Identification of Host Proteins Interacting with IBV S1 Based on Tracheal Organ Culture.

Infectious bronchitis virus (IBV) belongs to the gamma-coronavirus genus of Coronaviridae and causes serious infectious diseases in the poultry industry. However, only a few IBV strains can infect avian passage cell lines, seriously hindering the progress of basic research on IBV pathogenesis. Whereas IBV field strains can replicate in tracheal ring organ culture (TOC) without any previous adaptation in chicken embryos or primary cells. In this study, to investigate the potential use of TOC as an in vitro infection model for the study of IBV-host interaction, we first established a chicken embryo TOC culture system and carried out an investigation on the IBV replication kinetics in the system. We found that the selected strains of the IBV GI-1, GI-7, GI-13, GI-19, and GI-22 genotypes could successfully replicate in TOC and bring about damage to the infected trachea. Next, we identified host proteins of the chicken embryo trachea that interact with the IBV S1 protein by immunoprecipitation and protein mass spectrometry. A total of 127 candidate proteins were initially identified with major involvement in cell adhesion pathways and apoptosis- and autophagy-related pathways. The heat shock protein 70 (HSP70) was selected for further investigation in the interaction with IBV viral proteins. Our results showed that HSP70 interacted with IBV S1 in both TOC and CEK cells, whereas HSP70 overexpression inhibited viral replication. This study indicates that TOC is a good system for the elucidation of IBV-host interactions and HSP70 is a potential host antiviral factor.

Cold Exposure-Induced Up-Regulation of Hsp70 Positively Regulates PEDV mRNA Synthesis and Protein Expression In Vitro.

Porcine epidemic diarrhea (PED) is a highly contagious, intestinal infectious disease caused by porcine epidemic diarrhea virus (PEDV). PEDV as an emerging and re-emerging epizootic virus of swine causes substantial economic losses to the pig industry in China and other countries. In China, the occurrence of PED shows significant seasonal variations, usually outbreak during the winter season. The epidemic characteristics of PED may be highly correlated with the changes of ambient temperature. However, molecular mechanism on the seasonal occurrence of PED still remains unclear. It has been widely observed that low ambient temperature up-regulates the expression of host heat shock protein 70 (Hsp70). Here, we showed that nucleotide and protein levels of Hsp70 were up-regulated in the intestinal of cold exposed pig and cold exposed Vero E6 cells. We found that overexpression of Hsp70 could increase PEDV mRNA synthesis and protein expression in Vero E6 and IPEC-J2 cells, while the siRNAs mediated knockdown of Hsp70 and VER155008 mediated inhibition of Hsp70 resulted in inhibition of viral mRNA synthesis and protein expression in Vero E6 cells. These data suggested that Hsp70 positively regulated PEDV mRNA synthesis and protein expression, which being helpful for understanding the seasonality of PED epidemics and development of novel antiviral therapies in the future.

The Role of Hsp70 in Adaptation to Adverse Conditions and Its Possible Medical Application.

In the present era of global warming and dramatically increased environmental pollution posing a threat to animal life, the understanding and manipulation of organisms' resources of stress tolerance is apparently a question of survival. Heat stress and other forms of stressful factors induce a highly organized response of organisms at the cellular level where heat shock proteins (Hsps) and in particular Hsp70 family of chaperones are among the major players in the protection from the environmental challenge. The present review article summarizes the peculiarities of the Hsp70 family of proteins protective functions being a result of many millions of years of adaptive evolution. It discusses the molecular structure and specific details of hsp70 gene regulation in various organisms, living in diverse climatic zones, with a special emphasis on the protective role of Hsp70 in adverse conditions of the environment. The review discusses the molecular mechanisms underlying Hsp70-specific properties that emerged in the course of adaptation to harsh environmental conditions. This review also includes the data on the anti-inflammatory role of Hsp70 and the involvement of endogenous and recombinant Hsp70 (recHsp70) in proteostatic machinery in various pathologies including neurodegenerative ones such as Alzheimer's and Parkinson's diseases in rodent model organisms and humans in vivo and in vitro. Specifically, the role of Hsp70 as an indicator of disease type and severity and the use of recHsp70 in several pathologies are discussed. The review discusses different roles exhibited by Hsp70 in various diseases including the dual and sometimes antagonistic role of this chaperone in various forms of cancer and viral infection including the SARS-Cov-2 case. Since Hsp70 apparently plays an important role in many diseases and pathologies and has significant therapeutic potential there is a dire need to develop cheap recombinant Hsp70 production and further investigate the interaction of externally supplied and endogenous Hsp70 in chaperonotherapy.

