bims-proteo 2023-12-03 papers

bims-proteo

Biomed News

on Proteostasis

Issue of 2023–12–03
thirty-one papers selected by
Eric Chevet, INSERM

Cutting through the stress: RNA decay pathways at the endoplasmic reticulum.
Small molecule correctors divert CFTR-F508del from ERAD by stabilizing sequential folding states.
Methionine Restriction Impairs Degradation of a Protein that Aberrantly Engages the Endoplasmic Reticulum Translocon.
Pervasive aggregation and depletion of host and viral proteins in response to cysteine-reactive electrophilic compounds.
System-wide analysis of RNA and protein subcellular localization dynamics.
Discovery of a Drug-like, Natural Product-Inspired DCAF11 Ligand Chemotype.
Bimodular architecture of bacterial effector SAP05 that drives ubiquitin-independent targeted protein degradation.
Lipid-anchored proteasomes control membrane protein homeostasis.
SYK ubiquitination by CBL E3 ligases restrains cross-presentation of dead cell-associated antigens by type 1 dendritic cells.
Yeast lunapark regulates the formation of trans-Sey1p complexes for homotypic ER membrane fusion.
Xport-A functions as a chaperone by stabilizing the first five transmembrane domains of rhodopsin-1.
Lysine deserts and cullin-RING ligase receptors: Navigating untrodden paths in proteostasis.
TAC-tics for Leveraging Proximity Biology in Drug Discovery.
Expression of SARS-CoV-2 Nonstructural Proteins 3 and 4 Can Tune the Unfolded Protein Response in Cell Culture.
Canonical and Non-Canonical Roles of SNX1 and SNX2 in Endosomal Membrane Dynamics.
Inhibiting the endoplasmic reticulum stress response enhances the effect of doxorubicin by altering the lipid metabolism of liver cancer cells.
The mechanism of mRNA activation.
OsHLP1 is an endoplasmic-reticulum-phagy receptor in rice plants.
NGLY1 mutations cause protein aggregation in human neurons.
UBE2J1 is the E2 ubiquitin-conjugating enzyme regulating androgen receptor degradation and antiandrogen resistance.
Coordination of RAB-8 and RAB-11 during unconventional protein secretion.
Solid-to-liquid phase transition in the dissolution of cytosolic misfolded-protein aggregates.
Endosomal Arl4A attenuates EGFR degradation by binding to the ESCRT-II component VPS36.
A simple method for developing lysine targeted covalent protein reagents.
mRNA reading frame maintenance during eukaryotic ribosome translocation.
COPI vesicle formation and N-myristoylation are targetable vulnerabilities of senescent cells.
m6A modification negatively regulates translation by switching mRNA from polysome to P-body via IGF2BP3.
Fingerprint of the oxido-reductase ERO1: A protein disulfide bond producer and supporter of cancer.
Tumor endothelial cell autophagy is a key vascular-immune checkpoint in melanoma.
Enniatin A inhibits the chaperone Hsp90 and unleashes the immune system against triple-negative breast cancer.
Biochemically functionalized probes for cell-type-specific targeting and recording in the brain.

Trends Cell Biol. 2023 Nov 25. pii: S0962-8924(23)00236-2. [Epub ahead of print]

Cutting through the stress: RNA decay pathways at the endoplasmic reticulum.

Franziska Ottens, Sotirios Efstathiou, Thorsten Hoppe.

  The endoplasmic reticulum (ER) is central to the processing of luminal, transmembrane, and secretory proteins, and maintaining a functional ER is essential for organismal physiology and health. Increased protein-folding load on the ER causes ER stress, which activates quality control mechanisms to restore ER function and protein homeostasis. Beyond protein quality control, mRNA decay pathways have emerged as potent ER fidelity regulators, but their mechanistic roles in ER quality control and their interrelationships remain incompletely understood. Herein, we review ER-associated RNA decay pathways - including regulated inositol-requiring enzyme 1α (IRE1α)-dependent mRNA decay (RIDD), nonsense-mediated mRNA decay (NMD), and Argonaute-dependent RNA silencing - in ER homeostasis, and highlight the intricate coordination of ER-targeted RNA and protein decay mechanisms and their association with antiviral defense.

Keywords:  AGO2; ER quality control; RNA degradation; UPR; proteostasis; ubiquitin

DOI:  https://doi.org/10.1016/j.tcb.2023.11.003
Mol Biol Cell. 2023 Nov 29. mbcE23080336

Small molecule correctors divert CFTR-F508del from ERAD by stabilizing sequential folding states.

Celeste Riepe, Magda Wąchalska, Kirandeep K Deol, Anais K Amaya, Matthew H Porteus, James A Olzmann, Ron R Kopito.

Over 80% of people with cystic fibrosis (CF) carry the F508del mutation in the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride ion channel at the apical plasma membrane (PM) of epithelial cells. F508del impairs CFTR folding causing it to be destroyed by endoplasmic reticulum associated degradation (ERAD). Small molecule correctors, which act as pharmacological chaperones to divert CFTR-F508del from ERAD, are the primary strategy for treating CF, yet corrector development continues with only a rudimentary understanding of how ERAD targets CFTR-F508del. We conducted genome-wide CRISPR/Cas9 knockout screens to systematically identify the molecular machinery that underlies CFTR-F508del ERAD. Although the ER-resident ubiquitin ligase, RNF5 was the top E3 hit, knocking out RNF5 only modestly reduced CFTR-F508del degradation. Sublibrary screens in an RNF5 knockout background identified RNF185 as a redundant ligase and demonstrated that CFTR-F508del ERAD is robust. Gene-drug interaction experiments illustrated that correctors tezacaftor (VX-661) and elexacaftor (VX-445) stabilize sequential, RNF5-resistant folding states. We propose that binding of correctors to nascent CFTR-F508del alters its folding landscape by stabilizing folding states that are not substrates for RNF5-mediated ubiquitylation.

DOI: https://doi.org/10.1091/mbc.E23-08-0336
MicroPubl Biol. 2023 ;2023

Methionine Restriction Impairs Degradation of a Protein that Aberrantly Engages the Endoplasmic Reticulum Translocon.

Avery M Runnebohm, Christopher J Indovina, Samantha M Turk, Connor G Bailey, Cade J Orchard, Lauren Wade, Danielle L Overton, Brian J Snow, Eric M Rubenstein.

Proteins that persistently engage endoplasmic reticulum (ER) translocons are degraded by multiple translocon quality control (TQC) mechanisms. In Saccharomyces cerevisiae , the model translocon-associated protein Deg1 -Sec62 is subject to ER-associated degradation (ERAD) by the Hrd1 ubiquitin ligase and, to a lesser extent, proteolysis mediated by the Ste24 protease. In a recent screen, we identified nine methionine-biosynthetic genes as candidate TQC regulators. Here, we found methionine restriction impairs Hrd1-independent Deg1 -Sec62 degradation. Beyond revealing methionine as a novel regulator of TQC, our results urge caution when working with laboratory yeast strains with auxotrophic mutations, often presumed not to influence cellular processes under investigation.

