Startseite In vitro import experiments with semi-intact cells suggest a role of the Sec61 paralog Ssh1 in mitochondrial biogenesis
Artikel
Lizenziert
Nicht lizenziert Erfordert eine Authentifizierung

In vitro import experiments with semi-intact cells suggest a role of the Sec61 paralog Ssh1 in mitochondrial biogenesis

  • Janina Laborenz , Katja Hansen , Cristina Prescianotto-Baschong , Anne Spang und Johannes M. Herrmann EMAIL logo
Veröffentlicht/Copyright: 12. Juni 2019

Abstract

Mitochondrial biogenesis relies on the synthesis of hundreds of different precursor proteins in the cytosol and their subsequent import into the organelle. Recent studies suggest that the surface of the endoplasmic reticulum (ER) actively contributes to the targeting of some mitochondrial precursors. In the past, in vitro import experiments with isolated mitochondria proved to be extremely powerful to elucidate the individual reactions of the mitochondrial import machinery. However, this in vitro approach is not well suited to study the influence of non-mitochondrial membranes. In this study, we describe an in vitro system using semi-intact yeast cells to test a potential import relevance of the ER proteins Erg3, Lcb5 and Ssh1, all being required for efficient mitochondrial respiration. We optimized the conditions of this experimental test system and found that cells lacking Ssh1, a paralog of the Sec61 translocation pore, show a reduced import efficiency of mitochondrial precursor proteins. Our results suggest that Ssh1, directly or indirectly, increases the efficiency of the biogenesis of mitochondrial proteins. Our findings are compatible with a functional interdependence of the mitochondrial and the ER protein translocation systems.

Award Identifier / Grant number: DIP MitoBalance

Award Identifier / Grant number: IRTG1830

Award Identifier / Grant number: HE2803/8-2

Funding statement: We thank Sabine Knaus for technical assistance and Sandra Backes for comments on the manuscript. This study was funded by grants of the Deutsche Forschungsgemeinschaft (Funder Id: http://dx.doi.org/10.13039/501100001659, DIP MitoBalance, Funder Id: http://dx.doi.org/10.13039/501100001659, IRTG1830, Funder Id: http://dx.doi.org/10.13039/501100001659, HE2803/8-2), the Forschungsinitiative Rheinland Pfalz and the University of Basel.

References

Backes, S. and Herrmann, J.M. (2017). Protein translocation into the intermembrane space and matrix of mitochondria: mechanisms and driving forces. Front. Mol. Biosci. 4, 83.10.3389/fmolb.2017.00083Suche in Google Scholar PubMed PubMed Central

Backes, S., Hess, S., Boos, F., Woellhaf, M.W., Godel, S., Jung, M., Muhlhaus, T., and Herrmann, J.M. (2018). Tom70 enhances mitochondrial preprotein import efficiency by binding to internal targeting sequences. J. Cell Biol. 217, 1369–1382.10.1083/jcb.201708044Suche in Google Scholar PubMed PubMed Central

Becker, T., Bhushan, S., Jarasch, A., Armache, J.P., Funes, S., Jossinet, F., Gumbart, J., Mielke, T., Berninghausen, O., Schulten, K., et al. (2009). Structure of monomeric yeast and mammalian Sec61 complexes interacting with the translating ribosome. Science 326, 1369–1373.10.1126/science.1178535Suche in Google Scholar PubMed PubMed Central

Beckers, C.J., Keller, D.S., and Balch, W.E. (1987). Semi-intact cells permeable to macromolecules: use in reconstitution of protein transport from the endoplasmic reticulum to the Golgi complex. Cell 50, 523–534.10.1016/0092-8674(87)90025-0Suche in Google Scholar PubMed

Beckers, C., Block, M., Glick, B., Rothman, J., and Balch, W. (1989). Vesicular transport between the ER and the Golgi stack requires the NEM-sensitive fusion protein. Nature 339, 397–398.10.1038/339397a0Suche in Google Scholar PubMed