Bioaccumulation Pattern of the SARS-CoV-2 Spike Proteins in Pacific Oyster Tissues.

There is mounting evidence of the contamination of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the sewage, surface water, and even marine environment. Various studies have confirmed that bivalve mollusks can bioaccumulate SARS-CoV-2 RNA to detectable levels. However, these results do not provide sufficient evidence for the presence of infectious viral particles. To verify whether oysters can bind the viral capsid and bioaccumulate the viral particles, Pacific oysters were artificially contaminated with the recombinant SARS-CoV-2 spike protein S1 subunit (rS1). The bioaccumulation pattern of the rS1 in different tissues was investigated by immunohistological assays. The results revealed that the rS1 was bioaccumulated predominately in the digestive diverticula. The rS1 was also present in the epithelium of the nondigestive tract tissues, including the gills, mantle, and heart. In addition, three potential binding ligands, including angiotensin-converting enzyme 2 (ACE 2)-like substances, A-type histo-blood group antigen (HBGA)-like substances, and oyster heat shock protein 70 (oHSP 70), were confirmed to bind rS1 and were distributed in tissues with various patterns. The colocalization analysis of rS1 and those potential ligands indicated that the distributions of rS1 are highly consistent with those of ACE 2-like substances and oHSP 70. Both ligands are distributed predominantly in the secretory absorptive cells of the digestive diverticula and may serve as the primary ligands to bind rS1. Therefore, oysters are capable of bioaccumulating the SARS-CoV-2 capsid readily by filter-feeding behavior assisted by specific binding ligands, especially in digestive diverticula. IMPORTANCE This is the first article to investigate the SARS-CoV-2 spike protein bioaccumulation pattern and mechanism in Pacific oysters by the histochemical method. Oysters can bioaccumulate SARS-CoV-2 capsid readily by filter-feeding behavior assisted by specific binding ligands. The new possible foodborne transmission route may change the epidemic prevention strategies and reveal some outbreaks that current conventional epidemic transmission routes cannot explain. This original and interdisciplinary paper advances a mechanistic understanding of the bioaccumulation of SARS-CoV-2 in oysters inhabiting contaminated surface water.

Comparison of allosteric signaling in DnaK and BiP using mutual information between simulated residue conformations.

The heat shock protein 70 kDa (Hsp70) chaperone system serves as a critical component of protein quality control across a wide range of prokaryotic and eukaryotic organisms. Divergent evolution and specialization to particular organelles have produced numerous Hsp70 variants which share similarities in structure and general function, but differ substantially in regulatory aspects, including conformational dynamics and activity modulation by cochaperones. The human Hsp70 variant BiP (also known as GRP78 or HSPA5) is of therapeutic interest in the context of cancer, neurodegenerative diseases, and viral infection, including for treatment of the pandemic virus SARS-CoV-2. Due to the complex conformational rearrangements and high sequential variance within the Hsp70 protein family, it is in many cases poorly understood which amino acid mutations are responsible for biochemical differences between protein variants. In this study, we predicted residues associated with conformational regulation of human BiP and Escherichia coli DnaK. Based on protein structure networks obtained from molecular dynamics simulations, we analyzed the shared information between interaction timelines to highlight residue positions with strong conformational coupling to their environment. Our predictions, which focus on the binding processes of the chaperone's substrate and cochaperones, indicate residues filling potential signaling roles specific to either DnaK or BiP. By combining predictions of individual residues into conformationally coupled chains connecting ligand binding sites, we predict a BiP specific secondary signaling pathway associated with substrate binding. Our study sheds light on mechanistic differences in signaling and regulation between Hsp70 variants, which provide insights relevant to therapeutic applications of these proteins.