DOI: https://doi.org/10.17912/micropub.biology.001021
bioRxiv. 2023 Nov 16. pii: 2023.10.30.564067. [Epub ahead of print]

Pervasive aggregation and depletion of host and viral proteins in response to cysteine-reactive electrophilic compounds.

Ashley R Julio, Flowreen Shikwana, Cindy Truong, Nikolas R Burton, Emil Dominguez, Alexandra C Turmon, Jian Cao, Keriann Backus.

Protein homeostasis is tightly regulated, with damaged or misfolded proteins quickly eliminated by the proteasome and autophagosome pathways. By co-opting these processes, targeted protein degradation technologies enable pharmacological manipulation of protein abundance. Recently, cysteine-reactive molecules have been added to the degrader toolbox, which offer the benefit of unlocking the therapeutic potential of 'undruggable' protein targets. The proteome-wide impact of these molecules remains to be fully understood and given the general reactivity of many classes of cysteine-reactive electrophiles, on- and off-target effects are likely. Using chemical proteomics, we identified a cysteine-reactive small molecule degrader of the SARS-CoV-2 non- structural protein 14 (nsp14), which effects degradation through direct modification of cysteines in both nsp14 and in host chaperones together with activation of global cell stress response pathways. We find that cysteine-reactive electrophiles increase global protein ubiquitylation, trigger proteasome activation, and result in widespread aggregation and depletion of host proteins, including components of the nuclear pore complex. Formation of stress granules was also found to be a remarkably ubiquitous cellular response to nearly all cysteine-reactive compounds and degraders. Collectively, our study sheds light on complexities of covalent target protein degradation and highlights untapped opportunities in manipulating and characterizing proteostasis processes via deciphering the cysteine-centric regulation of stress response pathways.

DOI: https://doi.org/10.1101/2023.10.30.564067
Nat Methods. 2023 Nov 30.

System-wide analysis of RNA and protein subcellular localization dynamics.

Eneko Villanueva, Tom Smith, Mariavittoria Pizzinga, Mohamed Elzek, Rayner M L Queiroz, Robert F Harvey, Lisa M Breckels, Oliver M Crook, Mie Monti, Veronica Dezi, Anne E Willis, Kathryn S Lilley.

Although the subcellular dynamics of RNA and proteins are key determinants of cell homeostasis, their characterization is still challenging. Here we present an integrative framework to simultaneously interrogate the dynamics of the transcriptome and proteome at subcellular resolution by combining two methods: localization of RNA (LoRNA) and a streamlined density-based localization of proteins by isotope tagging (dLOPIT) to map RNA and protein to organelles (nucleus, endoplasmic reticulum and mitochondria) and membraneless compartments (cytosol, nucleolus and cytosolic granules). Interrogating all RNA subcellular locations at once enables system-wide quantification of the proportional distribution of RNA. We obtain a cell-wide overview of localization dynamics for 31,839 transcripts and 5,314 proteins during the unfolded protein response, revealing that endoplasmic reticulum-localized transcripts are more efficiently recruited to cytosolic granules than cytosolic RNAs, and that the translation initiation factor eIF3d is key to sustaining cytoskeletal function. Overall, we provide the most comprehensive overview so far of RNA and protein subcellular localization dynamics.

DOI: https://doi.org/10.1038/s41592-023-02101-9
Nat Commun. 2023 Nov 30. 14(1): 7908

Discovery of a Drug-like, Natural Product-Inspired DCAF11 Ligand Chemotype.

Gang Xue, Jianing Xie, Matthias Hinterndorfer, Marko Cigler, Lara Dötsch, Hana Imrichova, Philipp Lampe, Xiufen Cheng, Soheila Rezaei Adariani, Georg E Winter, Herbert Waldmann.

Targeted proteasomal and autophagic protein degradation, often employing bifunctional modalities, is a new paradigm for modulation of protein function. In an attempt to explore protein degradation by means of autophagy we combine arylidene-indolinones reported to bind the autophagy-related LC3B-protein and ligands of the PDEδ lipoprotein chaperone, the BRD2/3/4-bromodomain containing proteins and the BTK- and BLK kinases. Unexpectedly, the resulting bifunctional degraders do not induce protein degradation by means of macroautophagy, but instead direct their targets to the ubiquitin-proteasome system. Target and mechanism identification reveal that the arylidene-indolinones covalently bind DCAF11, a substrate receptor in the CUL4A/B-RBX1-DDB1-DCAF11 E3 ligase. The tempered α, β-unsaturated indolinone electrophiles define a drug-like DCAF11-ligand class that enables exploration of this E3 ligase in chemical biology and medicinal chemistry programs. The arylidene-indolinone scaffold frequently occurs in natural products which raises the question whether E3 ligand classes can be found more widely among natural products and related compounds.

DOI: https://doi.org/10.1038/s41467-023-43657-6
Proc Natl Acad Sci U S A. 2023 Dec 05. 120(49): e2310664120

Bimodular architecture of bacterial effector SAP05 that drives ubiquitin-independent targeted protein degradation.

Qun Liu, Abbas Maqbool, Federico G Mirkin, Yeshveer Singh, Clare E M Stevenson, David M Lawson, Sophien Kamoun, Weijie Huang, Saskia A Hogenhout.

  In eukaryotes, targeted protein degradation (TPD) typically depends on a series of interactions among ubiquitin ligases that transfer ubiquitin molecules to substrates leading to degradation by the 26S proteasome. We previously identified that the bacterial effector protein SAP05 mediates ubiquitin-independent TPD. SAP05 forms a ternary complex via interactions with the von Willebrand Factor Type A (vWA) domain of the proteasomal ubiquitin receptor Rpn10 and the zinc-finger (ZnF) domains of the SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE (SPL) and GATA BINDING FACTOR (GATA) transcription factors (TFs). This leads to direct TPD of the TFs by the 26S proteasome. Here, we report the crystal structures of the SAP05-Rpn10vWA complex at 2.17 Å resolution and of the SAP05-SPL5ZnF complex at 2.20 Å resolution. Structural analyses revealed that SAP05 displays a remarkable bimodular architecture with two distinct nonoverlapping surfaces, a "loop surface" with three protruding loops that form electrostatic interactions with ZnF, and a "sheet surface" featuring two β-sheets, loops, and α-helices that establish polar interactions with vWA. SAP05 binding to ZnF TFs involves single amino acids responsible for multiple contacts, while SAP05 binding to vWA is more stable due to the necessity of multiple mutations to break the interaction. In addition, positioning of the SAP05 complex on the 26S proteasome points to a mechanism of protein degradation. Collectively, our findings demonstrate how a small bacterial bimodular protein can bypass the canonical ubiquitin-proteasome proteolysis pathway, enabling ubiquitin-independent TPD in eukaryotic cells. This knowledge holds significant potential for the creation of TPD technologies.

