HSP90 Inhibition Attenuated Isoflurane-Induced Neurotoxicity in Mice and Human Neuroglioma Cells.

Isoflurane, a widely used inhalation anesthetic in clinical practice, is associated with an increased risk of neuronal injury. Heat shock protein 90 (HSP90) plays a crucial role in maintaining neuronal homeostasis under stress conditions; however, its role during isoflurane exposure remains poorly understood. In this study, we aimed to investigate the protective effects of HSP90 inhibition and explore the regulatory mechanisms underlying these effects during isoflurane exposure. We found that the HSP90 inhibitor 17-N-allylamino-17-demethoxygeldanamycin (17 AAG) has great protective effects in mitigating isoflurane-induced ferroptosis of mouse hippocampus and cultured neuronal cells. We focused on the activity of the crucial protein GPX4 in ferroptosis and found that 17 AAG exerted protective effects, preserving the physiological GPX4 activity under isoflurane exposure; further, 17 AAG restored the protein level of GPX4. Further, we observed that the chaperone-mediated autophagy (CMA) pathway was activated; 17 AAG also mediated GPX4 degradation under isoflurane exposure. Additionally, it interfered with the formation of complexes between HSP90 and Lamp-2a, inhibiting CMA activity, followed by the blockade of GPX4 degradation, further affecting the isoflurane-induced ferroptosis. Based on these findings, we proposed HSP90 inhibition as a protective mechanism against isoflurane-induced ferroptosis in neurons.

Chaperone-mediated autophagy degrade Dicer to promote breast cancer metastasis.

Metastasis in breast cancer usually lead to the majority of deaths on clinical patients. Accordingly, diagnosis of metastasis at the early stage in breast cancer is important to improve the prognosis. We observed that Dicer protein levels are significant decrease in highly invasive breast cancer cells and usually correlated with poor clinical outcomes. Following, we aim to clarify the molecular regulatory mechanism of this phenomenon in breast cancer to provide a new therapeutic target. In this study, we obtained that Dicer expression correlated with metastasis and invasion without affect cell stability in breast cancer cells. Importantly, we identified the regulatory mechanism of Dicer protein degradation, the chaperone-mediated autophagy (CMA)-mediated degradation that is major mechanism to decrease Dicer protein expression and lead to cancer metastasis. We discovered that heat shock cognate 71-kDa protein (Hsc70) which as a CMA-related factor interacts with the CMA-targeting motif I333A/K334A on Dicer to promote degradation through CMA. Taken together, our findings hint that Dicer highly correlated with cancer metastasis, we reveal the tumor-promoting effect of CMA-mediated Dicer degradation in breast cancer.

CMA mediates resistance in breast cancer models.

BACKGROUND: Breast cancer (BC) is the most common malignancy in women and the second leading cause of cancer-related death; chemoresistance is still a clinical challenge mainly because of the different molecular features of this kind of tumour. Doxorubicin (Doxo) is widely used despite its adverse effects and the common onset of resistance. Chaperone-Mediated Autophagy (CMA) has been identified as an important mechanism through which chemotherapeutics can exert their cytotoxic effects and, in this context, LAMP-2A, the key player of CMA, can be a useful biomarker. METHODS: A cohort of patients and breast cancer cells have been screened for Doxo effect and CMA activation by analysing the LAMP-2A level. Molecular silencing has been used to clarify CMA role in BC responsiveness to treatments. Low Doxo doses were combined with other drugs (TMZ or PX-478, a HIF-1α inhibitor) to evaluate their cytotoxic ability and their role in modulating CMA. RESULTS: In this paper, we showed that CMA is an important mechanism mediating the responsiveness of breast cancer cell to different treatments (Doxo and TMZ, as suggested by triple negative cells that are TMZ-resistant and fails to activate CMA). The LAMP-2A expression level was specific for different cell lines and patient-derived tumour subtypes, and was also useful in discriminating patients for their survival rates. Moreover, molecular silencing or pharmacological blockage of HIF-1α activity reverted BC resistance to TMZ. The combination of low-dose Doxo with TMZ or PX-478 showed that the drug associations have synergistic behaviours. CONCLUSION: Here, we demonstrated that CMA activity exerts a fundamental role in the responsiveness to different treatments, and LAMP-2A can be proposed as a reliable prognostic biomarker in breast cancer. In this context, HIF-1α, a potential target of CMA, can also be assessed as a valuable therapeutic target in BC in view of identifying new, more efficient and less toxic therapeutic drug combinations. Moreover, the possibility to combine Doxo with other drugs acting on different but coherent molecular targets could help overcome resistance and open the way to a decrease in the dose of the single drugs.