Bonnefoy, N., Chalvet, F., Hamel, P., Slominski, P.P., and Dujardin, G. (1994). OXA1, a Saccharomyces cerevisiae nuclear gene whose sequence is conserved from prokaryotes to eukaryotes controls cytochrome oxidase biogenesis. J. Mol. Biol. 239, 201–212.10.1006/jmbi.1994.1363Suche in Google Scholar PubMed

Boos, F., Kramer, L., Groh, C., Jung, F., Haberkant, P., Stein, F., Wollweber, F., Gackstatter, A., Zoller, E., van der Laan, M., et al. (2019). Mitochondrial protein-induced stress triggers a global adaptive transcriptional programme. Nat. Cell Biol. 21, 442–451.10.1038/s41556-019-0294-5Suche in Google Scholar PubMed

Calvo, S.E., Julien, O., Clauser, K.R., Shen, H., Kamer, K.J., Wells, J.A., and Mootha, V.K. (2017). Comparative analysis of mitochondrial N-termini from mouse, human, and yeast. Mol. Cell. Proteomics 16, 512–523.10.1074/mcp.M116.063818Suche in Google Scholar PubMed PubMed Central

Chacinska, A., Lind, M., Frazier, A.E., Dudek, J., Meisinger, C., Geissler, A., Sickmann, A., Meyer, H.E., Truscott, K.N., Guiard, B., et al. (2005). Mitochondrial presequence translocase: switching between TOM tethering and motor recruitment involves Tim21 and Tim17. Cell 120, 817–829.10.1016/j.cell.2005.01.011Suche in Google Scholar PubMed

Cheng, Z., Jiang, Y., Mandon, E.C., and Gilmore, R. (2005). Identification of cytoplasmic residues of Sec61p involved in ribosome binding and cotranslational translocation. J. Cell Biol. 168, 67–77.10.1083/jcb.200408188Suche in Google Scholar PubMed PubMed Central

Costa, E.A., Subramanian, K., Nunnari, J., and Weissman, J.S. (2018). Defining the physiological role of SRP in protein-targeting efficiency and specificity. Science 359, 689–692.10.1126/science.aar3607Suche in Google Scholar PubMed PubMed Central

Daum, G., Gasser, S., and Schatz, G. (1982). Import of proteins into mitochondria: energy-dependent, two-step processing of the intermembrane space enzyme cytochrome b2 by isolated yeast mitochondria. J. Biol. Chem. 257, 13075–13080.10.1016/S0021-9258(18)33624-XSuche in Google Scholar

Finke, K., Plath, K., Panzner, S., Prehn, S., Rapoport, T.A., Hartmann, E., and Sommer, T. (1996). A second trimeric complex containing homologs of the Sec61p complex functions in protein transport across the ER membrane of S. cerevisiae. EMBO J. 15, 1482–1494.10.1002/j.1460-2075.1996.tb00492.xSuche in Google Scholar

Fünfschilling, U. and Rospert, S. (1999). Nascent polypeptide-associated complex stimulates protein import into yeast mitochondria. Mol. Biol. Cell. 10, 3289–3299.10.1091/mbc.10.10.3289Suche in Google Scholar PubMed PubMed Central

Gamerdinger, M., Hanebuth, M.A., Frickey, T., and Deuerling, E. (2015). The principle of antagonism ensures protein targeting specificity at the endoplasmic reticulum. Science 348, 201–207.10.1126/science.aaa5335Suche in Google Scholar PubMed

Gasser, S.M., Daum, G., and Schatz, G. (1982). Import of proteins into mitochondria: energy-dependent uptake of precursors into isolated mitochondria. J. Biol. Chem. 257, 13034–13041.10.1016/S0021-9258(18)33618-4Suche in Google Scholar

Hansen, K.G., Aviram, N., Laborenz, J., Bibi, C., Meyer, M., Spang, A., Schuldiner, M., and Herrmann, J.M. (2018). An ER surface retrieval pathway safeguards the import of mitochondrial membrane proteins in yeast. Science 361, 1118–1122.10.1126/science.aar8174Suche in Google Scholar PubMed

