The natural history of Get3‐like chaperones

Farkas, Ákos; Laurentiis, Evelina Ines De; Schwappach, Blanche

doi:10.1111/tra.12643

Cited by 23 publications

(26 citation statements)

References 53 publications

(126 reference statements)

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“…Thus, the cytosolic interaction, and hence the diffuse staining pattern observed for substrates in the presence of TRC40 D74E , correlated with an intact TA-binding groove, which is consistent with our rationale to stabilize the TA-protein-interacting form of TRC40. It should, however, be noted that much less is currently known about the molecular details of how Get3-like chaperones interact with their clients (Farkas et al, 2019; Voth et al, 2014). Hence, we cannot exclude that some of the effects of overexpressing TRC40 D74E may not directly relate to membrane targeting of TA protein precursors during biogenesis.…”

Section: Resultsmentioning

confidence: 99%

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A trap mutant reveals the physiological client spectrum of TRC40

Coy-Vergara

Rivera‐Monroy

Urlaub

et al. 2019

Journal of Cell Science

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The transmembrane recognition complex (TRC) pathway targets tail-anchored (TA) proteins to the membrane of the endoplasmic reticulum (ER). While many TA proteins are known to be able to use this pathway, it is essential for the targeting of only a few. Here, we uncover a large number of TA proteins that engage with TRC40 when other targeting machineries are fully operational. We use a dominant-negative ATPase-impaired mutant of TRC40 in which aspartate 74 was replaced by a glutamate residue to trap TA proteins in the cytoplasm. Manipulation of the hydrophobic TA-binding groove in TRC40 (also known as ASNA1) reduces interaction with most, but not all, substrates suggesting that co-purification may also reflect interactions unrelated to precursor protein targeting. We confirm known TRC40 substrates and identify many additional TA proteins interacting with TRC40. By using the trap approach in combination with quantitative mass spectrometry, we show that Golgi-resident TA proteins such as the golgins golgin-84, CASP and giantin as well as the vesicle-associated membrane-protein-associated proteins VAPA and VAPB interact with TRC40. Thus, our results provide new avenues to assess the essential role of TRC40 in metazoan organisms.

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Section: Resultsmentioning

confidence: 99%

“…At the same time, our data suggest that the observed co-purification of TA proteins with TRC40 may partially reflect a molecular interaction unrelated to precursor protein targeting. Hence, our analysis motivates future focus on the chaperone function of proteins homologous to TRC40 and its yeast counterpart Get3 (Farkas et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

A trap mutant reveals the physiological client spectrum of TRC40

Coy-Vergara

Rivera‐Monroy

Urlaub

et al. 2019

Journal of Cell Science

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“…C. elegans asna-1 has all the structural features required for TAP insertion 10 , 36 , 37 . However, since not all ASNA-1 homologs have TAP-targeting activity 38 , it was important to examine whether C. elegans ASNA-1 has this function. In addition, parallel pathways like EMC, HSP40/HSC70, and SND participate in TAP targeting 23 , 39 – 41 .…”

Section: Discussionmentioning

confidence: 99%

Alternative redox forms of ASNA-1 separate insulin signaling from tail-anchored protein targeting and cisplatin resistance in C. elegans

Raj

Billing

Podraza-Farhanieh

et al. 2021

Sci Rep

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Cisplatin is a frontline cancer therapeutic, but intrinsic or acquired resistance is common. We previously showed that cisplatin sensitivity can be achieved by inactivation of ASNA-1/TRC40 in mammalian cancer cells and in Caenorhabditis elegans. ASNA-1 has two more conserved functions: in promoting tail-anchored protein (TAP) targeting to the endoplasmic reticulum membrane and in promoting insulin secretion. However, the relation between its different functions has remained unknown. Here, we show that ASNA-1 exists in two redox states that promote TAP-targeting and insulin secretion separately. The reduced state is the one required for cisplatin resistance: an ASNA-1 point mutant, in which the protein preferentially was found in the oxidized state, was sensitive to cisplatin and defective for TAP targeting but had no insulin secretion defect. The same was true for mutants in wrb-1, which we identify as the C. elegans homolog of WRB, the ASNA1/TRC40 receptor. Finally, we uncover a previously unknown action of cisplatin induced reactive oxygen species: cisplatin induced ROS drives ASNA-1 into the oxidized form, and selectively prevents an ASNA-1-dependent TAP substrate from reaching the endoplasmic reticulum. Our work suggests that ASNA-1 acts as a redox-sensitive target for cisplatin cytotoxicity and that cisplatin resistance is likely mediated by ASNA-1-dependent TAP substrates. Treatments that promote an oxidizing tumor environment should be explored as possible means to combat cisplatin resistance.

