Translationally controlled tumor protein (TCTP) is a multifunctional and highly conserved protein from yeast to humans. Recently, its role in non-selective autophagy has been reported with controversial results in mammalian and human cells. Herein we examine the effect of Mmi1, the yeast ortholog of TCTP, on non-selective autophagy in budding yeast Saccharomyces cerevisiae, a well-established model system to monitor autophagy. We induced autophagy by nitrogen starvation or rapamycin addition and measured autophagy by using the Pho8Δ60 and GFP-Atg8 processing assays in WT, mmi1Δ, and in autophagy-deficient strains atg8Δ or atg1Δ. Our results demonstrate that Mmi1 does not affect basal or nitrogen starvation-induced autophagy. However, an increased rapamycin-induced autophagy is detected in mmi1Δ strain when the cells enter the post-diauxic growth phase, and this phenotype can be rescued by inserted wild-type MMI1 gene. Further, the mmi1Δ cells exhibit significantly lower amounts of reactive oxygen species (ROS) in the post-diauxic growth phase compared to WT cells. In summary, our study suggests that Mmi1 negatively affects rapamycin-induced autophagy in the post-diauxic growth phase and supports the role of Mmi1/TCTP as a negative autophagy regulator in eukaryotic cells.

Institute of Microbiology of the Czech Academy of Sciences Videnska 1083 Prague 4 142 20 Czech Republic

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Chen S.H., Wu P.-S., Chou C.-H., Yan Y.-T., Liu H., Weng S.-Y., Yang-Yen H.-F. A Knockout Mouse Approach Reveals that TCTP Functions as an Essential Factor for Cell Proliferation and Survival in a Tissue- or Cell Type–specific Manner. Mol. Boil. Cell. 2007;18:2525–2532. doi: 10.1091/mbc.e07-02-0188. PubMed DOI PMC

Hsu Y.-C., Chern J.J., Cai Y., Liu M., Choi K.-W. Drosophila TCTP is essential for growth and proliferation through regulation of dRheb GTPase. Nature. 2007;445:785–788. doi: 10.1038/nature05528. PubMed DOI

Liu Z.-L., Xu J., Ling L., Zhang R., Shang P., Huang Y.-P. CRISPR disruption of TCTP gene impaired normal development in the silkworm Bombyx mori. Insect Sci. 2018;26:973–982. doi: 10.1111/1744-7917.12567. PubMed DOI PMC

Susini L., Besse S., Duflaut D., Lespagnol A., Beekman C., Fiucci G., Atkinson A.R., Busso D., Poussin P., Marine J.-C., et al. TCTP protects from apoptotic cell death by antagonizing bax function. Cell Death Differ. 2008;15:1211–1220. doi: 10.1038/cdd.2008.18. PubMed DOI

Rho S.B., Lee J.H., Park M.S., Byun H.J., Kang S., Seo S.S., Kim J.Y., Park S.Y. Anti-Apoptotic Protein Tctp Controls the Stability of the Tumor Suppressor P53. FEBS Lett. 2011;585:29–35. doi: 10.1016/j.febslet.2010.11.014. PubMed DOI

Li F., Zhang D., Fujise K. Characterization of Fortilin, a Novel Antiapoptotic Protein. J. Boil. Chem. 2001;276:47542–47549. doi: 10.1074/jbc.M108954200. PubMed DOI

Zhang D., Li F., Weidner D., Mnjoyan Z.H., Fujise K. Physical and Functional Interaction between Myeloid Cell Leukemia 1 Protein (MCL1) and Fortilin. J. Boil. Chem. 2002;277:37430–37438. doi: 10.1074/jbc.M207413200. PubMed DOI

Bommer U.-A. The Translational Controlled Tumour Protein TCTP: Biological Functions and Regulation. Results Probl. Cell Differ. 2017;64:69–126. PubMed

Thomas G., Luther H. Transcriptional and translational control of cytoplasmic proteins after serum stimulation of quiescent Swiss 3T3 cells. Proc. Natl. Acad. Sci. USA. 1981;78:5712–5716. doi: 10.1073/pnas.78.9.5712. PubMed DOI PMC

Chen K., Huang C., Yuan J., Cheng H., Zhou R. Long-Term Artificial Selection Reveals a Role of TCTP in Autophagy in Mammalian Cells. Mol. Boil. Evol. 2014;31:2194–2211. doi: 10.1093/molbev/msu181. PubMed DOI