Targeting heat shock proteins 90 and 70: A promising remedy for both autoimmune bullous diseases and COVID-19.

Heat shock proteins (Hsps), including Hsp90 and Hsp70, are intra- and extracellular molecules implicated in cellular homeostasis and immune processes and are induced by cell stress such as inflammation and infection. Autoimmune bullous disorders (AIBDs) and COVID-19 represent potentially life-threatening inflammatory and infectious diseases, respectively. A significant portion of AIBDs remain refractory to currently available immunosuppressive therapies, which may represent a risk factor for COVID-19, and suffer from treatment side-effects. Despite advances in vaccination, there is still a need to develop new therapeutic approaches targeting SARS-CoV-2, especially considering vaccine hesitancy, logistical distribution challenges, and breakthrough infections. In this mini review, we briefly summarize the role of targeting Hsp90/70 as a promising double-edged sword in the therapy of AIBDs and COVID-19.

Therapeutic potential of green tea catechin, (-)-epigallocatechin-3-O-gallate (EGCG) in SARS-CoV-2 infection: Major interactions with host/virus proteases.

Background: The current COVID-19 pandemic from the human pathogenic virus SARS-CoV-2 has resulted in a major health hazard globally. The morbidity and transmission modality of this disease are severe and uncontrollable. As no effective clinical drugs are available for treatment of COVID-19 infection till to date and only vaccination is used as prophylaxis and its efficacy is restricted due to emergent of new variants of SARS-CoV-2, there is an urgent need for effective drugs for its treatment. Purpose: The aim of this review was to provide a detailed analysis of anti-SARS-CoV-2 efficacy of (-)-epigallocatechin-3-O-gallate (EGCG), a major catechin constituent of green tea (Camellia sinensis (L.) Kuntze) beverage to highlight the scope of EGCG in clinical medicine as both prophylaxis and treatment of present COVID-19 infection. In addition, the factors related to poor oral bioavailabilty of EGCG was also analysed for a suggestion for future research in this direction. Study design: We collected the published articles related to anti-SARS-CoV-2 activity of EGCG against the original strain (Wuhan type) and its newly emerged variants of SARS-CoV-2 virus. Methods: A systematic search on the published literature was conducted in various databases including Google Scholar, PubMed, Science Direct and Scopus to collect the relevant literature. Results: The findings of this search demonstrate that EGCG shows potent antiviral activity against SARS-CoV-2 virus by preventing viral entry and replication in host cells in vitro models. The studies on the molecular mechanisms of EGCG in inhibition of SARS-CoV-2 infection in host cells reveal that EGCG blocks the entry of the virus particles by interaction with the receptor binding domain (RBD) of viral spike (S) protein to host cell surface receptor protease angiotensin-converting enzyme 2 (ACE2) as well as suppression of the expressions of host proteases, ACE2, TMPRSS2 and GRP78, required for viral entry, by Nrf2 activation in host cells. Moreover, EGCG inhibits the activities of SARS-CoV-2 main protease (Mpro), papain-like protease (PLpro), endoribonuclease Nsp15 in vitro models and of RNA-dependent RNA polymerase (RdRp) in molecular docking model for suppression of viral replication. In addition, EGCG significantly inhibits viral inflammatory cytokine production by stimulating Nrf2- dependent host immune response in virus-infected cells. EGCG significantly reduces the elevated levels of HMGB1, a biomarker of sepsis, lung fibrosis and thrombotic complications in viral infections. EGCG potentially inhibits the infection of original (Wuhan type) strain of SARS-CoV-2 and other newly emerged variants as well as the infections of SARS-CoV-2 virus spike-protein of WT and its mutants-mediated pseudotyped viruses . EGCG shows maximum inhibitory effect against SARS-CoV-2 infection when the host cells are pre-incubated with the drug prior to viral infection. A sorbitol/lecithin-based throat spray containing concentrated green tea extract rich in EGCG content significantly reduces SARS-CoV-2 infectivity in oral mucosa. Several factors including degradation in gastrointestinal environment, low absorption in small intestine and extensive metabolism of EGCG are responsible for its poor bioavailability in humans. Pharmacokinetic and metabolism studies of EGCG in humans reveal poor bioavailability of EGCG in human plasma and EGCG-4"-sulfate is its major metabolite. The concentration of EGCG-4"-sulfate in human plasma is almost equivalent to that of free EGCG (Cmax 177.9 vs 233.5 nmol/L). These findings suggest that inhibition of sulfation of EGCG is a crucial factor for improvement of its bioavailability. In vitro study on the mechanism of EGCG sulfonation indicates that sulfotransferases, SULT1A1 and SULT1A3 are responsible for sulfonation in human liver and small intestine, respectively. Some attempts including structural modifications, and nanoformulations of EGCG and addition of nutrients with EGCG have been made to improve the bioavailability of EGCG. Conclusions: The findings of this study suggest that EGCG has strong antiviral activity against SARS-CoV-2 infection independent of viral strains (Wuhan type (WT), other variants) by inhibition of viral entry and replication in host cells in vitro models. EGCG may be useful in reduction of this viral load in salivary glands of COVID-19 patients, if it is applied in mouth and throat wash formulations in optimal concentrations. EGCG could be a promising candidate in the development of effective vaccine for prevention of the infections of newly emergent strains of SARS-CoV-2 virus. EGCG might be useful also as a clinical medicine for treatment of COVID-19 patients if its bioavailability in human plasma is enhanced.