Harty, C. and Romisch, K. (2013). Analysis of Sec61p and Ssh1p interactions in the ER membrane using the split-ubiquitin system. BMC Cell Biol. 14, 14.10.1186/1471-2121-14-14Suche in Google Scholar PubMed PubMed Central

Hoseini, H., Pandey, S., Jores, T., Schmitt, A., Franz-Wachtel, M., Macek, B., Buchner, J., Dimmer, K.S., and Rapaport, D. (2016). The cytosolic cochaperone Sti1 is relevant for mitochondrial biogenesis and morphology. FEBS J. 283, 3338–3352.10.1111/febs.13813Suche in Google Scholar PubMed

Itakura, E., Zavodszky, E., Shao, S., Wohlever, M.L., Keenan, R.J., and Hegde, R.S. (2016). Ubiquilins chaperone and triage mitochondrial membrane proteins for degradation. Mol. Cell 63, 21–33.10.1016/j.molcel.2016.05.020Suche in Google Scholar PubMed PubMed Central

Jan, C.H., Williams, C.C., and Weissman, J.S. (2014). Principles of ER cotranslational translocation revealed by proximity-specific ribosome profiling. Science 346, 1257521.10.1126/science.1257521Suche in Google Scholar PubMed PubMed Central

Jiang, Y., Cheng, Z., Mandon, E.C., and Gilmore, R. (2008). An interaction between the SRP receptor and the translocon is critical during cotranslational protein translocation. J. Cell Biol. 180, 1149–1161.10.1083/jcb.200707196Suche in Google Scholar PubMed PubMed Central

Jores, T., Lawatscheck, J., Beke, V., Franz-Wachtel, M., Yunoki, K., Fitzgerald, J.C., Macek, B., Endo, T., Kalbacher, H., Buchner, J., et al. (2018). Cytosolic Hsp70 and Hsp40 chaperones enable the biogenesis of mitochondrial β-barrel proteins. J Cell Biol. 217, 3091–3108.10.1083/jcb.201712029Suche in Google Scholar PubMed PubMed Central

Kowalski, L., Bragoszewski, P., Khmelinskii, A., Glow, E., Knop, M., and Chacinska, A. (2018). Determinants of the cytosolic turnover of mitochondrial intermembrane space proteins. BMC Biol. 16, 66.10.1186/s12915-018-0536-1Suche in Google Scholar PubMed PubMed Central

Merz, S. and Westermann, B. (2009). Genome-wide deletion mutant analysis reveals genes required for respiratory growth, mitochondrial genome maintenance and mitochondrial protein synthesis in Saccharomyces cerevisiae. Genome Biol. 10, R95.10.1186/gb-2009-10-9-r95Suche in Google Scholar PubMed PubMed Central

Mootha, V.K., Bunkenborg, J., Olsen, J.V., Hjerrild, M., Wisniewski, J.R., Stahl, E., Bolouri, M.S., Ray, H.N., Sihag, S., Kamal, M., et al. (2003). Integrated analysis of protein composition, tissue diversity, and gene regulation in mouse mitochondria. Cell 115, 629–640.10.1016/S0092-8674(03)00926-7Suche in Google Scholar PubMed

Morgenstern, M., Stiller, S.B., Lubbert, P., Peikert, C.D., Dannenmaier, S., Drepper, F., Weill, U., Hoss, P., Feuerstein, R., Gebert, M., et al. (2017). Definition of a high-confidence mitochondrial proteome at quantitative scale. Cell Rep. 19, 2836–2852.10.1016/j.celrep.2017.06.014Suche in Google Scholar PubMed PubMed Central