Integrated stress response in hepatitis C promotes Nrf2-related chaperone-mediated autophagy: A novel mechanism for host-microbe survival and HCC development in liver cirrhosis.

The molecular mechanism(s) how liver damage during the chronic hepatitis C virus (HCV) infection evolve into cirrhosis and hepatocellular carcinoma (HCC) is unclear. HCV infects hepatocyte, the major cell types in the liver. During infection, large amounts of viral proteins and RNA replication intermediates accumulate in the endoplasmic reticulum (ER) of the infected hepatocyte, which creates a substantial amount of stress response. Infected hepatocyte activates a different type of stress adaptive mechanisms such as unfolded protein response (UPR), antioxidant response (AR), and the integrated stress response (ISR) to promote virus-host cell survival. The hepatic stress is also amplified by another layer of innate and inflammatory response associated with cellular sensing of virus infection through the production of interferon (IFN) and inflammatory cytokines. The interplay between various types of cellular stress signal leads to different forms of cell death such as apoptosis, necrosis, and autophagy depending on the intensity of the stress and nature of the adaptive cellular response. How do the adaptive cellular responses decode such death programs that promote host-microbe survival leading to the establishment of chronic liver disease? In this review, we discuss how the adaptive cellular response through the Nrf2 pathway that promotes virus and cell survival. Furthermore, we provide a glimpse of novel stress-induced Nrf2 mediated compensatory autophagy mechanisms in virus-cell survival that degrade tumor suppressor gene and activation of oncogenic signaling during HCV infection. Based on these facts, we hypothesize that the balance between hepatic stress, inflammation and different types of cell death determines liver disease progression outcomes. We propose that a more nuanced understanding of virus-host interactions under excessive cellular stress may provide an answer to the fundamental questions why some individuals with chronic HCV infection remain at risk of developing cirrhosis, cancer and some do not.

Methyltransferase-like 21c methylates and stabilizes the heat shock protein Hspa8 in type I myofibers in mice.

Protein methyltransferases mediate posttranslational modifications of both histone and nonhistone proteins. Whereas histone methylation is well-known to regulate gene expression, the biological significance of nonhistone methylation is poorly understood. Methyltransferase-like 21c (Mettl21c) is a newly classified nonhistone lysine methyltransferase whose in vivo function has remained elusive. Using a Mettl21cLacZ knockin mouse model, we show here that Mettl21c expression is absent during myogenesis and restricted to mature type I (slow) myofibers in the muscle. Using co-immunoprecipitation, MS, and methylation assays, we demonstrate that Mettl21c trimethylates heat shock protein 8 (Hspa8) at Lys-561 to enhance its stability. As such, Mettl21c knockout reduced Hspa8 trimethylation and protein levels in slow muscles, and Mettl21c overexpression in myoblasts increased Hspa8 trimethylation and protein levels. We further show that Mettl21c-mediated stabilization of Hspa8 enhances its function in chaperone-mediated autophagy, leading to degradation of client proteins such as the transcription factors myocyte enhancer factor 2A (Mef2A) and Mef2D. In contrast, Mettl21c knockout increased Mef2 protein levels in slow muscles. These results identify Hspa8 as a Mettl21c substrate and reveal that nonhistone methylation has a physiological function in protein stabilization.

Impairment of chaperone-mediated autophagy affects neuronal homeostasis through altered expression of DJ-1 and CRMP-2 proteins.