Sandwich-like electrochemical aptasensing of heat shock protein 70 kDa (HSP70): Application in diagnosis/prognosis of coronavirus disease 2019 (COVID-19).

In this research, by using aptamer-conjugated gold nanoparticles (aptamer-AuNPs) and a modified glassy carbon electrode (GCE) with reduced graphene oxide (rGO) and Acropora-like gold (ALG) nanostructure, a sandwich-like system provided for sensitive detection of heat shock protein 70 kDa (HSP70), which applied as a functional biomarker in diagnosis/prognosis of COVID-19. Initially, the surface of the GCE was improved with rGO and ALG nanostructures, respectively. Then, an aptamer sequence as the first part of the bioreceptor was covalently bound on the surface of the GCE/rGO/ALG nanostructures. After adding the analyte, the second part of the bioreceptor (aptamer-AuNPs) was immobilized on the electrode surface to improve the diagnostic performance. The designed aptasensor detected HSP70 in a wide linear range, from 5 pg mL-1 to 75 ng mL-1, with a limit of detection (LOD) of ∼2 pg mL-1. The aptasensor was stable for 3 weeks and applicable in detecting 40 real plasma samples of COVID-19 patients. The diagnostic sensitivity and specificity were 90% and 85%, respectively, compared with the reverse transcription-polymerase chain reaction (RT-PCR) method.

Rationale for 1068 nm Photobiomodulation Therapy (PBMT) as a Novel, Non-Invasive Treatment for COVID-19 and Other Coronaviruses: Roles of NO and Hsp70.