Keywords:  26S proteasome; bacterial effector protein; phytoplasma; targeted protein degradation; ubiquitin-independent

DOI:  https://doi.org/10.1073/pnas.2310664120
Sci Adv. 2023 Dec;9(48): eadj4605

Lipid-anchored proteasomes control membrane protein homeostasis.

Ruizhu Zhang, Shuxian Pan, Suya Zheng, Qingqing Liao, Zhaodi Jiang, Dixian Wang, Xuemei Li, Ao Hu, Xinran Li, Yezhang Zhu, Xiaoqi Shen, Jing Lei, Siming Zhong, Xiaomei Zhang, Lingyun Huang, Xiaorong Wang, Lan Huang, Li Shen, Bao-Liang Song, Jing-Wei Zhao, Zhiping Wang, Bing Yang, Xing Guo.

Protein degradation in eukaryotic cells is mainly carried out by the 26S proteasome, a macromolecular complex not only present in the cytosol and nucleus but also associated with various membranes. How proteasomes are anchored to the membrane and the biological meaning thereof have been largely unknown in higher organisms. Here, we show that N-myristoylation of the Rpt2 subunit is a general mechanism for proteasome-membrane interaction. Loss of this modification in the Rpt2-G2A mutant cells leads to profound changes in the membrane-associated proteome, perturbs the endomembrane system, and undermines critical cellular processes such as cell adhesion, endoplasmic reticulum-associated degradation and membrane protein trafficking. Rpt2G2A/G2A homozygous mutation is embryonic lethal in mice and is sufficient to abolish tumor growth in a nude mice xenograft model. These findings have defined an evolutionarily conserved mechanism for maintaining membrane protein homeostasis and underscored the significance of compartmentalized protein degradation by myristoyl-anchored proteasomes in health and disease.

DOI: https://doi.org/10.1126/sciadv.adj4605
Cell Rep. 2023 Nov 28. pii: S2211-1247(23)01518-8. [Epub ahead of print]42(12): 113506

SYK ubiquitination by CBL E3 ligases restrains cross-presentation of dead cell-associated antigens by type 1 dendritic cells.

Conor M Henry, Carlos A Castellanos, Michael D Buck, Evangelos Giampazolias, Bruno Frederico, Ana Cardoso, Neil C Rogers, Oliver Schulz, Sonia Lee, Johnathan Canton, Peter Faull, Ambrosius P Snijders, Bhopal Mohapatra, Hamid Band, Caetano Reis E Sousa.

  Cross-presentation of dead cell-associated antigens by conventional dendritic cells type 1 (cDC1s) is critical for CD8+ T cells response against many tumors and viral infections. It is facilitated by DNGR-1 (CLEC9A), an SYK-coupled cDC1 receptor that detects dead cell debris. Here, we report that DNGR-1 engagement leads to rapid activation of CBL and CBL-B E3 ligases to cause K63-linked ubiquitination of SYK and terminate signaling. Genetic deletion of CBL E3 ligases or charge-conserved mutation of target lysines within SYK abolishes SYK ubiquitination and results in enhanced DNGR-1-dependent antigen cross-presentation. We also find that cDC1 deficient in CBL E3 ligases are more efficient at cross-priming CD8+ T cells to dead cell-associated antigens and promoting host resistance to tumors. Our findings reveal a role for CBL-dependent ubiquitination in limiting cross-presentation of dead cell-associated antigens and highlight an axis of negative regulation of cDC1 activity that could be exploited to increase anti-tumor immunity.

Keywords:  CBL E3 ligase; CD8(+) T cells; CP: Immunology; DNGR-1/CLEC9A; SYK; anti-tumor immunity; cross-presentation; cross-priming; dendritic cells; spleen tyrosine kinase

DOI:  https://doi.org/10.1016/j.celrep.2023.113506
iScience. 2023 Dec 15. 26(12): 108386

Yeast lunapark regulates the formation of trans-Sey1p complexes for homotypic ER membrane fusion.

Eunhong Jang, Miriam Lee, So Young Yoon, Sang Soo Lee, Jongseo Park, Mi Sun Jin, Soo Hyun Eom, Changwook Lee, Youngsoo Jun.

  The endoplasmic reticulum (ER) consists of the nuclear envelope and a connected peripheral network of tubules and interspersed sheets. The structure of ER tubules is generated and maintained by various proteins, including reticulons, DP1/Yop1p, atlastins, and lunapark. Reticulons and DP1/Yop1p stabilize the high membrane curvature of ER tubules, and atlastins mediate homotypic membrane fusion between ER tubules; however, the exact role of lunapark remains poorly characterized. Here, using isolated yeast ER microsomes and reconstituted proteoliposomes, we directly examined the function of the yeast lunapark Lnp1p for yeast atlastin Sey1p-mediated ER fusion and found that Lnp1p inhibits Sey1p-driven membrane fusion. Furthermore, by using a newly developed assay for monitoring trans-Sey1p complex assembly, a prerequisite for ER fusion, we found that assembly of trans-Sey1p complexes was increased by the deletion of LNP1 and decreased by the overexpression of Lnp1p, indicating that Lnp1p inhibits Sey1p-mediated fusion by interfering with assembly of trans-Sey1p complexes.

Keywords:  Biochemistry; Cell biology; Molecular biology

DOI:  https://doi.org/10.1016/j.isci.2023.108386
iScience. 2023 Dec 15. 26(12): 108309

Xport-A functions as a chaperone by stabilizing the first five transmembrane domains of rhodopsin-1.

Catarina J Gaspar, Tiago Gomes, Joana C Martins, Manuel N Melo, Colin Adrain, Tiago N Cordeiro, Pedro M Domingos.

  Rhodopsin-1 (Rh1), the main photosensitive protein of Drosophila, is a seven-transmembrane domain protein, which is inserted co-translationally in the endoplasmic reticulum (ER) membrane. Biogenesis of Rh1 occurs in the ER, where various chaperones interact with Rh1 to aid in its folding and subsequent transport from the ER to the rhabdomere, the light-sensing organelle of the photoreceptors. Xport-A has been proposed as a chaperone/transport factor for Rh1, but the exact molecular mechanism for Xport-A activity upon Rh1 is unknown. Here, we propose a model where Xport-A functions as a chaperone during the biogenesis of Rh1 in the ER by stabilizing the first five transmembrane domains (TMDs) of Rh1.