Okamoto, K., Brinker, A., Paschen, S.A., Moarefi, I., Hayer-Hartl, M., Neupert, W., and Brunner, M. (2002). The protein import motor of mitochondria: a targeted molecular ratchet driving unfolding and translocation. EMBO J. 21, 3659–3671.10.1093/emboj/cdf358Suche in Google Scholar PubMed PubMed Central

Opalinski, L., Song, J., Priesnitz, C., Wenz, L.S., Oeljeklaus, S., Warscheid, B., Pfanner, N., and Becker, T. (2018). Recruitment of cytosolic J-proteins by TOM receptors promotes mitochondrial protein biogenesis. Cell Rep. 25, 2036–2043.e2035.10.1016/j.celrep.2018.10.083Suche in Google Scholar PubMed PubMed Central

Papic, D., Elbaz-Alon, Y., Koerdt, S.N., Leopold, K., Worm, D., Jung, M., Schuldiner, M., and Rapaport, D. (2013). The role of Djp1 in import of the mitochondrial protein Mim1 demonstrates specificity between a cochaperone and its substrate protein. Mol. Cell Biol. 33, 4083–4094.10.1128/MCB.00227-13Suche in Google Scholar PubMed PubMed Central

Peleh, V., Ramesh, A., and Herrmann, J.M. (2015). Import of proteins into isolated yeast mitochondria. Methods Mol. Biol. 1270, 37–50.10.1007/978-1-4939-2309-0_3Suche in Google Scholar PubMed

Ponce-Rojas, J.C., Avendano-Monsalve, M.C., Yanez-Falcon, A.R., Jaimes-Miranda, F., Garay, E., Torres-Quiroz, F., DeLuna, A., and Funes, S. (2017). αβ′-NAC cooperates with Sam37 to mediate early stages of mitochondrial protein import. FEBS J. 284, 814–830.10.1111/febs.14024Suche in Google Scholar PubMed

Prescianotto-Baschong, C. and Riezman, H. (2002). Ordering of compartments in the yeast endocytic pathway. Traffic 3, 37–49.10.1034/j.1600-0854.2002.30106.xSuche in Google Scholar PubMed

Ramesh, A., Peleh, V., Martinez-Caballero, S., Wollweber, F., Sommer, F., van der Laan, M., Schroda, M., Alexander, R.T., Campo, M.L., and Herrmann, J.M. (2016). A disulfide bond in the TIM23 complex is crucial for voltage gating and mitochondrial protein import. J. Cell Biol. 214, 417–431.10.1083/jcb.201602074Suche in Google Scholar PubMed PubMed Central

Rhee, H.W., Zou, P., Udeshi, N.D., Martell, J.D., Mootha, V.K., Carr, S.A., and Ting, A.Y. (2013). Proteomic mapping of mitochondria in living cells via spatially restricted enzymatic tagging. Science 339, 1328–1331.10.1126/science.1230593Suche in Google Scholar PubMed PubMed Central

Schlenstedt, G., Hurt, E., Doye, V., and Silver, P.A. (1993). Reconstitution of nuclear protein transport with semi-intact yeast cells. J. Cell Biol. 123, 785–798.10.1083/jcb.123.4.785Suche in Google Scholar PubMed PubMed Central

Shieh, H.L. and Chiang, H.L. (1998). In vitro reconstitution of glucose-induced targeting of fructose-1,6-bisphosphatase into the vacuole in semi-intact yeast cells. J. Biol. Chem. 273, 3381–3387.10.1074/jbc.273.6.3381Suche in Google Scholar PubMed

Shiota, T., Imai, K., Qiu, J., Hewitt, V.L., Tan, K., Shen, H.H., Sakiyama, N., Fukasawa, Y., Hayat, S., Kamiya, M., et al. (2015). Molecular architecture of the active mitochondrial protein gate. Science 349, 1544–1548.10.1126/science.aac6428Suche in Google Scholar PubMed

Spiller, M.P. and Stirling, C.J. (2011). Preferential targeting of a signal recognition particle-dependent precursor to the Ssh1p translocon in yeast. J. Biol. Chem. 286, 21953–21960.10.1074/jbc.M111.219568Suche in Google Scholar PubMed PubMed Central