Researchers from across the world are seeking to develop effective treatments for the ongoing coronavirus disease 2019 (COVID-19) outbreak, which arose as a major public health issue in 2019, and was declared a pandemic in early 2020. The pro-inflammatory cytokine storm, acute respiratory distress syndrome (ARDS), multiple-organ failure, neurological problems, and thrombosis have all been linked to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) fatalities. The purpose of this review is to explore the rationale for using photobiomodulation therapy (PBMT) of the particular wavelength 1068 nm as a therapy for COVID-19, investigating the cellular and molecular mechanisms involved. Our findings illustrate the efficacy of PBMT 1068 nm for cytoprotection, nitric oxide (NO) release, inflammation changes, improved blood flow, and the regulation of heat shock proteins (Hsp70). We propose, therefore, that PBMT 1068 is a potentially effective and innovative approach for avoiding severe and critical illness in COVID-19 patients, although further clinical evidence is required.

The Beneficial Role of Physical Exercise on Anthracyclines Induced Cardiotoxicity in Breast Cancer Patients

Simple SummaryBreast cancer is one of the most common cancers worldwide and the leading cause of cancer death in women. Screening, early diagnosis, and surgical techniques might increase patients’ survival, leading to a rise in the health-related consequences of anti-neoplastic therapies, such as cardiotoxicity following anthracycline treatments. Alongside conventional therapies, physical activity seems to reduce treatment side effects, improving quality of life either after a breast cancer diagnosis or in the early steps post-surgery. This review offers a general framework for the role of anthracycline in the physio-pathological mechanisms of cardiotoxicity and the effect of exercise on cancer treatment side effects. Moreover, we propose the type and the timing of exercise to better assist patients and reduce the pressure on the health care system in breast cancer patients undergoing anthracycline.The increase in breast cancer (BC) survival has determined a growing survivor population that seems to develop several comorbidities and, specifically, treatment-induced cardiovascular disease (CVD), especially those patients treated with anthracyclines. Indeed, it is known that these compounds act through the induction of supraphysiological production of reactive oxygen species (ROS), which appear to be central mediators of numerous direct and indirect cardiac adverse consequences. Evidence suggests that physical exercise (PE) practised before, during or after BC treatments could represent a viable non-pharmacological strategy as it increases heart tolerance against many cardiotoxic agents, and therefore improves several functional, subclinical, and clinical parameters. At molecular level, the cardioprotective effects are mainly associated with an exercise-induced increase of stress response proteins (HSP60 and HSP70) and antioxidant (SOD activity, GSH), as well as a decrease in lipid peroxidation, and pro-apoptotic proteins such as Bax, Bax-to-Bcl-2 ratio. Moreover, this protection can potentially be explained by a preservation of myosin heavy chain (MHC) isoform distribution. Despite this knowledge, it is not clear which type of exercise should be suggested in BC patient undergoing anthracycline treatment. This highlights the lack of special guidelines on how affected patients should be managed more efficiently. This review offers a general framework for the role of anthracyclines in the physio-pathological mechanisms of cardiotoxicity and the potential protective role of PE. Finally, potential exercise-based strategies are discussed on the basis of scientific findings.

Possible Therapeutic Intervention Strategies for COVID-19 by Manipulating the Cellular Proteostasis Network.

INTRODUCTION: The recent outbreak of coronavirus infection by SARS-CoV-2 that started from the Wuhan Province of China in 2019 has spread to most parts of the world infecting millions of people. Although the case fatality rate of SARS-CoV-2 infection is less than the previous epidemics by other closely related coronaviruses, due to its high infectivity, the total number of SARS-CoV-2 infection-associated disease, called Covid-19, is a matter of global concern. Despite drastic preventive measures, the number of Covid-19 cases are steadily increasing, and the future course of this pandemic is highly unpredictable. The most concerning fact about Covid-19 is the absence of specific and effective preventive or therapeutic agents against the disease. Finding an immediate intervention against Covid-19 is the need of the hour. In this chapter, we have discussed the role of different branches of the cellular proteostasis network, represented by Hsp70-Hsp40 chaperone system, Ubiquitin-Proteasome System (UPS), autophagy, and endoplasmic reticulum-Unfolded Protein Response (ER-UPR) pathway in the pathogenesis of coronavirus infections and in the host antiviral defense mechanisms. RESULTS: Based on scientific literature, we present that pharmacological manipulation of proteostasis network can alter the fate of coronavirus infections and may help to prevent the resulting pathologies like Covid-19.