Keywords:  Biochemistry; Biological sciences; Molecular biology; Molecular interaction

DOI:  https://doi.org/10.1016/j.isci.2023.108309
iScience. 2023 Nov 17. 26(11): 108344

Lysine deserts and cullin-RING ligase receptors: Navigating untrodden paths in proteostasis.

Natalia A Szulc, Małgorzata Piechota, Lilla Biriczová, Pankaj Thapa, Wojciech Pokrzywa.

  The ubiquitin-proteasome system (UPS) governs the degradation of proteins by ubiquitinating their lysine residues. Our study focuses on lysine deserts - regions in proteins conspicuously low in lysine residues - in averting ubiquitin-dependent proteolysis. We spotlight the prevalence of lysine deserts among bacteria leveraging the pupylation-dependent proteasomal degradation, and in the UPS of eukaryotes. To further scrutinize this phenomenon, we focused on human receptors VHL and SOCS1 to ascertain if lysine deserts could limit their ubiquitination within the cullin-RING ligase (CRL) complex. Our data indicate that the wild-type and lysine-free variants of VHL and SOCS1 maintain consistent turnover rates, unaltered by CRL-mediated ubiquitination, hinting at a protective mechanism facilitated by lysine deserts. Nonetheless, we noted their ubiquitination at non-lysine sites, alluding to alternative regulation by the UPS. Our research underscores the role of lysine deserts in limiting CRL-mediated ubiquitin tagging while promoting non-lysine ubiquitination, thereby advancing our understanding of proteostasis.

Keywords:  Enzymology; Evolutionary biology; Molecular interaction; Properties of biomolecules; Protein

DOI:  https://doi.org/10.1016/j.isci.2023.108344
Chembiochem. 2023 Nov 28. e202300712

TAC-tics for Leveraging Proximity Biology in Drug Discovery.

Dhanusha A Nalawansha, Kyle Mangano, Willem den Besten, Patrick Ryan Potts.

  Chemically induced proximity (CIP) refers to co-opting naturally occurring biological pathways using synthetic molecules to recruit neosubstrates that are not normally encountered or to enhance the affinity of naturally occurring interactions. Leveraging proximity biology through CIPs has become a rapidly evolving field and has garnered considerable interest in basic research and drug discovery. PROteolysis Targeting Chimera (PROTAC) is a well-established CIP modality that induces the proximity between a target protein and an E3 ubiquitin ligase, causing target protein degradation via the ubiquitin-proteasome system. Inspired by PROTACs, several other induced proximity modalities have emerged to modulate both proteins and RNA over recent years. In this review, we summarize the critical advances and opportunities in the field, focusing on protein degraders, RNA degraders and non-degrader modalities such as post-translational modification (PTM) and protein-protein interaction (PPI) modulators. We envision that these emerging proximity-based drug modalities will be valuable resources for both biological research and therapeutic discovery in the future.

Keywords:  Chemically induced Proximity; Drug discovery; PROTAC; induced proximity; proximity biology

DOI:  https://doi.org/10.1002/cbic.202300712
J Proteome Res. 2023 Dec 01.

Expression of SARS-CoV-2 Nonstructural Proteins 3 and 4 Can Tune the Unfolded Protein Response in Cell Culture.

Jonathan P Davies, Athira Sivadas, Katherine R Keller, Brynn K Roman, Richard J H Wojcikiewicz, Lars Plate.

  Coronaviruses (CoV), including SARS-CoV-2, modulate host proteostasis through the activation of stress-responsive signaling pathways such as the Unfolded Protein Response (UPR), which remedies misfolded protein accumulation by attenuating translation and increasing protein folding capacity. While CoV nonstructural proteins (nsps) are essential for infection, little is known about the role of nsps in modulating the UPR. We characterized the impact of overexpression of SARS-CoV-2 nsp4, a key driver of replication, on the UPR in cell culture using quantitative proteomics to sensitively detect pathway-wide upregulation of effector proteins. We find that nsp4 preferentially activates the ATF6 and PERK branches of the UPR. Previously, we found that an N-terminal truncation of nsp3 (nsp3.1) can suppress pharmacological ATF6 activation. To determine how nsp3.1 and nsp4 tune the UPR, their coexpression demonstrated that nsp3.1 suppresses nsp4-mediated PERK, but not ATF6 activation. Reanalysis of SARS-CoV-2 infection proteomics data revealed time-dependent activation of PERK targets early in infection, which subsequently fades. This temporal regulation suggests a role for nsp3 and nsp4 in tuning the PERK pathway to attenuate host translation beneficial for viral replication while avoiding later apoptotic signaling caused by chronic activation. This work furthers our understanding of CoV-host proteostasis interactions and highlights the power of proteomic methods for systems-level analysis of the UPR.

Keywords:  ATF6; PERK; coronavirus; nonstructural protein; proteomics; stress response

DOI:  https://doi.org/10.1021/acs.jproteome.3c00600
Contact (Thousand Oaks). 2023 Jan-Dec;6:6 25152564231217867

Canonical and Non-Canonical Roles of SNX1 and SNX2 in Endosomal Membrane Dynamics.

Juliane Da Graça, Etienne Morel.

  Sorting nexins (SNXs) are a family of membrane-binding proteins known to play a critical role in regulating endocytic pathway sorting and endosomal membrane trafficking. Among them, SNX1 and SNX2 are members of the SNX-BAR subfamily and possess a membrane-curvature domain and a phosphoinositide-binding domain, which enables their stabilization at the phosphatidylinositol-3-phosphate (PI3P)-positive surface of endosomes. While their binding to PI3P-positive platforms facilitates interaction with endosomal partners and stabilization at the endosomal membrane, their SNX-BAR region is pivotal for generating membrane tubulation from endosomal compartments. In this context, their primary identified biological roles-and their partnership-are tightly associated with the retromer and endosomal SNX-BAR sorting complex for promoting exit 1 complex trafficking, facilitating the transport of cargoes from early endosomes to the secretory pathway. However, recent literature indicates that these proteins also possess biological functions in other aspects of endosomal features and sorting processes. Notably, SNX2 has been found to regulate endosome-endoplasmic reticulum (ER) contact sites through its interaction with VAP proteins at the ER membrane. Furthermore, data from our laboratory show that SNX1 and SNX2 are involved in the tubulation of early endosomes toward ER sites associated with autophagy initiation during starvation. These findings shed light on a novel role of SNXs in inter-organelle tethering and communication. In this concise review, we will explore the non-retromer functions of SNX1 and SNX2, specifically focusing on their involvement in endosomal membrane dynamics during stress sensing and autophagy-associated processes.