Chaperone-mediated autophagy (CMA) is a substrate-specific mode of lysosomal proteolysis, with multiple lines of evidence connecting its dysfunction to both ageing and disease. We have recently shown that CMA impairment through knock-down of the lysosomal receptor LAMP2A is detrimental to neuronal viability in vivo; however, it is not clear which subset of proteins regulated by the CMA pathway mediate such changes. In this study, we have manipulated CMA function through alterations of LAMP2A abundance in primary rat cortical neurons, to identify potential changes to the neuronal proteome occurring prior to neurotoxic effects. We have identified a list of proteins with significant, >2-fold change in abundance following our manipulations, of which PARK7/DJ-1 - an anti-oxidant implicated in hereditary forms of Parkinson's Disease (PD), and DPYSL2/CRMP-2 - a microtubule-binding phosphoprotein involved in schizophrenia pathogenesis - were both found to have measurable effects on neuronal homeostasis and phenotype. Taken together, this study describes alterations in the abundance of neuronal proteins involved in neuropsychiatric disorders upon CMA manipulation, and suggests that such alterations may in part be responsible for the neurodegeneration observed upon CMA impairment in vivo.

Secretion of misfolded cytosolic proteins from mammalian cells is independent of chaperone-mediated autophagy.

In eukaryotic cells, elimination of misfolded proteins is essential for maintaining protein homeostasis and cell viability. Misfolding-associated protein secretion (MAPS) is a protein quality-control mechanism that exports misfolded cytosolic proteins via a compartment characteristic of late endosomes, but how cytosolic proteins enter this compartment is unclear. Because chaperone-mediated autophagy (CMA) is a known mechanism that imports cytosolic proteins bearing a specific CMA motif to lysosomes for degradation and because late endosomes and lysosomes overlap significantly in mammalian cells, we determined here whether CMA is involved in targeting protein cargoes to the lumen of late endosomes in MAPS. Using HEK293T and COS-7 cells and immunoblotting and -staining and coimmunoprecipitation methods, we show that, unlike CMA, the secretion of misfolded proteins in MAPS does not require cargo unfolding, is inhibited by serum starvation, and is not dependent on the CMA motif in cargo. Intriguingly, knockdown of lysosome-associated membrane protein 2 (LAMP2), which consists of three isoforms, including a variant proposed to form a protein channel on lysosomes for CMA, attenuated MAPS. However, this could not be attributed to the proposed channel function of the LAMP2a isoform because overexpression of a cytosolic MAPS stimulator, DnaJ heat shock protein family (Hsp40) member C5 (DNAJC5), fully rescued the secretion defect associated with LAMP2 deficiency. We conclude that, in MAPS, cargoes use a CMA-independent mechanism to enter a nondegradative prelysosomal compartment.

Chaperone-Mediated Autophagy.

Protein homeostasis is essential for maintaining cell survival. Protein synthesis and degradation coordinately regulate protein homeostasis. Chaperone-mediated autophagy (CMA) was the first lysosomal process to be discovered by which intracellular components are selectively degraded. This process involves the recognition of the substrate, the unfolding and translocation of the substrate, and the degradation of the substrate. By degrading specific target proteins in a timely manner, CMA is involved in a variety of cellular activities. In the past few years, we have acquired a better understanding of how CMA is regulated. It has been reported that peroxide accumulation, aging and/or other pathological signals interfere with CMA function, which in turn induces neurodegenerative diseases, cancer, and other diseases. Combining results from the current research, we summarize the basic processes, regulatory mechanisms, and physiological functions of CMA and discuss its critical role in the development of diseases.

Lysosomal alterations in peripheral blood mononuclear cells of Parkinson's disease patients.

BACKGROUND: Reduced expression of lysosomal-associated membrane protein 2a and heatshock-cognate 70 proteins, involved in chaperone-mediated autophagy and of glucocerebrosidase, is reported in PD brains. The aim of this study was to identify systemic alterations in lysosomal-associated membrane protein 2a, heatshock cognate-70, and glucocerebrosidase levels/activity in peripheral blood mononuclear cells from PD patients. METHODS: Protein/mRNA levels were assessed in PD patients from genetically undetermined background, alpha-synuclein (G209A/A53T), or glucocerebrosidase mutation carriers and age-/sex-matched controls. RESULTS: Heatshock cognate 70 protein levels were reduced in all PD groups, whereas its mRNA levels were decreased only in the genetically undetermined group. Glucocerebrosidase protein levels were decreased only in the genetic PD groups, whereas increased mRNA levels and decreased activity were detected only in the glucocerebrosidase mutation group. CONCLUSIONS: Reduced heatshock cognate-70 levels are suggestive of an apparent systemic chaperone-mediated autophagy dysfunction irrespective of genetic background. Glucocerebrosidase activity may serve as a screening tool to identify glucocerebrosidase mutation carriers with PD.