Bae S.-Y., Byun S., Bae S.H., Min S., Woo H.A., Lee K. TPT1 (tumor protein, translationally-controlled 1) negatively regulates autophagy through the BECN1 interactome and an MTORC1-mediated pathway. Autophagy. 2017;13:820–833. doi: 10.1080/15548627.2017.1287650. PubMed DOI PMC

Tsukada M., Ohsumi Y. Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae. FEBS Lett. 1993;333:169–174. doi: 10.1016/0014-5793(93)80398-E. PubMed DOI

Torggler R., Papinski D., Kraft C. Assays to Monitor Autophagy in Saccharomyces cerevisiae. Cells. 2017;6:23. doi: 10.3390/cells6030023. PubMed DOI PMC

Nakatogawa H., Suzuki K., Kamada Y., Ohsumi Y. Dynamics and diversity in autophagy mechanisms: Lessons from yeast. Nat. Rev. Mol. Cell Boil. 2009;10:458–467. doi: 10.1038/nrm2708. PubMed DOI

Farré J.-C., Subramani S. Mechanistic insights into selective autophagy pathways: Lessons from yeast. Nat. Rev. Mol. Cell Boil. 2016;17:537–552. doi: 10.1038/nrm.2016.74. PubMed DOI PMC

Galdieri L., Mehrotra S., Yu S., Vancura A. Transcriptional Regulation in Yeast during Diauxic Shift and Stationary Phase. OMICS: A J. Integr. Boil. 2010;14:629–638. doi: 10.1089/omi.2010.0069. PubMed DOI PMC

Swinnen E., Wanke V., Roosen J., Smets B., Dubouloz F., Pedruzzi I., Cameroni E., De Virgilio C., Winderickx J. Rim15 and the crossroads of nutrient signalling pathways in Saccharomyces cerevisiae. Cell Div. 2006;1:3. doi: 10.1186/1747-1028-1-3. PubMed DOI PMC

Fleischer T.C., Weaver C.M., McAfee K.J., Jennings J.L., Link A.J. Systematic identification and functional screens of uncharacterized proteins associated with eukaryotic ribosomal complexes. Genome Res. 2006;20:1294–1307. doi: 10.1101/gad.1422006. PubMed DOI PMC

Rinnerthaler M., Jarolim S., Heeren G., Palle E., Perju S., Klinger H., Bogengruber E., Madeo F., Braun R.J., Breitenbach-Koller L., et al. Mmi1 (Ykl056c, Tma19), the Yeast Orthologue of the Translationally Controlled Tumor Protein (Tctp) Has Apoptotic Functions and Interacts with Both Microtubules and Mitochondria. Biochim. Et Biophys. Acta. 2006;1757:631–638. doi: 10.1016/j.bbabio.2006.05.022. PubMed DOI

A Kulak N., Pichler G., Paron I., Nagaraj N., Mann M. Minimal, encapsulated proteomic-sample processing applied to copy-number estimation in eukaryotic cells. Nat. Methods. 2014;11:319–324. doi: 10.1038/nmeth.2834. PubMed DOI

Murphy J.P., Stepanova E., Everley R.A., Paulo J.A., Gygi S.P. Comprehensive Temporal Protein Dynamics during the Diauxic Shift in Saccharomyces cerevisiae. Mol. Cell. Proteom. 2015;14:2454–2465. doi: 10.1074/mcp.M114.045849. PubMed DOI PMC

Delaney J.R., Murakami C.J., Olsen B., Kennedy B.K., Kaeberlein M. Quantitative evidence for early life fitness defects from 32 longevity-associated alleles in yeast. Cell Cycle. 2011;10:156–165. doi: 10.4161/cc.10.1.14457. PubMed DOI PMC

Chong Y.T., Koh J.L., Friesen H., Duffy S.K., Cox M.J., Moses A., Moffat J., Boone C., Andrews B.J. Yeast Proteome Dynamics from Single Cell Imaging and Automated Analysis. Cell. 2015;161:1413–1424. doi: 10.1016/j.cell.2015.04.051. PubMed DOI

Bommer U.A., Iadevaia V., Chen J., Knoch B., Engel M., Proud C.G. Growth-Factor Dependent Expression of the Translationally Controlled Tumour Protein Tctp Is Regulated through the Pi3-K/Akt/Mtorc1 Signalling Pathway. Cell. Signal. 2015;27:1557–1568. doi: 10.1016/j.cellsig.2015.04.011. PubMed DOI