Keywords:  autophagy; endosomes; membrane dynamics; membrane tubulation; sorting nexins

DOI:  https://doi.org/10.1177/25152564231217867
Mol Metab. 2023 Nov 27. pii: S2212-8778(23)00180-1. [Epub ahead of print] 101846

Inhibiting the endoplasmic reticulum stress response enhances the effect of doxorubicin by altering the lipid metabolism of liver cancer cells.

Maria Kopsida, Ada Lerma Clavero, Jaafar Khaled, David Balgoma, Clara Luna-Marco, Azazul Chowdhury, Sofi Sennefelt Nyman, Fredrik Rorsman, Charlotte Ebeling Barbier, Peter Bergsten, Hans Lennernäs, Mikael Hedeland, Femke Heindryckx.

  Hepatocellular carcinoma (HCC) is characterized by a low and variable response to chemotherapeutic treatments. One contributing factor to the overall pharmacodynamics is the activation of endoplasmic reticulum (ER) stress pathways. This is a cellular stress mechanism that becomes activated when the cell´s need for protein synthesis surpasses the ER´s capacity to maintain accurate protein folding, and has been implicated in creating drug-resistance in several solid tumors.
OBJECTIVE: to identify the role of ER-stress and lipid metabolism in mediating drug response in HCC.
METHODS: By using a chemically-induced mouse model for HCC, we administered the ER-stress inhibitor 4μ8C and/or DOX twice weekly for three weeks post-tumor initiation. Histological analyses were performed alongside comprehensive molecular biology and lipidomics assessments of isolated liver samples. In vitro models, including HCC cells, spheroids, and patient-derived liver organoids were subjected to 4μ8C and/or DOX, enabling us to assess their synergistic effects on cellular viability, lipid metabolism, and oxygen consumption rate.
RESULTS: we reveal a pivotal synergy between ER-stress modulation and drug response in HCC. The inhibition of ER-stress using 4μ8C not only enhances the cytotoxic effect of DOX, but also significantly reduces cellular lipid metabolism. This intricate interplay culminates in the deprivation of energy reserves essential for the sustenance of tumor cells.
CONCLUSIONS: This study elucidates the interplay between lipid metabolism and ER-stress modulation in enhancing doxorubicin efficacy in HCC. This novel approach not only deepens our understanding of the disease, but also uncovers a promising avenue for therapeutic innovation. The long-term impact of our study could open the possibility of ER-stress inhibitors and/or lipase inhibitors as adjuvant treatments for HCC-patients.

Keywords:  Lipidomics; chemotherapy; endoplasmic reticulum stress; hepatocellular carcinoma

DOI:  https://doi.org/10.1016/j.molmet.2023.101846
bioRxiv. 2023 Nov 15. pii: 2023.11.15.567265. [Epub ahead of print]

The mechanism of mRNA activation.

Riley C Gentry, Nicholas A Ide, Victoria M Comunale, Erik W Hartwick, Colin D Kinz-Thompson, Ruben L Gonzalez.

During translation initiation, messenger RNA molecules must be identified and activated for loading into a ribosome. In this rate-limiting step, the heterotrimeric protein eukaryotic initiation factor eIF4F must recognize and productively interact with the 7-methylguanosine cap at the 5' end of the messenger RNA and subsequently activate the message. Despite its fundamental, regulatory role in gene expression, the molecular events underlying cap recognition and messenger RNA activation remain mysterious. Here, we generate a unique, single-molecule fluorescence imaging system to interrogate the dynamics with which eIF4F discriminates productive and non-productive locations on full-length, native messenger RNA molecules. At the single-molecule level, we observe stochastic sampling of eIF4F along the length of the messenger RNA and identify allosteric communication between the eIF4F subunits which ultimately drive cap-recognition and subsequent activation of the message. Our experiments uncover novel functions for each subunit of eIF4F and we conclude by presenting a model for messenger RNA activation which precisely defines the composition of the activated message. This model provides a general framework for understanding how messenger RNA molecules may be discriminated from one another, and how other RNA-binding proteins may control the efficiency of translation initiation.

DOI: https://doi.org/10.1101/2023.11.15.567265
Cell Rep. 2023 Nov 28. pii: S2211-1247(23)01492-4. [Epub ahead of print]42(12): 113480

OsHLP1 is an endoplasmic-reticulum-phagy receptor in rice plants.

Yingbo Liang, Fanwei Meng, Xia Zhao, Xinyi He, Jun Liu.

  The endoplasmic reticulum (ER) is the largest intracellular endomembrane system; it shows dynamic changes upon environmental stress. To maintain ER morphology and homeostasis under stress, the excessive ER membrane and the associated unwanted proteins can be removed via ER-phagy. Although a few ER-phagy receptors have been reported in mammals and yeast, their functional counterparts in plants remain largely unexplored. Here, we report that the HVA22 family protein OsHLP1 is an uncharacterized ER-phagy receptor in rice (Oryza sativa L.). OsHLP1 interacts with OsATG8b and recruits ER subdomains and the cargo protein OsNTL6, a negative immune regulator, to autophagosomes upon infection with the fungus Magnaporthe oryzae, which substantially activates disease resistance in rice. AtHVA22J, an Arabidopsis thaliana OsHLP1 ortholog, induced similar ER-phagy in plants. Altogether, we unraveled a conservative protein family that may act as ER-phagy receptors in higher plants, and in particular, we highlighted their roles in rice immune responses.

Keywords:  CP: Plants; ER-phagy; OsHLP1; OsNTL6; blast disease; rice

DOI:  https://doi.org/10.1016/j.celrep.2023.113480
Cell Rep. 2023 Nov 30. pii: S2211-1247(23)01478-X. [Epub ahead of print]42(12): 113466

NGLY1 mutations cause protein aggregation in human neurons.

Andreea Manole, Thomas Wong, Amanda Rhee, Sammy Novak, Shao-Ming Chin, Katya Tsimring, Andres Paucar, April Williams, Traci Fang Newmeyer, Simon T Schafer, Idan Rosh, Susmita Kaushik, Rene Hoffman, Songjie Chen, Guangwen Wang, Michael Snyder, Ana Maria Cuervo, Leo Andrade, Uri Manor, Kevin Lee, Jeffrey R Jones, Shani Stern, Maria C Marchetto, Fred H Gage.