Targeting of HSP70/HSF1 Axis Abrogates In Vitro Ibrutinib-Resistance in Chronic Lymphocytic Leukemia.

The Btk inhibitor ibrutinib has significantly changed the management of chronic lymphocytic leukemia (CLL) patients. Despite its clinical efficacy, relapses occur, and outcomes after ibrutinib failure are poor. Although BTK and PLCγ2 mutations have been found to be associated with ibrutinib resistance in a fair percentage of CLL patients, no information on resistance mechanisms is available in patients lacking these mutations. The heat shock protein of 70 kDa (HSP70) and its transcription factor heat shock factor 1 (HSF1) play a role in mediating the survival and progression of CLL, as well as taking part in drug resistance in various cancers. We demonstrated that resveratrol and related phenols were able to induce apoptosis in vitro in leukemic cells from CLL untreated patients by acting on the HSP70/HSF1 axis. The same was achieved in cells recovered from 13 CLL patients failing in vivo ibrutinib treatment. HSP70 and HSF1 levels decreased following in vitro treatment, correlating to apoptosis induction. We suggest an involvement of HSP70/HSF1 axis in controlling resistance to ibrutinib in CLL cells, since their inhibition is effective in inducing in vitro apoptosis in cells from ibrutinib refractory patients. The targeting of HSP70/HSF1 axis could represent a novel rational therapeutic strategy for CLL, also for relapsing patients.

Role of Heat Shock Proteins (HSP70 and HSP90) in Viral Infection.

Heat shock proteins (HSPs) are a large group of chaperones found in most eukaryotes and bacteria. They are responsible for the correct protein folding, protection of the cell against stressors, presenting immune and inflammatory cytokines; furthermore, they are important factors in regulating cell differentiation, survival and death. Although the biological function of HSPs is to maintain cell homeostasis, some of them can be used by viruses both to fold their proteins and increase the chances of survival in unfavorable host conditions. Folding viral proteins as well as replicating many different viruses are carried out by, among others, proteins from the HSP70 and HSP90 families. In some cases, the HSP70 family proteins directly interact with viral polymerase to enhance viral replication or they can facilitate the formation of a viral replication complex and/or maintain the stability of complex proteins. It is known that HSP90 is important for the expression of viral genes at both the transcriptional and the translational levels. Both of these HSPs can form a complex with HSP90 and, consequently, facilitate the entry of the virus into the cell. Current studies have shown the biological significance of HSPs in the course of infection SARS-CoV-2. A comprehensive understanding of chaperone use during viral infection will provide new insight into viral replication mechanisms and therapeutic potential. The aim of this study is to describe the molecular basis of HSP70 and HSP90 participation in some viral infections and the potential use of these proteins in antiviral therapy.

First Virtual International Congress on Cellular and Organismal Stress Responses, November 5-6, 2020.

Members of the Cell Stress Society International (CSSI), Patricija van Oosten-Hawle (University of Leeds, UK), Mehdi Mollapour (SUNY Upstate Medical University, USA), Andrew Truman (University of North Carolina at Charlotte, USA) organized a new virtual meeting format which took place on November 5-6, 2020. The goal of this congress was to provide an international platform for scientists to exchange data and ideas among the Cell Stress and Chaperones community during the Covid-19 pandemic. Here we will highlight the summary of the meeting and acknowledge those who were honored by the CSSI.

Novel Broad-Spectrum Antiviral Inhibitors Targeting Host Factors Essential for Replication of Pathogenic RNA Viruses.