Lysosomal-associated membrane protein 2 isoforms are differentially affected in early Parkinson's disease.

Lysosomes are the primary catabolic compartment for the degradation of intracellular proteins through autophagy. The presence of abnormal intracellular α-synuclein-positive aggregates in Parkinson's disease (PD) indicates that the degradative capacity of lysosomes is impaired in PD. Specific dysfunction of chaperone-mediated autophagy (CMA) in PD is suggested by reductions in the CMA membrane receptor, lysosomal-associated membrane protein (LAMP) 2A, although whether LAMP2A is the only LAMP2 isoform affected by PD is unknown. Messenger RNA (mRNA) and protein expression of all three LAMP2 isoforms was assessed in brain extracts from regions with and without PD-related increases in α-synuclein in autopsy samples from subjects in the early pathological stage of PD (n = 9), compared to age- and postmortem delay-matched controls (n = 10). In the early stages of PD, mRNA expression of all LAMP2 isoforms was not different from controls, with LAMP2B and LAMP2C protein levels also unchanged in PD. The selective loss of LAMP2A protein directly correlated with the increased levels of α-synuclein and decreased levels of the CMA chaperone heat shock cognate protein 70 in the same PD samples, as well as with the accumulation of cytosolic CMA substrate proteins. Our data show that LAMP2 protein isoforms are differentially affected in the early stages of PD, with LAMP2A selectively reduced in association with increased α-synuclein, and suggests that dysregulation of CMA-mediated protein degradation occurs before substantial α-synuclein aggregation in PD.

Role of chaperone-mediated autophagy in degrading Huntington's disease-associated huntingtin protein.

Mutant N-terminal huntingtin (Htt) protein resulting from Huntington's disease (HD) with expanded polyglutamine accumulates and forms aggregates in vulnerable neurons. Both ubiquitin proteasomal and autophagic pathways contribute to the degradation of mutant Htt. Here, we focus on the involvement of chaperone-mediated autophagy (CMA), a selective form of autophagy in the clearance of Htt. Selective catabolism in CMA is conferred by the presence of a KFERQ-like targeting motif in the substrates, by which molecular chaperones recognize the hydrophobic surfaces of the misfolded substrates, and transfer them to the lysosomal membrane protein type-2A, LAMP-2A. The substrates are taken into the lysosomes through LAMP-2A and are rapidly degraded by the lysosomal enzymes. Taken together, we summarize the recent evidence to elucidate that Htt is also a potential substrate of CMA. We propose that the manipulation of CMA could be a therapeutic strategy for HD.

TRPML1 contributes to antimony-induced nephrotoxicity by initiating ferroptosis via chaperone-mediated autophagy.

Evidence suggests that ferroptosis participates in kidney injury. However, the role of ferroptosis in antimony (Sb) induced nephrotoxicity and the mechanism are unknown. Here, we demonstrated that Sb induced injury in renal tubular epithelial cells (RTECs) and ferroptosis. Inhibition of ferroptosis reduced RTECs injury. Besides, elimination of reactive oxygen species (ROS) alleviated ferroptosis and RTECs injury. Moreover, exposure to Sb not only increased the co-localization of glutathione peroxidase 4 (GPX4) and LAMP1, but also decreased the levels of MEF2D and LRRK2, while increased the levels of HSC70, HSP90, and LAMP2a. These findings suggest that Sb activates chaperone-mediated autophagy (CMA), enhances lysosomal transport and subsequent degradation of GPX4, ultimately leads to ferroptosis. Additionally, up-regulation of lysosomal cationic channel, TRPML1, mitigated RTECs injury and ferroptosis. Mechanistically, up-regulation of TRPML1 mitigated the changes in CMA-associated proteins induced by Sb, diminished the binding of HSC70, HSP90, and TRPML1 with LAMP2a. Furthermore, NAC restored the decreased TRPML1 level caused by Sb. In summary, deficiency of TRPML1, secondary to increased ROS induced by Sb, facilitates the CMA-dependent degradation of GPX4, thereby leading to ferroptosis and RTECs injury. These findings provide insights into the mechanism underlying Sb-induced nephrotoxicity and propose TRPML1 as a promising therapeutic target.