  Biallelic mutations in the gene that encodes the enzyme N-glycanase 1 (NGLY1) cause a rare disease with multi-symptomatic features including developmental delay, intellectual disability, neuropathy, and seizures. NGLY1's activity in human neural cells is currently not well understood. To understand how NGLY1 gene loss leads to the specific phenotypes of NGLY1 deficiency, we employed direct conversion of NGLY1 patient-derived induced pluripotent stem cells (iPSCs) to functional cortical neurons. Transcriptomic, proteomic, and functional studies of iPSC-derived neurons lacking NGLY1 function revealed several major cellular processes that were altered, including protein aggregate-clearing functionality, mitochondrial homeostasis, and synaptic dysfunctions. These phenotypes were rescued by introduction of a functional NGLY1 gene and were observed in iPSC-derived mature neurons but not astrocytes. Finally, laser capture microscopy followed by mass spectrometry provided detailed characterization of the composition of protein aggregates specific to NGLY1-deficient neurons. Future studies will harness this knowledge for therapeutic development.

Keywords:  CP: Neuroscience; NGLY1 deficiency; chaperones; fragmented mitochondria; neural cells; organoids; protein aggregates

DOI:  https://doi.org/10.1016/j.celrep.2023.113466
Oncogene. 2023 Nov 29.

UBE2J1 is the E2 ubiquitin-conjugating enzyme regulating androgen receptor degradation and antiandrogen resistance.

Carla Rodriguez Tirado, Choushi Wang, Xiaoling Li, Su Deng, Julisa Gonzalez, Nickolas A Johnson, Yaru Xu, Lauren A Metang, Medha Sundar Rajan, Yuqiu Yang, Yi Yin, Mia Hofstad, Ganesh V Raj, Song Zhang, Andrew Lemoff, Wei He, Jie Fan, Yunguan Wang, Tao Wang, Ping Mu.

Prostate cancer (PCa) is primarily driven by aberrant Androgen Receptor (AR) signaling. Although there has been substantial advancement in antiandrogen therapies, resistance to these treatments remains a significant obstacle, often marked by continuous or enhanced AR signaling in resistant tumors. While the dysregulation of the ubiquitination-based protein degradation process is instrumental in the accumulation of oncogenic proteins, including AR, the molecular mechanism of ubiquitination-driven AR degradation remains largely undefined. We identified UBE2J1 as the critical E2 ubiquitin-conjugating enzyme responsible for guiding AR ubiquitination and eventual degradation. The absence of UBE2J1, found in 5-15% of PCa patients, results in disrupted AR ubiquitination and degradation. This disruption leads to an accumulation of AR proteins, promoting resistance to antiandrogen treatments. By employing a ubiquitination-based AR degrader to adeptly restore AR ubiquitination, we reestablished AR degradation and inhibited the proliferation of antiandrogen-resistant PCa tumors. These findings underscore the fundamental role of UBE2J1 in AR degradation and illuminate an uncharted mechanism through which PCa maintains heightened AR protein levels, fostering resistance to antiandrogen therapies.

DOI: https://doi.org/10.1038/s41388-023-02890-5
J Cell Biol. 2024 Feb 05. pii: e202306107. [Epub ahead of print]223(2):

Coordination of RAB-8 and RAB-11 during unconventional protein secretion.

Xinxin Li, Bowen Liu, Yue Wen, Jiabin Wang, Yusong R Guo, Anbing Shi, Long Lin.

Multiple physiology-pertinent transmembrane proteins reach the cell surface via the Golgi-bypassing unconventional protein secretion (UcPS) pathway. By employing C. elegans-polarized intestine epithelia, we recently have revealed that the small GTPase RAB-8/Rab8 serves as an important player in the process. Nonetheless, its function and the relevant UcPS itinerary remain poorly understood. Here, we show that deregulated RAB-8 activity resulted in impaired apical UcPS, which increased sensitivity to infection and environmental stress. We also identified the SNARE VTI-1/Vti1a/b as a new RAB-8-interacting factor involved in the apical UcPS. Besides, RAB-11/Rab11 was capable of recruiting RABI-8/Rabin8 to reduce the guanine nucleotide exchange activity of SMGL-1/GEF toward RAB-8, indicating the necessity of a finely tuned RAB-8/RAB-11 network. Populations of RAB-8- and RAB-11-positive endosomal structures containing the apical UcPS cargo moved toward the apical side. In the absence of RAB-11 or its effectors, the cargo was retained in RAB-8- and RAB-11-positive endosomes, respectively, suggesting that these endosomes are utilized as intermediate carriers for the UcPS.

DOI: https://doi.org/10.1083/jcb.202306107
iScience. 2023 Dec 15. 26(12): 108334

Solid-to-liquid phase transition in the dissolution of cytosolic misfolded-protein aggregates.

Alexis Tomaszewski, Rebecca Wang, Eduardo Sandoval, Jin Zhu, Jian Liu, Rong Li.

  Accumulation of protein aggregates is a hallmark of cellular aging and degenerative disorders. This could result from either increased protein misfolding and aggregation or impaired dissolution of aggregates formed under stress, the latter of which is poorly understood. In this study, we employed quantitative live-cell imaging to investigate the dynamic process of protein disaggregation in yeast. We show that protein aggregates formed upon heat stress are solid condensates, but after stress attenuation these protein aggregates first transition into a liquid-like state during their dissolution. This solid-to-liquid phase transition (SLPT) accompanies the reduction in aggregate number due to the fusion of the liquid condensates. The chaperone activity of Hsp104, a Clp/HSP100 family chaperone, is required for both SLPT and subsequent dispersal of the liquid condensates. Sse1, a yeast HSP110 chaperone, also facilitates SLPT. These results illuminate an unexpected mechanistic framework of cellular control over protein disaggregation upon stress attenuation.

Keywords:  Biochemistry; Biological sciences; Cell biology

DOI:  https://doi.org/10.1016/j.isci.2023.108334
Nat Commun. 2023 Nov 29. 14(1): 7859

Endosomal Arl4A attenuates EGFR degradation by binding to the ESCRT-II component VPS36.

Shin-Jin Lin, Ming-Chieh Lin, Tsai-Jung Liu, Yueh-Tso Tsai, Ming-Ting Tsai, Fang-Jen S Lee.

Ligand-induced epidermal growth factor receptor (EGFR) endocytosis followed by endosomal EGFR signaling and lysosomal degradation plays important roles in controlling multiple biological processes. ADP-ribosylation factor (Arf)-like protein 4 A (Arl4A) functions at the plasma membrane to mediate cytoskeletal remodeling and cell migration, whereas its localization at endosomal compartments remains functionally unknown. Here, we report that Arl4A attenuates EGFR degradation by binding to the endosomal sorting complex required for transport (ESCRT)-II component VPS36. Arl4A plays a role in prolonging the duration of EGFR ubiquitinylation and deterring endocytosed EGFR transport from endosomes to lysosomes under EGF stimulation. Mechanistically, the Arl4A-VPS36 direct interaction stabilizes VPS36 and ESCRT-III association, affecting subsequent recruitment of deubiquitinating-enzyme USP8 by CHMP2A. Impaired Arl4A-VPS36 interaction enhances EGFR degradation and clearance of EGFR ubiquitinylation. Together, we discover that Arl4A negatively regulates EGFR degradation by binding to VPS36 and attenuating ESCRT-mediated late endosomal EGFR sorting.