Recent RNA virus outbreaks such as Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Ebola virus (EBOV) have caused worldwide health emergencies highlighting the urgent need for new antiviral strategies. Targeting host cell pathways supporting viral replication is an attractive approach for development of antiviral compounds, especially with new, unexplored viruses where knowledge of virus biology is limited. Here, we present a strategy to identify host-targeted small molecule inhibitors using an image-based phenotypic antiviral screening assay followed by extensive target identification efforts revealing altered cellular pathways upon antiviral compound treatment. The newly discovered antiviral compounds showed broad-range antiviral activity against pathogenic RNA viruses such as SARS-CoV-2, EBOV and Crimean-Congo hemorrhagic fever virus (CCHFV). Target identification of the antiviral compounds by thermal protein profiling revealed major effects on proteostasis pathways and disturbance in interactions between cellular HSP70 complex and viral proteins, illustrating the supportive role of HSP70 on many RNA viruses across virus families. Collectively, this strategy identifies new small molecule inhibitors with broad antiviral activity against pathogenic RNA viruses, but also uncovers novel virus biology urgently needed for design of new antiviral therapies.

Moderate Fever Cycles as a Potential Mechanism to Protect the Respiratory System in COVID-19 Patients.

Mortality in COVID-19 patients predominantly results from an acute respiratory distress syndrome (ARDS), in which lungs alveolar cells undergo programmed cell death. Mortality in a sepsis-induced ARDS rat model is reduced by adenovirus over-expression of the HSP70 chaperone. A natural rise of body temperature during mild fever can naturally accumulate high cellular levels of HSP70 that can arrest apoptosis and protect alveolar lung cells from inflammatory damages. However, beyond 1-2 h of fever, no HSP70 is being further produced and a decreased in body temperature required to the restore cell's ability to produce more HSP70 in a subsequent fever cycle. We suggest that antipyretics may be beneficial in COVID-19 patients subsequent to several hours of mild (<38.8°C) advantageous fever, allowing lung cells to accumulate protective HSP70 against damages from the inflammatory response to the virus SARS-CoV-2. With age, the ability to develop fever and accumulate HSP70 decreases. This could be ameliorated, when advisable to do so, by thermotherapies and/or physical training.

Interaction of HSPA5 (Grp78, BIP) with negatively charged phospholipid membranes via oligomerization involving the N-terminal end domain.

Heat shock proteins (HSPs) are ubiquitous polypeptides expressed in all living organisms that participate in several basic cellular processes, including protein folding, from which their denomination as molecular chaperones originated. There are several HSPs, including HSPA5, also known as 78-kDa glucose-regulated protein (GRP78) or binding immunoglobulin protein (BIP) that is an ER resident involved in the folding of polypeptides during their translocation into this compartment prior to the transition to the Golgi network. HSPA5 is detected on the surface of cells or secreted into the extracellular environment. Surface HSPA5 has been proposed to have various roles, such as receptor-mediated signal transduction, a co-receptor for soluble ligands, as well as a participant in tumor survival, proliferation, and resistance. Recently, surface HSPA5 has been reported to be a potential receptor of some viruses, including the novel SARS-CoV-2. In spite of these observations, the association of HSPA5 within the plasma membrane is still unclear. To gain information about this process, we studied the interaction of HSPA5 with liposomes made of different phospholipids. We found that HSPA5 has a high affinity for negatively charged phospholipids, such as palmitoyl-oleoyl phosphoserine (POPS) and cardiolipin (CL). The N-terminal and C-terminal domains of HSPA5 were independently capable of interacting with negatively charged phospholipids, but to a lesser extent than the full-length protein, suggesting that both domains are required for the maximum insertion into membranes. Interestingly, we found that the interaction of HSPA5 with negatively charged liposomes promotes an oligomerization process via intermolecular disulfide bonds in which the N-terminus end of the protein plays a critical role.