Bischof J., Salzmann M., Streubel M.K., Hasek J., Geltinger F., Duschl J., Bresgen N., Briza P., Haskova D., Lejskova R., et al. Clearing the outer mitochondrial membrane from harmful proteins via lipid droplets. Cell Death Discov. 2017;3:17016. doi: 10.1038/cddiscovery.2017.16. PubMed DOI PMC

Rinnerthaler M., Lejskova R., Groušl T., Stradalova V., Heeren G., Richter K., Breitenbach-Koller L., Malínský J., Hasek J., Breitenbach M. Mmi1, the Yeast Homologue of Mammalian TCTP, Associates with Stress Granules in Heat-Shocked Cells and Modulates Proteasome Activity. PLoS ONE. 2013;8:e77791. doi: 10.1371/journal.pone.0077791. PubMed DOI PMC

Stark C., Breitkreutz B.-J., Reguly T., Boucher L., Breitkreutz A., Tyers M. BioGRID: A general repository for interaction datasets. Nucleic Acids Res. 2006;34:D535–D539. doi: 10.1093/nar/gkj109. PubMed DOI PMC

Cheong H., Klionsky D.J. Chapter 1 Biochemical Methods to Monitor Autophagy-Related Processes in Yeast. Methods Enzymol. 2008;451:1–26. PubMed

Wang X., Li S., Liu Y., Ma C. Redox Regulated Peroxisome Homeostasis. Redox Biol. 2015:104–108. doi: 10.1016/j.redox.2014.12.006. PubMed DOI PMC

Yorimitsu T., Klionsky D.J. Autophagy: Molecular machinery for self-eating. Cell Death Differ. 2005;12:1542–1552. doi: 10.1038/sj.cdd.4401765. PubMed DOI PMC

Takeshige K., Baba M., Tsuboi S., Noda T., Ohsumi Y. Autophagy in yeast demonstrated with proteinase-deficient mutants and conditions for its induction. J. Cell Boil. 1992;119:301–311. doi: 10.1083/jcb.119.2.301. PubMed DOI PMC

Noda T. Tor, a Phosphatidylinositol Kinase Homologue, Controls Autophagy in Yeast. J. Boil. Chem. 1998;273:3963–3966. doi: 10.1074/jbc.273.7.3963. PubMed DOI

Barbet N.C., Schneider U., Helliwell S.B., Stansfield I., Tuite M.F., Hall M.N. TOR controls translation initiation and early G1 progression in yeast. Mol. Boil. Cell. 1996;7:25–42. doi: 10.1091/mbc.7.1.25. PubMed DOI PMC

Thomas G., Hall M.N. TOR signalling and control of cell growth. Curr. Opin. Cell Boil. 1997;9:782–787. doi: 10.1016/S0955-0674(97)80078-6. PubMed DOI

Parzych K.R., Klionsky D.J. An Overview of Autophagy: Morphology, Mechanism, and Regulation. Antioxid. Redox Signal. 2014;20:460–473. doi: 10.1089/ars.2013.5371. PubMed DOI PMC

Ohsumi Y. Historical Landmarks of Autophagy Research. Cell Res. 2014;24:9–23. doi: 10.1038/cr.2013.169. PubMed DOI PMC

Kawamata T., Kamada Y., Suzuki K., Kuboshima N., Akimatsu H., Ota S., Ohsumi M., Ohsumi Y. Characterization of a novel autophagy-specific gene, ATG29. Biochem. Biophys. Res. Commun. 2005;338:1884–1889. doi: 10.1016/j.bbrc.2005.10.163. PubMed DOI

Kiel J.A.K.W. Autophagy in unicellular eukaryotes. Philos. Trans. R. Soc. B: Boil. Sci. 2010;365:819–830. doi: 10.1098/rstb.2009.0237. PubMed DOI PMC

Liu E.Y., Ryan K.M. Autophagy and cancer-issues we need to digest. J. Cell Sci. 2012;125:2349–2358. doi: 10.1242/jcs.093708. PubMed DOI

Brachmann C.B., Davies A., Cost G.J., Caputo E., Li J., Hieter P., Boeke J.D. Designer deletion strains derived fromSaccharomyces cerevisiae S288C: A useful set of strains and plasmids for PCR-mediated gene disruption and other applications. Yeast. 1998;14:115–132. doi: 10.1002/(SICI)1097-0061(19980130)14:2<115::AID-YEA204>3.0.CO;2-2. PubMed DOI