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“…C. elegans asna-1 has all the structural features required for TAP insertion (Chio et al, 2019)(Mateja and Keenan, 2018; Mateja et al, 2009). However, since not all ASNA-1 homologs have TAP-targeting activity (Farkas et al, 2019), it was important to examine whether C. elegans ASNA-1 has this function. In addition, parallel pathways like EMC, HSP70/HSC40, and SND participate in TAP targeting (Aviram et al, 2016; Casson et al, 2017; Cho and Shan, 2018; Guna et al, 2018).…”

Section: Discussionmentioning

confidence: 99%

ASNA-1 oxidation induced by cisplatin exposure enhances its cytotoxicity by selectively perturbing tail anchored protein targeting

Raj

Billing

Podraza

et al. 2019

Preprint

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protein SUMMARY STATEMENTSensitizing tumors to cisplatin would be of considerable therapeutic benefit. Here we show a novel mechanism of cisplatin sensitization via oxidation of ASNA-1 in a Caenorhabditis elegans model. ABSTRACTCisplatin is a frontline cancer treatment, but intrinsic or acquired resistance is common. We previously showed that ASNA-1/TRC40 inactivation increases cisplatin sensitivity in mammalian cells and a Caenorhabditis elegans asna-1 knockdown model. ASNA-1 has conserved tail-anchored protein (TAP) targeting and insulin secretion functions. Here we examined the mechanism of ASNA-1 action. We show that ASNA-1 exists in two physiologically-responsive redox states with separable TAP-targeting and insulin secretion functions. Cisplatin-generated ROS targeted ASNA-1 oxidation, resulting in a selective targeting defect of an ASNA-1-dependent TAP. Increased ASNA-1 oxidation sensitized worms to cisplatin cytotoxicity. Mutants with a redox balance favoring oxidized ASNA-1 were cisplatin sensitive as null mutants by diverting ASNA-1 away from its TAP-targeting role and 2 instead perturbing endoplasmic reticulum (ER) function. Mutations in the ASNA-1 receptor required for TAP insertion induced equivalent cisplatin sensitivity. We reveal a previously undescribed cellular dysfunction induced by cisplatin, identify a cisplatin target, and show that drug exposure causes TAP targeting-induced ER dysfunction. Therapeutic oxidation of ASNA-1 could be a clinically useful means to increase cisplatin sensitivity, reduce cytotoxic drug doses, and counteract cisplatin resistance. AUTHOR SUMMARYCisplatin is a very effective anti-cancer drug and is widely used as a frontline treatment.However, tumor resistance limits its use. Tumor re-sensitization would improve cancer treatment. ASNA-1/TRC40 knockdown in Caenorhabditis elegans and mammals results in cisplatin hypersensitivity, but the underlying mechanistic details are largely unknown. We show that in C. elegans ASNA-1 mutants, increased cisplatin killing is coupled with delocalization of a tail-anchored protein, SEC-61b, a membrane protein that should reach the ER and is instead mistargeted. Like its homologs, the reduced form of worm ASNA-1 is needed for targeting activity. Targeting is blocked upon ASNA-1 oxidation after cisplatin treatment, likely via reactive oxygen species (ROS) generated by cisplatin treatment. Nevertheless, the oxidized form of the protein can execute other functions like insulin secretion. We show also that mutants with high oxidized ASNA-1 levels are cisplatin sensitive. Additionally, cisplatin induced mistargeting strictly acts through ASNA-1 inactivation. Thus, we define a pathway from cisplatin exposure that targets protein (ASNA-1) inactivation, consequently leading to mis-targeting of proteins that need ASNA-1 for their maturation. This multi-step process provides vital information about likely proteins that can be targeted by drugs to enhance cisplatin mediated killing and improve chemotherapy.

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The natural history of Get3‐like chaperones

Cited by 23 publications

References 53 publications

A trap mutant reveals the physiological client spectrum of TRC40

A trap mutant reveals the physiological client spectrum of TRC40

Alternative redox forms of ASNA-1 separate insulin signaling from tail-anchored protein targeting and cisplatin resistance in C. elegans

ASNA-1 oxidation induced by cisplatin exposure enhances its cytotoxicity by selectively perturbing tail anchored protein targeting

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