Topf, U., Suppanz, I., Samluk, L., Wrobel, L., Boser, A., Sakowska, P., Knapp, B., Pietrzyk, M.K., Chacinska, A., and Warscheid, B. (2018). Quantitative proteomics identifies redox switches for global translation modulation by mitochondrially produced reactive oxygen species. Nat. Commun. 9, 324.10.1038/s41467-017-02694-8Suche in Google Scholar PubMed PubMed Central

Verleur, N., Hettema, E.H., van Roermund, C.W., Tabak, H.F., and Wanders, R.J. (1997). Transport of activated fatty acids by the peroxisomal ATP-binding-cassette transporter Pxa2 in a semi-intact yeast cell system. Eur. J. Biochem. 249, 657–661.10.1111/j.1432-1033.1997.00657.xSuche in Google Scholar PubMed

Vögtle, F.N., Wortelkamp, S., Zahedi, R.P., Becker, D., Leidhold, C., Gevaert, K., Kellermann, J., Voos, W., Sickmann, A., Pfanner, N., et al. (2009). Global analysis of the mitochondrial N-proteome identifies a processing peptidase critical for protein stability. Cell 139, 428–439.10.1016/j.cell.2009.07.045Suche in Google Scholar PubMed

Weidberg, H. and Amon, A. (2018). MitoCPR-A surveillance pathway that protects mitochondria in response to protein import stress. Science 360, pii: eaan4146.10.1126/science.aan4146Suche in Google Scholar PubMed PubMed Central

Wiedemann, N. and Pfanner, N. (2017). Mitochondrial machineries for protein import and assembly. Annu. Rev. Biochem. 86, 685–714.10.1146/annurev-biochem-060815-014352Suche in Google Scholar PubMed

Wilkinson, B.M., Tyson, J.R., and Stirling, C.J. (2001). Ssh1p determines the translocation and dislocation capacities of the yeast endoplasmic reticulum. Dev. Cell 1, 401–409.10.1016/S1534-5807(01)00043-0Suche in Google Scholar PubMed

Wrobel, L., Topf, U., Bragoszewski, P., Wiese, S., Sztolsztener, M.E., Oeljeklaus, S., Varabyova, A., Lirski, M., Chroscicki, P., Mroczek, S., et al. (2015). Mistargeted mitochondrial proteins activate a proteostatic response in the cytosol. Nature 524, 485–488.10.1038/nature14951Suche in Google Scholar PubMed

Yamamoto, H., Itoh, N., Kawano, S., Yatsukawa, Y., Momose, T., Makio, T., Matsunaga, M., Yokota, M., Esaki, M., Shodai, T., et al. (2011). Dual role of the receptor Tom20 in specificity and efficiency of protein import into mitochondria. Proc. Natl. Acad. Sci. USA 108, 91–96.10.1073/pnas.1014918108Suche in Google Scholar PubMed PubMed Central

Young, J.C., Hoogenraad, N.J., and Hartl, F.U. (2003). Molecular chaperones Hsp90 and Hsp70 deliver preproteins to the mitochondrial import receptor Tom70. Cell 112, 41–50.10.1016/S0092-8674(02)01250-3Suche in Google Scholar PubMed

Zimmermann, R. and Neupert, W. (1980). Transport of proteins to mitochondria: posttranslational transfer of ADP/ATP carrier into mitochondria. Eur. J. Biochem. 109, 217–229.10.1111/j.1432-1033.1980.tb04787.xSuche in Google Scholar PubMed

Received: 2019-03-21
Accepted: 2019-05-03
Published Online: 2019-06-12
Published in Print: 2019-08-27

©2019 Walter de Gruyter GmbH, Berlin/Boston

Heruntergeladen am 23.9.2025 von https://www.degruyterbrill.com/document/doi/10.1515/hsz-2019-0196/html
Button zum nach oben scrollen