Noda T., Matsuura A., Wada Y., Ohsumi Y. Novel System for Monitoring Autophagy in the Yeast Saccharomyces cerevisiae. Biochem. Biophys. Res. Commun. 1995;210:126–132. doi: 10.1006/bbrc.1995.1636. PubMed DOI

Bicknell A.A., Tourtellotte J., Niwa M. Late Phase of the Endoplasmic Reticulum Stress Response Pathway Is Regulated by Hog1 MAP Kinase*. J. Boil. Chem. 2010;285:17545–17555. doi: 10.1074/jbc.M109.084681. PubMed DOI PMC

Noda T., Klionsky D.J. The Quantitative Pho8delta60 Assay of Nonspecific Autophagy. Methods Enzymol. 2018;451:33–42. PubMed

Kramer M.H., Farré J.-C., Mitra K., Yu M.K., Ono K., Demchak B., Licon K., Flagg M., Balakrishnan R., Cherry J.M., et al. Active Interaction Mapping Reveals the Hierarchical Organization of Autophagy. Mol. Cell. 2017;65:761–774. doi: 10.1016/j.molcel.2016.12.024. PubMed DOI PMC

Suzuki K., Kirisako T., Kamada Y., Mizushima N., Noda T., Ohsumi Y. The pre-autophagosomal structure organized by concerted functions of APG genes is essential for autophagosome formation. EMBO J. 2001;20:5971–5981. doi: 10.1093/emboj/20.21.5971. PubMed DOI PMC

Rueden C.T., Schindelin J., Hiner M.C., DeZonia B.E., Walter A.E., Arena E.T., Eliceiri K.W. Imagej2: Imagej for the Next Generation of Scientific Image Data. BMC Bioinform. 2017;18:529. doi: 10.1186/s12859-017-1934-z. PubMed DOI PMC

Neklesa T.K., Davis R.W. Superoxide anions regulate TORC1 and its ability to bind Fpr1:rapamycin complex. Proc. Natl. Acad. Sci. USA. 2008;105:15166–15171. doi: 10.1073/pnas.0807712105. PubMed DOI PMC

Robinson K.M., Janes M.S., Pehar M., Monette J.S., Ross M.F., Hagen T.M., Murphy M.P., Beckman J.S. Selective fluorescent imaging of superoxide in vivo using ethidium-based probes. Proc. Natl. Acad. Sci. USA. 2006;103:15038–15043. doi: 10.1073/pnas.0601945103. PubMed DOI PMC

Murakami C., Kaeberlein M. Quantifying Yeast Chronological Life Span by Outgrowth of Aged Cells. J. Vis. Exp. 2009 doi: 10.3791/1156. PubMed DOI PMC

Kirisako T., Ichimura Y., Okada H., Kabeya Y., Mizushima N., Yoshimori T., Ohsumi M., Takao T., Noda T., Ohsumi Y. The Reversible Modification Regulates the Membrane-Binding State of Apg8/Aut7 Essential for Autophagy and the Cytoplasm to Vacuole Targeting Pathway. J. Cell Boil. 2000;151:263–276. doi: 10.1083/jcb.151.2.263. PubMed DOI PMC

Matsuura A., Tsukada M., Wada Y., Ohsumi Y. Apg1p, a novel protein kinase required for the autophagic process in Saccharomyces cerevisiae. Gene. 1997;192:245–250. doi: 10.1016/S0378-1119(97)00084-X. PubMed DOI

An Z., Tassa A., Thomas C., Zhong R., Xiao G., Fotedar R., Tu B.P., Klionsky D.J., Levine B. Autophagy Is Required for G(1)/G(0) Quiescence in Response to Nitrogen Starvation in Saccharomyces Cerevisiae. Autophagy. 2014;10:1702–1711. doi: 10.4161/auto.32122. PubMed DOI PMC

Rallis C., Codlin S., Bahler J. TORC1 signaling inhibition by rapamycin and caffeine affect lifespan, global gene expression, and cell proliferation of fission yeast. Aging Cell. 2013;12:563–573. doi: 10.1111/acel.12080. PubMed DOI PMC