DOI: https://doi.org/10.1038/s41467-023-42979-9
Nat Commun. 2023 Dec 01. 14(1): 7933

A simple method for developing lysine targeted covalent protein reagents.

Ronen Gabizon, Barr Tivon, Rambabu N Reddi, Maxime C M van den Oetelaar, Hadar Amartely, Peter J Cossar, Christian Ottmann, Nir London.

Peptide-based covalent probes can target shallow protein surfaces not typically addressable using small molecules, yet there is a need for versatile approaches to convert native peptide sequences into covalent binders that can target a broad range of residues. Here we report protein-based thio-methacrylate esters-electrophiles that can be installed easily on unprotected peptides and proteins via cysteine side chains, and react efficiently and selectively with cysteine and lysine side chains on the target. Methacrylate phosphopeptides derived from 14-3-3-binding proteins irreversibly label 14-3-3σ via either lysine or cysteine residues, depending on the position of the electrophile. Methacrylate peptides targeting a conserved lysine residue exhibit pan-isoform binding of 14-3-3 proteins both in lysates and in extracellular media. Finally, we apply this approach to develop protein-based covalent binders. A methacrylate-modified variant of the colicin E9 immunity protein irreversibly binds to the E9 DNAse, resulting in significantly higher thermal stability relative to the non-covalent complex. Our approach offers a simple and versatile route to convert peptides and proteins into potent covalent binders.

DOI: https://doi.org/10.1038/s41467-023-42632-5
Nature. 2023 Nov 29.

mRNA reading frame maintenance during eukaryotic ribosome translocation.

Nemanja Milicevic, Lasse Jenner, Alexander Myasnikov, Marat Yusupov, Gulnara Yusupova.

One of the most critical steps of protein synthesis is coupled translocation of messenger RNA (mRNA) and transfer RNAs (tRNAs) required to advance the mRNA reading frame by one codon. In eukaryotes, translocation is accelerated and its fidelity is maintained by elongation factor 2 (eEF2)1,2. At present, only a few snapshots of eukaryotic ribosome translocation have been reported3-5. Here we report ten high-resolution cryogenic-electron microscopy (cryo-EM) structures of the elongating eukaryotic ribosome bound to the full translocation module consisting of mRNA, peptidyl-tRNA and deacylated tRNA, seven of which also contained ribosome-bound, naturally modified eEF2. This study recapitulates mRNA-tRNA2-growing peptide module progression through the ribosome, from the earliest states of eEF2 translocase accommodation until the very late stages of the process, and shows an intricate network of interactions preventing the slippage of the translational reading frame. We demonstrate how the accuracy of eukaryotic translocation relies on eukaryote-specific elements of the 80S ribosome, eEF2 and tRNAs. Our findings shed light on the mechanism of translation arrest by the anti-fungal eEF2-binding inhibitor, sordarin. We also propose that the sterically constrained environment imposed by diphthamide, a conserved eukaryotic posttranslational modification in eEF2, not only stabilizes correct Watson-Crick codon-anticodon interactions but may also uncover erroneous peptidyl-tRNA, and therefore contribute to higher accuracy of protein synthesis in eukaryotes.

DOI: https://doi.org/10.1038/s41586-023-06780-4
Nat Cell Biol. 2023 Nov 27.

COPI vesicle formation and N-myristoylation are targetable vulnerabilities of senescent cells.

Domhnall McHugh, Bin Sun, Carmen Gutierrez-Muñoz, Fernanda Hernández-González, Massimiliano Mellone, Romain Guiho, Imanol Duran, Joaquim Pombo, Federico Pietrocola, Jodie Birch, Wouter W Kallemeijn, Sanjay Khadayate, Gopuraja Dharmalingam, Santiago Vernia, Edward W Tate, Juan Pedro Martínez-Barbera, Dominic J Withers, Gareth J Thomas, Manuel Serrano, Jesús Gil.

Drugs that selectively kill senescent cells (senolytics) improve the outcomes of cancer, fibrosis and age-related diseases. Despite their potential, our knowledge of the molecular pathways that affect the survival of senescent cells is limited. To discover senolytic targets, we performed RNAi screens and identified coatomer complex I (COPI) vesicle formation as a liability of senescent cells. Genetic or pharmacological inhibition of COPI results in Golgi dispersal, dysfunctional autophagy, and unfolded protein response-dependent apoptosis of senescent cells, and knockdown of COPI subunits improves the outcomes of cancer and fibrosis in mouse models. Drugs targeting COPI have poor pharmacological properties, but we find that N-myristoyltransferase inhibitors (NMTi) phenocopy COPI inhibition and are potent senolytics. NMTi selectively eliminated senescent cells and improved outcomes in models of cancer and non-alcoholic steatohepatitis. Our results suggest that senescent cells rely on a hyperactive secretory apparatus and that inhibiting trafficking kills senescent cells with the potential to treat various senescence-associated diseases.

DOI: https://doi.org/10.1038/s41556-023-01287-6
Mol Cell. 2023 Nov 24. pii: S1097-2765(23)00911-5. [Epub ahead of print]

m6A modification negatively regulates translation by switching mRNA from polysome to P-body via IGF2BP3.

Ting Shan, Feiyan Liu, Miaomiao Wen, Zonggui Chen, Shaopeng Li, Yafen Wang, Hong Cheng, Yu Zhou.

  In the cytoplasm, mRNAs are dynamically partitioned into translating and non-translating pools, but the mechanism for this regulation has largely remained elusive. Here, we report that m6A regulates mRNA partitioning between polysome and P-body where a pool of non-translating mRNAs resides. By quantifying the m6A level of polysomal and cytoplasmic mRNAs with m6A-LAIC-seq and m6A-LC-MS/MS in HeLa cells, we observed that polysome-associated mRNAs are hypo-m6A-methylated, whereas those enriched in P-body are hyper-m6A-methylated. Downregulation of the m6A writer METTL14 enhances translation by switching originally hyper-m6A-modified mRNAs from P-body to polysome. Conversely, by proteomic analysis, we identify a specific m6A reader IGF2BP3 enriched in P-body, and via knockdown and molecular tethering assays, we demonstrate that IGF2BP3 is both necessary and sufficient to switch target mRNAs from polysome to P-body. These findings suggest a model for the dynamic regulation of mRNA partitioning between the translating and non-translating pools in an m6A-dependent manner.

