolded proteins have the inherent capability of translocation from one neuron to another, thus having far-reaching implications for understanding the processes involved in the onset and progression of NDs, as well as the development of innovative therapy and diagnostic options. These misfolded proteins can also influence the transcription of other proteins and form aggregates, tangles, plaques, and inclusion bodies, which then accumulate in the CNS, leading to neuronal dysfunction and neurodegeneration. This review demonstrates protein misfolding and aggregation in NDs, and similarities and differences between different protein aggregates have been discussed. Furthermore, we have also reviewed the disposal of protein aggregates, the various molecular machinery involved in the process, their regulation, and how these molecular mechanisms are targeted to build innovative therapeutic and diagnostic procedures. In addition, the landscape of various therapeutic interventions for targeting protein aggregation for the effective prevention or treatment of NDs has also been discussed.

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Protein quality control is essential for cellular homeostasis. In this study, we examined the effect of improperly folded proteins that do not form amyloid fibrils on mitochondria, which play important roles in ATP production and cell death. First, we prepared domain 3 of the dengue envelope protein in wild type and four mutants with widely different biophysical properties in misfolded/aggregated or destabilized states. The effects of the different proteins were detected using fluorescence microscopy and Western blotting, which revealed that three of the five proteins disrupted both inner and outer membrane integrity, while the other two proteins, including the wild type, did not. Next, we examined the common characteristics of the proteins that displayed toxicity against mitochondria by measuring oligomer size, molten globule-like properties, and thermal stability. The common feature of all three toxic proteins was thermal instability. Therefore, our data strongly suggest that thermally unstable proteins generated in the cytosol can cause cellular damage by coming into direct contact with mitochondria. More importantly, we revealed that this damage is not amyloid-specific.

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Neurodegenerative diseases are inseparably linked with aging and increase as life expectancy extends. There are common dysfunctions in various cellular events shared among neurogenerative diseases, such as calcium dyshomeostasis, neuroinflammation, and age-associated decline in the autophagy-lysosome system. However, most of all, the prominent pathological feature of neurodegenerative diseases is the toxic buildup of misfolded protein aggregates and inclusion bodies accompanied by an impairment in proteostasis. Recent studies have suggested a close association between endoplasmic reticulum (ER) stress and neurodegenerative pathology in cellular and animal models as well as in human patients. The contribution of mutant or misfolded protein-triggered ER stress and its associated signaling events, such as unfolded protein response (UPR), to the pathophysiology of various neurodegenerative disorders, including Alzheimer's, Parkinson's, and Huntington's disease, amyotrophic lateral sclerosis, and prion disease, is described here. Impaired UPR action is commonly attributed to exacerbated ER stress, pathogenic protein aggregate accumulation, and deteriorating neurodegenerative pathologies. Thus, activating certain UPR components has been shown to alleviate ER stress and its associated neurodegeneration. However, uncontrolled activation of some UPR factors has also been demonstrated to worsen neurodegenerative phenotypes, suggesting that detailed molecular mechanisms around ER stress and its related neurodegenerations should be understood to develop effective therapeutics against aging-associated neurological syndromes. We also discuss current therapeutic endeavors, such as the development of small molecules that selectively target individual UPR components and address ER stress in general.

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Limb-girdle muscular dystrophy type R3 (LGMD R3) is a rare genetic disorder characterized by a progressive proximal muscle weakness and caused by mutations in the SGCA gene encoding alpha-sarcoglycan (α-SG). Here, we report the results of a mechanistic screening ascertaining the molecular mechanisms involved in the degradation of the most prevalent misfolded R77C-α-SG protein. We performed a combinatorial study to identify drugs potentializing the effect of a low dose of the proteasome inhibitor bortezomib on the R77C-α-SG degradation inhibition. Analysis of the screening associated to artificial intelligence-based predictive ADMET characterization of the hits led to identification of the HDAC inhibitor givinostat as potential therapeutical candidate. Functional characterization revealed that givinostat effect was related to autophagic pathway inhibition, unveiling new theories concerning degradation pathways of misfolded SG proteins. Beyond the identification of a new therapeutic option for LGMD R3 patients, our results shed light on the potential repurposing of givinostat for the treatment of other genetic diseases sharing similar protein degradation defects such as LGMD R5 and cystic fibrosis.

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BACKGROUND: The Z-type variant of human α1-antitrypsin is involved in liver cirrhosis and pulmonary emphysema. Due to its slow folding characteristics, this variant accumulates folding intermediates and forms protein aggregates within hepatocytes. Misfolded proteins may induce oxidative stress and subsequent cell death. OBJECTIVE: The potential application of antioxidant response signaling pathway and antioxidants to cope with Z-type α1-antitrypsin-induced oxidative stress was evaluated. METHODS: Overexpression of Z-type α1-antitrypsin in Saccharomyces cerevisiae provoked oxidative stress and increased susceptibility to oxidative challenges such as hydrogen peroxide treatment. Deletion of antioxidant-response genes, including yap1, skn7, sod2, tsa1, and pst2, exacerbated the slow growth phenotype of Z-type α1-antitrypsin-expressing cells. Antioxidant treatment alleviated oxidative stress and cytotoxicity induced by Z-type α1-antitrypsin. RESULTS: Our results show that cellular antioxidant capacity is crucial to protection against misfolded Z-type α1-antitrypsin. CONCLUSION: The information obtained here may be used to prevent oxidative stress caused by misfolded proteins, which are associated with several degenerative diseases, including amyotrophic lateral sclerosis and Parkinson's disease.

