Substrate relay in an Hsp70‐cochaperone cascade safeguards tail‐anchored membrane protein targeting

Cho, Hyunjun; Shan, Shu‐ou

doi:10.15252/embj.201899264

Cited by 47 publications

(128 citation statements)

References 61 publications

Supporting

Mentioning

120

Contrasting

Order By: Relevance

“…To test the effect of the SR MoRF deletion on SRP-dependent protein targeting, we measured the in vivo targeting and translocation of a model substrate, DHC-αF, in which the signal sequence of prepro-α-factor is replaced by the hydrophobic core of the dipeptidyl aminopeptidase B signal sequence to convert it into an SRP-dependent substrate protein (Fig. S2 D; Ng et al, 1996; Cho and Shan, 2018). DHC-αF was efficiently glycosylated upon insertion into the ER (Yabal et al, 2003; Wang et al, 2010; Rao et al, 2016), providing a quantitative readout for its targeting and translocation.…”

Section: Resultsmentioning

confidence: 99%

“…DHC-αF was efficiently glycosylated upon insertion into the ER (Yabal et al, 2003; Wang et al, 2010; Rao et al, 2016), providing a quantitative readout for its targeting and translocation. To measure the targeting kinetics of newly synthesized proteins, we performed pulse-chase assays coupled to immunoprecipitation of HA-tagged substrate proteins (see Materials and methods; Cho and Shan, 2018). The results showed that, while DHC-αF was rapidly and nearly completely translocated in SRP101 cells, the translocation of DHC-αF was substantially delayed and plateaued at <50% in srp101 dM cells (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The following observations suggested additional defects in srp101 dM cells. In control reactions, we tested the insertion of a posttranslationally targeted model protein substrate, BirA-Bos1, into the ER (Cho and Shan, 2018). BirA-Bos1 is a model tail-anchored membrane protein substrate in which the transmembrane domain of the SNARE protein Bos1 is fused to the C-terminus of BirA (Fig.…”

Section: Resultsmentioning

confidence: 99%

See 2 more Smart Citations

A molecular recognition feature mediates ribosome-induced SRP-receptor assembly during protein targeting

Chandrasekar

Lee

et al. 2019

Journal of Cell Biology

Self Cite

View full text Add to dashboard Cite

Molecular recognition features (MoRFs) provide interaction motifs in intrinsically disordered protein regions to mediate diverse cellular functions. Here we report that a MoRF element, located in the disordered linker domain of the mammalian signal recognition particle (SRP) receptor and conserved among eukaryotes, plays an essential role in sensing the ribosome during cotranslational protein targeting to the endoplasmic reticulum. Loss of the MoRF in the SRP receptor (SR) largely abolishes the ability of the ribosome to activate SRP-SR assembly and impairs cotranslational protein targeting. These results demonstrate a novel role for MoRF elements and provide a mechanism for the ribosome-induced activation of the mammalian SRP pathway. Kinetic analyses and comparison with the bacterial SRP further suggest that the SR MoRF functionally replaces the essential GNRA tetraloop in the bacterial SRP RNA, providing an example for the replacement of RNA function by proteins during the evolution of ancient ribonucleoprotein particles.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

A molecular recognition feature mediates ribosome-induced SRP-receptor assembly during protein targeting

Chandrasekar

Lee

et al. 2019

Journal of Cell Biology

Self Cite

View full text Add to dashboard Cite

show abstract

“…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). It was possible that compensated TAP targeting by these pathways would make it difficult to detect a targeting defect in asna-1 single mutants.…”

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

View full text Add to dashboard Cite

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.

show abstract

“…Altogether, TA proteins represent 3–5% of the eukaryotic membrane proteome (Hegde and Keenan, ). It is well known that the targeting signal of TA proteins is embedded in their C‐terminal domain; however, in spite of a progress in understanding of the mechanisms involved in the sorting of TA proteins into ER (Mateja et al ., ; Cho and Shan, ), the targeting into the OMM, is still poorly understood.…”

Section: Introductionmentioning

confidence: 99%

Targeting of tail‐anchored proteins to Trichomonas vaginalis hydrogenosomes

Rada

Makki

Žárský

et al. 2019

Molecular Microbiology

View full text Add to dashboard Cite

Summary Tail‐anchored (TA) proteins are membrane proteins that are found in all domains of life. They consist of an N‐terminal domain that performs various functions and a single transmembrane domain (TMD) near the C‐terminus. In eukaryotes, TA proteins are targeted to the membranes of mitochondria, the endoplasmic reticulum (ER), peroxisomes and in plants, chloroplasts. The targeting of these proteins to their specific destinations correlates with the properties of the C‐terminal domain, mainly the TMD hydrophobicity and the net charge of the flanking regions. Trichomonas vaginalis is a human parasite that has adapted to oxygen‐poor environment. This adaptation is reflected by the presence of highly modified mitochondria (hydrogenosomes) and the absence of peroxisomes. The proteome of hydrogenosomes is considerably reduced; however, our bioinformatic analysis predicted 120 putative hydrogenosomal TA proteins. Seven proteins were selected to prove their localization. The elimination of the net positive charge in the C‐tail of the hydrogenosomal TA4 protein resulted in its dual localization to hydrogenosomes and the ER, causing changes in ER morphology. Domain mutation and swap experiments with hydrogenosomal (TA4) and ER (TAPDI) proteins indicated that the general principles for specific targeting are conserved across eukaryotic lineages, including T. vaginalis; however, there are also significant lineage‐specific differences.

show abstract

Substrate relay in an Hsp70‐cochaperone cascade safeguards tail‐anchored membrane protein targeting

Cited by 47 publications

References 61 publications

A molecular recognition feature mediates ribosome-induced SRP-receptor assembly during protein targeting

A molecular recognition feature mediates ribosome-induced SRP-receptor assembly during protein targeting

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

Targeting of tail‐anchored proteins to Trichomonas vaginalis hydrogenosomes

Contact Info

Product

Resources

About