Keywords:  IGF2BP3; P-body; m(6)A modification; mRNA partitioning; polysome profiling; translation

DOI:  https://doi.org/10.1016/j.molcel.2023.10.040
Biochim Biophys Acta Rev Cancer. 2023 Nov 24. pii: S0304-419X(23)00176-2. [Epub ahead of print]1879(1): 189027

Fingerprint of the oxido-reductase ERO1: A protein disulfide bond producer and supporter of cancer.

Ester Zito, Luca Guarrera, Yvonne M W Janssen-Heininger.

  Endoplasmic reticulum oxidoreductin 1 (ERO1) alpha (ERO1A) is an endoplasmic reticulum (ER)-localized protein disulfide oxidoreductase, involved in the disulfide bond formation of proteins. ERO1's activity in oxidative protein folding is redundant in higher eukaryotes and its loss is well compensated. Although it is dispensable in non-cancer cells, high ERO1 levels are seen with different cancers and predict their malignant phenotype. ERO1 fosters tumor aggressiveness and the response to drug therapy in hypoxic and highly metastatic tumors. It regulates vascular endothelial growth factor (VEGF) levels, oxidative folding and N-glycosylation in hypoxic conditions, boosting tumor fitness and angiogenesis on multiple levels. In addition, ERO1 regulates protein death ligand-1 (PD-L1) on tumors, interfering with the related immune surveillance mechanism, hence acting on the tumors' response to immune check-point inhibitors (ICI). This all points to inhibition of ERO1 as an effective pharmacological tool, selectively targeting tumors while sparing non-cancer cells from cytotoxicity. The critical discussion here closely examines the molecular basis for ERO1's involvement in tumors and ERO1 inhibition strategies for their treatment.

Keywords:  Angiogenesis; ERO1 alpha; Endoplasmic reticulum stress; Hypoxia; Metastasis; UPR (unfolded protein response); cancer

DOI:  https://doi.org/10.1016/j.bbcan.2023.189027
EMBO Mol Med. 2023 Nov 27. e18028

Tumor endothelial cell autophagy is a key vascular-immune checkpoint in melanoma.

Jelle Verhoeven, Kathryn A Jacobs, Francesca Rizzollo, Francesca Lodi, Yichao Hua, Joanna Poźniak, Adhithya Narayanan Srinivasan, Diede Houbaert, Gautam Shankar, Sanket More, Marco B Schaaf, Nikolina Dubroja Lakic, Maarten Ganne, Jochen Lamote, Johan Van Weyenbergh, Louis Boon, Oliver Bechter, Francesca Bosisio, Yasuo Uchiyama, Mathieu Jm Bertrand, Jean Christophe Marine, Diether Lambrechts, Gabriele Bergers, Madhur Agrawal, Patrizia Agostinis.

  Tumor endothelial cells (TECs) actively repress inflammatory responses and maintain an immune-excluded tumor phenotype. However, the molecular mechanisms that sustain TEC-mediated immunosuppression remain largely elusive. Here, we show that autophagy ablation in TECs boosts antitumor immunity by supporting infiltration and effector function of T-cells, thereby restricting melanoma growth. In melanoma-bearing mice, loss of TEC autophagy leads to the transcriptional expression of an immunostimulatory/inflammatory TEC phenotype driven by heightened NF-kB and STING signaling. In line, single-cell transcriptomic datasets from melanoma patients disclose an enriched InflammatoryHigh /AutophagyLow TEC phenotype in correlation with clinical responses to immunotherapy, and responders exhibit an increased presence of inflamed vessels interfacing with infiltrating CD8+ T-cells. Mechanistically, STING-dependent immunity in TECs is not critical for the immunomodulatory effects of autophagy ablation, since NF-kB-driven inflammation remains functional in STING/ATG5 double knockout TECs. Hence, our study identifies autophagy as a principal tumor vascular anti-inflammatory mechanism dampening melanoma antitumor immunity.

Keywords:  autophagy; cancer; immunotherapy; inflammation; tumor endothelial cells

DOI:  https://doi.org/10.15252/emmm.202318028
iScience. 2023 Dec 15. 26(12): 108308

Enniatin A inhibits the chaperone Hsp90 and unleashes the immune system against triple-negative breast cancer.

Nada H Eisa, Vincent M Crowley, Asif Elahi, Vamsi Krishna Kommalapati, Michael A Serwetnyk, Taoufik Llbiyi, Sumin Lu, Kashish Kainth, Yasmeen Jilani, Daniela Marasco, Abdeljabar El Andaloussi, Sukyeong Lee, Francis T F Tsai, Paulo C Rodriguez, David Munn, Esteban Celis, Hasan Korkaya, Abdessamad Debbab, Brian Blagg, Ahmed Chadli.

  Low response rates and immune-related adverse events limit the remarkable impact of cancer immunotherapy. To improve clinical outcomes, preclinical studies have shown that combining immunotherapies with N-terminal Hsp90 inhibitors resulted in improved efficacy, even though induction of an extensive heat shock response (HSR) and less than optimal dosing of these inhibitors limited their clinical efficacy as monotherapies. We discovered that the natural product Enniatin A (EnnA) targets Hsp90 and destabilizes its client oncoproteins without inducing an HSR. EnnA triggers immunogenic cell death in triple-negative breast cancer (TNBC) syngeneic mouse models and exhibits superior antitumor activity compared to Hsp90 N-terminal inhibitors. EnnA reprograms the tumor microenvironment (TME) to promote CD8+ T cell-dependent antitumor immunity by reducing PD-L1 levels and activating the chemokine receptor CX3CR1 pathway. These findings provide strong evidence for transforming the immunosuppressive TME into a more tumor-hostile milieu by engaging Hsp90 with therapeutic agents involving novel mechanisms of action.

Keywords:  Biological sciences; Cancer; Microenvironment; Molecular biology

DOI:  https://doi.org/10.1016/j.isci.2023.108308
Sci Adv. 2023 Dec;9(48): eadk1050

Biochemically functionalized probes for cell-type-specific targeting and recording in the brain.

Anqi Zhang, Theodore J Zwang, Charles M Lieber.

Selective targeting and modulation of distinct cell types and neuron subtypes is central to understanding complex neural circuitry and could enable electronic treatments that target specific circuits while minimizing off-target effects. However, current brain-implantable electronics have not yet achieved cell-type specificity. We address this challenge by functionalizing flexible mesh electronic probes, which elicit minimal immune response, with antibodies or peptides to target specific cell markers. Histology studies reveal selective association of targeted neurons, astrocytes, and microglia with functionalized probe surfaces without accumulating off-target cells. In vivo chronic electrophysiology further yields recordings consistent with selective targeting of these cell types. Last, probes functionalized to target dopamine receptor 2 expressing neurons show the potential for neuron-subtype-specific targeting and electrophysiology.

DOI: https://doi.org/10.1126/sciadv.adk1050