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CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy) is the most common familial form of stroke, which is caused by mutations located in the epidermal growth factor (EGF)-like repeats of the NOTCH3 gene. Mutations cause the NOTCH3 (N3) protein to misfold and aggregate. These aggregates will be a component of granular osmiophilic material, which when accumulated around the arteries and arterioles is believed to cause the degradation of vascular smooth muscle cells (VSMC). VSMC degradation affects blood flow regulation and leads to white matter and neuronal death. Currently, there is no treatment for CADASIL. The dementia-relevant BRICHOS domain is a small multitalented protein with functions that include ATP-independent chaperone-like properties. BRICHOS has been shown to prevent the aggregation of both fibrillar and non-fibrillar structures. Therefore, the objective of this study is to investigate whether BRICHOS exhibits anti-aggregating properties on a recombinant CADASIL-mutated N3 protein consisting of the first five repeats of EGF (EGF1-5), harboring a cysteine instead of an arginine in the position 133, (R133C). We found that the N3 EGF1-5 R133C mutant is more prone to aggregate, while the wildtype is more stable. Recombinant human Bri2 BRICHOS is able to interact and stabilize the R133C-mutated N3 protein in a dose-dependent manner. These results suggest an anti-aggregating impact of BRICHOS on the N3 EGF1-5 R133C protein, which could be a potential treatment for CADASIL.

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The major histocompatibility complex (MHC) is a central molecule in the immune system. The MHC is also a highly polymorphic gene that is the most strongly involved in susceptibility to many autoimmune diseases. Therefore, the elucidation of the mechanism by which specific MHC alleles are involved in autoimmune disease susceptibility is important to our understanding of the pathogenesis of autoimmune diseases. Specific autoantibody production is observed in many autoimmune diseases, but the mechanism underlying their production remains unclear. We have found that MHC class II molecules exhibit a chaperone-like function to transport misfolded proteins in the endoplasmic reticulum to the outside of cells. Furthermore, misfolded proteins transported to the cell surface by MHC class II molecules of autoimmune disease risk alleles are major autoantibody targets. In this article, I propose a novel mechanism underlying autoimmune diseases mediated by misfolded proteins complexed with MHC class II molecules.

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Background: The association between misfolded proteins presented in the urine of pregnant women and pregnancy outcomes associated with early-onset pre-eclampsia (PE) remains unclear. This study aimed to investigate this association to examine the predictive value of urinary congophilia in the prognostication of pregnancy outcomes in this patient group in the Chinese population. Materials and Methods: This study included 1,397 patients, of which 46, 147, and 8 patients had gestational hypertension, PE, and chronic hypertension, respectively, and 1,196 were healthy controls undergoing the CapCord test for urinary congophilia. Patients with PE were divided into early- and late-onset groups. Patients with early-onset PE were further divided into iatrogenic prematurity and full-term delivery groups, the rates of urinary congophilia were compared between the groups; additionally, this patient group was divided into positive and negative urinary congophilia groups, clinical characteristics and pregnancy outcomes were compared between the groups. Univariate and multivariate logistic regression analyses were performed. Results: A total of 113 (76.9%) of 147 patients in the PE group had urinary congophilia; this rate was higher than that observed in the other three groups (χ2 = 780.892, p < 0.001). Gestational age in the early-onset PE group at both onset and delivery was lower than that in the late-onset PE group (p < 0.001). The rates of iatrogenic prematurity and hemolysis, elevated liver enzymes, and low platelet count syndrome were both higher in the early-onset PE group than in the late-onset PE group (p < 0.001, p < 0.05). In addition, the rate of urinary congophilia in the early-onset PE group was higher than that in the late-onset PE group (χ2 = 13.297, p < 0.001). Urinary congophilia was an independent risk factor for iatrogenic prematurity among patients with early-onset PE in both univariate [odds ratio (OR) 17.143, 95% confidence interval (CI): 4.719-62.271; p < 0.001] and multivariate (OR 18.174; 95% CI: 4.460-74.063; p < 0.001) analyses. Patients with early-onset PE and urinary congophilia were more likely than their counterparts without urinary congophilia to deliver at a lower gestational age, present with iatrogenic prematurity, and have a shorter latency period between onset and delivery. Conclusion: Urinary congophilia confirmed with the CapCord test may help predict pregnancy outcomes in patients with early-onset PE.

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Most cancer mortality results from metastatic tumor cells and not the localized tumor. Overcoming anoikis is one of the most important steps for detached tumor cells to migrate and metastasize. However, the molecular mechanisms remain to be fully deciphered. Herein, our study revealed upregulation of vacuolar ATPase (V-ATPase) in cancer cells during ECM detachment plays a key role in anoikis evasion. V-ATPase is an enzyme complex that utilizes energy from ATP hydrolysis to maintain cellular homeostasis and had been reported to enhance cancer progression. In this study, V-ATPase inhibition sensitized human cervical cancer, breast cancer, and murine melanoma cells to anoikis via increased ROS production, accumulation of misfolded protein, and impaired pulmonary metastasis in vivo. Scavenging ROS restored anoikis resistance and clearance of misfolded protein accumulation in the tumor cells. Mechanistically, STAT3 upregulates V-ATPase expression while blockade of STAT3 activity repressed V-ATPase expression in these tumor cells as well as sensitized cells to anoikis, increased ROS production, and misfolded protein accumulation. Altogether, our data demonstrate an unreported role of STAT3 in mediating the upregulation of V-ATPase to promote anoikis resistance, thus provides an alternative option to target cancer metastasis.