Recent Advances on Mutant p53: Unveiling Novel Oncogenic Roles, Degradation Pathways, and Therapeutic Interventions.

The p53 protein is the master regulator of cellular integrity, primarily due to its tumor-suppressing functions. Approximately half of all human cancers carry mutations in the TP53 gene, which not only abrogate the tumor-suppressive functions but also confer p53 mutant proteins with oncogenic potential. The latter is achieved through so-called gain-of-function (GOF) mutations that promote cancer progression, metastasis, and therapy resistance by deregulating transcriptional networks, signaling pathways, metabolism, immune surveillance, and cellular compositions of the microenvironment. Despite recent progress in understanding the complexity of mutp53 in neoplastic development, the exact mechanisms of how mutp53 contributes to cancer development and how they escape proteasomal and lysosomal degradation remain only partially understood. In this review, we address recent findings in the field of oncogenic functions of mutp53 specifically regarding, but not limited to, its implications in metabolic pathways, the secretome of cancer cells, the cancer microenvironment, and the regulating scenarios of the aberrant proteasomal degradation. By analyzing proteasomal and lysosomal protein degradation, as well as its connection with autophagy, we propose new therapeutical approaches that aim to destabilize mutp53 proteins and deactivate its oncogenic functions, thereby providing a fundamental basis for further investigation and rational treatment approaches for TP53-mutated cancers.

The novel lnc-HZ12 suppresses autophagy degradation of BBC3 by preventing its interactions with HSPA8 to induce trophoblast cell apoptosis.

Abnormal expression of long non-coding RNAs (lncRNAs) is associated with the dysfunctions of human trophoblast cells and the occurrence of miscarriage (abnormal early embryo loss). BBC3/PUMA (BCL2 binding component 3) plays significant roles in regulation of cell apoptosis. However, whether specific lncRNAs might regulate BBC3 in trophoblast cells and further induce apoptosis and miscarriage remains completely unclear. Through screening, we identified a novel lnc-HZ12, which was significantly highly expressed in villous tissues of recurrent miscarriage (RM) patients relative to their healthy control (HC) group. Lnc-HZ12 suppressed chaperone-mediated autophagy (CMA) degradation of BBC3, promoted trophoblast cell apoptosis, and was associated with miscarriage. In mechanism, lnc-HZ12 downregulated the expression levels of chaperone molecules HSPA8 and LAMP2A in trophoblast cells. Meanwhile, lnc-HZ12 (mainly lnc-HZ12-SO2 region in F2 fragment) and HSPA8 competitively bound with the 169RVLYNL174 patch on BBC3, which prevented BBC3 from interactions with HSPA8 and impaired the formation of BBC3-HSPA8-LAMP2A complex for CMA degradation of BBC3. Thus, lnc-HZ12 upregulated the BBC3-CASP9-CASP3 pathway and induced trophoblast cell apoptosis. In villous tissues, lnc-HZ12 was highly expressed, CMA degradation of BBC3 was suppressed, and the apoptosis levels were higher in RM vs HC villous tissues, all of which were associated with miscarriage. Interestingly, knockdown of murine Bbc3 could efficiently suppress placental apoptosis and alleviate miscarriage in a mouse miscarriage model. Taken together, our results indicated that lnc-HZ12 and BBC3 played important roles in trophoblast cell apoptosis and miscarriage and might act as attractive targets for miscarriage treatment.Abbreviation: 7-AAD: 7-aminoactinomycin D; BaP: benzopyrene; BBC3/PUMA: BCL2 binding component 3; ChIP: chromatin immunoprecipitation; CHX: cycloheximide; CMA: chaperone-mediated autophagy; CQ: chloroquine; DMSO: dimethyl sulfoxide; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HC: healthy control; HSPA8: heat shock protein family A (Hsp70) member 8; IP: immunoprecipitation; LAMP2A: lysosomal associated membrane protein 2; LncRNA: long non-coding RNA; mRNA: messenger RNA; MT: mutant-type; NC: negative control; NSO: nonspecific oligonucleotide; PARP1: poly(ADP-ribose) polymerase 1; RIP: RNA immunoprecipitation; RM: recurrent miscarriage; TBP: TATA-box binding protein; WT: wild-type.