Evans S.K., Burgess K.E., Gray J.V. Recovery from Rapamycin: Drug-Insensitive Activity of Yeast Target of Rapamycin Complex 1 (Torc1) Supports Residual Proliferation That Dilutes Rapamycin among Progeny Cells. J. Biol. Chem. 2014;289:26554–26565. doi: 10.1074/jbc.M114.589754. PubMed DOI PMC

Dikicioglu D., Eke E.D., Eraslan S., Oliver S.G., Kirdar B. Saccharomyces cerevisiae adapted to grow in the presence of low-dose rapamycin exhibit altered amino acid metabolism. Cell Commun. Signal. 2018;16:85. doi: 10.1186/s12964-018-0298-y. PubMed DOI PMC

Morgan J.T., Fink G.R., Bartel D.P. Excised linear introns regulate growth in yeast. Nature. 2019;565:606–611. doi: 10.1038/s41586-018-0828-1. PubMed DOI PMC

Thoreen C.C., Kang S.A., Chang J.W., Liu Q., Zhang J., Gao Y., Reichling L.J., Sim T., Sabatini D.M., Gray N.S. An ATP-competitive Mammalian Target of Rapamycin Inhibitor Reveals Rapamycin-resistant Functions of mTORC1. J. Boil. Chem. 2009;284:8023–8032. doi: 10.1074/jbc.M900301200. PubMed DOI PMC

Feldman M.E., Apsel B., Uotila A., Loewith R., Knight Z.A., Ruggero D., Shokat K.M. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLoS Biol. 2009;7:e1000038. doi: 10.1371/journal.pbio.1000038. PubMed DOI PMC

Gray J.V., Petsko G.A., Johnston G.C., Ringe D., Singer R.A., Werner-Washburne M. “Sleeping Beauty”: Quiescence in Saccharomyces cerevisiae†. Microbiol. Mol. Boil. Rev. 2004;68:187–206. doi: 10.1128/MMBR.68.2.187-206.2004. PubMed DOI PMC

Krause S.A., Gray J.V. The protein kinase C pathway is required for viability in quiescence in Saccharomyces cerevisiae. Curr. Boil. 2002;12:588–593. doi: 10.1016/S0960-9822(02)00760-1. PubMed DOI

Adachi A., Koizumi M., Ohsumi Y. Autophagy induction under carbon starvation conditions is negatively regulated by carbon catabolite repression. J. Boil. Chem. 2017;292:19905–19918. doi: 10.1074/jbc.M117.817510. PubMed DOI PMC

Kayikci Ö., Nielsen J. Glucose repression in Saccharomyces cerevisiae. FEMS Yeast Res. 2015;15 PubMed PMC

Scherz-Shouval R., Shvets E., Fass E., Shorer H., Gil L., Elazar Z. Reactive oxygen species are essential for autophagy and specifically regulate the activity of Atg4. EMBO J. 2007;26:1749–1760. doi: 10.1038/sj.emboj.7601623. PubMed DOI PMC

Lee J., Giordano S., Zhang J. Autophagy, Mitochondria and Oxidative Stress: Cross-Talk and Redox Signalling. Biochem. J. 2011;441:523–540. doi: 10.1042/BJ20111451. PubMed DOI PMC

Pérez-Pérez M.E., Zaffagnini M., Marchand C.H., Crespo J.L., Lemaire S.D. The yeast autophagy protease Atg4 is regulated by thioredoxin. Autophagy. 2014;10:1953–1964. doi: 10.4161/auto.34396. PubMed DOI PMC

Jing H., Liu H., Zhang L., Gao J., Song H., Tan X. Ethanol induces autophagy regulated by mitochondrial ROS in Saccharomyces cerevisiae. J. Microbiol. Biotechnol. 2018;28:1982–1991. doi: 10.4014/jmb.1806.06014. PubMed DOI

Horie T., Kawamata T., Matsunami M., Ohsumi Y. Recycling of iron via autophagy is critical for the transition from glycolytic to respiratory growth. J. Boil. Chem. 2017;292:8533–8543. doi: 10.1074/jbc.M116.762963. PubMed DOI PMC

Iwama R., Ohsumi Y. Analysis of autophagy activated during changes in carbon source availability in yeast cells. J. Boil. Chem. 2019;294:5590–5603. doi: 10.1074/jbc.RA118.005698. PubMed DOI PMC

A fully automated morphological analysis of yeast mitochondria from wide-field fluorescence images