Chaperone-Mediated Autophagy: A Potential Target for Metabolic Diseases.

Autophagy is the process by which cells selectively remove damaged organelles or proteins. Chaperone-mediated autophagy (CMA) is a type of autophagy that degrades proteins containing the KFERQ pentapeptide in cells. CMA can degrade damaged or excess proteins and therefore plays an important role in maintaining protein balance in cells. CMA can also play a regulatory role by degrading key proteins in life activities, such as lipid and glucose metabolism. This review introduces the CMA process and describes the current commonly used CMA detection methods. In addition, we describe the role of CMA in glucose and lipid metabolism. Finally, we summarize the current role of CMA in metabolic diseases such as diabetic nephropathy (DN), alcoholic liver disease (ALD) and nonalcoholic fatty liver disease (NAFLD) and discuss the role of CMA as a potential therapeutic target for metabolic diseases.

Palmitic and Stearic Acids Inhibit Chaperone-Mediated Autophagy (CMA) in POMC-like Neurons In Vitro.

The intake of food with high levels of saturated fatty acids (SatFAs) is associated with the development of obesity and insulin resistance. SatFAs, such as palmitic (PA) and stearic (SA) acids, have been shown to accumulate in the hypothalamus, causing several pathological consequences. Autophagy is a lysosomal-degrading pathway that can be divided into macroautophagy, microautophagy, and chaperone-mediated autophagy (CMA). Previous studies showed that PA impairs macroautophagy function and insulin response in hypothalamic proopiomelanocortin (POMC) neurons. Here, we show in vitro that the exposure of POMC neurons to PA or SA also inhibits CMA, possibly by decreasing the total and lysosomal LAMP2A protein levels. Proteomics of lysosomes from PA- and SA-treated cells showed that the inhibition of CMA could impact vesicle formation and trafficking, mitochondrial components, and insulin response, among others. Finally, we show that CMA activity is important for regulating the insulin response in POMC hypothalamic neurons. These in vitro results demonstrate that CMA is inhibited by PA and SA in POMC-like neurons, giving an overview of the CMA-dependent cellular pathways that could be affected by such inhibition and opening a door for in vivo studies of CMA in the context of the hypothalamus and obesity.

Role of Chaperone-Mediated Autophagy in Ageing Biology and Rejuvenation of Stem Cells.

What lies at the basis of the mechanisms that regulate the maintenance and self-renewal of pluripotent stem cells is still an open question. The control of stemness derives from a fine regulation between transcriptional and metabolic factors. In the last years, an emerging topic has concerned the involvement of Chaperone-Mediated Autophagy (CMA) as a key mechanism in stem cell pluripotency control acting as a bridge between epigenetic, transcriptional and differentiation regulation. This review aims to clarify this new and not yet well-explored horizon discussing the recent studies regarding the CMA impact on embryonic, mesenchymal, and haematopoietic stem cells. The review will discuss how CMA influences embryonic stem cell activity promoting self-renewal or differentiation, its involvement in maintaining haematopoietic stem cell function by increasing their functionality during the normal ageing process and its effects on mesenchymal stem cells, in which modulation of CMA regulates immunosuppressive and differentiation properties. Finally, the importance of these new discoveries and their relevance for regenerative medicine applications, from transplantation to cell rejuvenation, will be addressed.

Placental Antioxidant Defenses and Autophagy-Related Genes in Maternal Obesity and Gestational Diabetes Mellitus.

Maternal obesity and gestational diabetes mellitus (GDM) are increasing worldwide, representing risk factors for both mother and child short/long-term outcomes. Oxidative stress, lipotoxicity and altered autophagy have already been reported in obesity, but few studies have focused on obese pregnant women with GDM. Antioxidant and macro/chaperone-mediated autophagy (CMA)-related gene expressions were evaluated herein in obese and GDM placentas. A total of 47 women with singleton pregnancies delivered by elective cesarean section were enrolled: 16 normal weight (NW), 18 obese with no comorbidities (OB GDM(-)), 13 obese with GDM (OB GDM(+)). Placental gene expression was assessed by real-time PCR. Antioxidant gene expression (CAT, GPX1, GSS) decreased, the pro-autophagic ULK1 gene increased and the chaperone-mediated autophagy regulator PHLPP1 decreased in OB GDM(-) vs. NW. On the other hand, PHLPP1 expression increased in OB GDM(+) vs. OB GDM(-). When analyzing results in relation to fetal sex, we found sexual dimorphism for both antioxidant and CMA-related gene expressions. These preliminary results can pave the way for further analyses aimed at elucidating the placental autophagy role in metabolic pregnancy disorders and its potential targetability for the treatment of diabetes outcomes.

Chaperone-Mediated Autophagy Markers LAMP2A and HSPA8 in Advanced Non-Small Cell Lung Cancer after Neoadjuvant Therapy.

In recent years autophagy has attracted the attention of researchers from many medical fields, including cancer research, and certain anti-macroautophagy drugs in combination with cytotoxic or targeted therapies have entered clinical trials. In the present study, we focused on a less explored subtype of autophagy, i.e., chaperone-mediated autophagy (CMA), with the key proteins LAMP2A and HSPA8 (HSC70), and their immunohistochemical evaluation with previously extensively validated antibodies. We were interested in whether the marker expression is influenced by the antecedent therapy, and its correlation with survival on a cohort of patients with non-small cell lung cancer (NSCLC) after neoadjuvant therapy and matched primary resected tumors. In concordance with our previous study, we did not find any intratumoral heterogeneity, nor correlation between the two parameters, nor correlation between the markers and any included pathological parameters. Surprisingly, the expression of both markers was also independent to tumor response or administered neoadjuvant treatment. In the survival analysis, the results were only significant for LAMP2A, where higher levels were associated with longer 5-year overall survival and disease-free survival for the mixed group of adenocarcinomas and squamous cell carcinomas (p < 0.0001 and p = 0.0019 respectively) as well as the squamous cell carcinoma subgroup (p = 0.0001 and p = 0.0001 respectively). LAMP2A was also an independent prognostic marker in univariate and multivariate analysis.

Zika virus NS2A protein induces the degradation of KPNA2 (karyopherin subunit alpha 2) via chaperone-mediated autophagy.

KPNA2/importin-alpha1 (karyopherin subunit alpha 2) is the primary nucleocytoplasmic transporter for some transcription factors to activate cellular proliferation and differentiation. Aberrant increase of KPNA2 level is identified as a prognostic marker in a variety of cancers. Yet, the turnover mechanism of KPNA2 remains unknown. Here, we demonstrate that KPNA2 is degraded via the chaperone-mediated autophagy (CMA) and that Zika virus (ZIKV) enhances the KPNA2 degradation. KPNA2 contains a CMA motif, which possesses an indispensable residue Gln109 for the CMA-mediated degradation. RNAi-mediated knockdown of LAMP2A, a vital component of the CMA pathway, led to a higher level of KPNA2. Moreover, ZIKV reduced KPNA2 via the viral NS2A protein, which contains an essential residue Thr100 for inducing the CMA-mediated KPNA2 degradation. Notably, mutant ZIKV with T100A alteration in NS2A replicates much weaker than the wild-type virus. Also, knockdown of KPNA2 led to a higher ZIKV viral yield, which indicates that KPNA2 mediates certain antiviral effects. These data provide insights into the KPNA2 turnover and the ZIKV-cell interactions.