Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic>
ABSTRACT The 90-kDa heat shock protein (HSP90) of eukaryotes is a highly abundant and essential chaperone required for the maturation of regulatory and signal proteins. In the protozoan parasite Leishmania donovani, causative agent of the fatal visceral leishmaniasis, HSP90 activity is essential for...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2018-10-01
|
| Series: | mSystems |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mSystems.00214-18 |
| _version_ | 1818405121268121600 |
|---|---|
| author | Eugenia Bifeld Stephan Lorenzen Katharina Bartsch Juan-José Vasquez T. Nicolai Siegel Joachim Clos |
| author_facet | Eugenia Bifeld Stephan Lorenzen Katharina Bartsch Juan-José Vasquez T. Nicolai Siegel Joachim Clos |
| author_sort | Eugenia Bifeld |
| collection | DOAJ |
| description | ABSTRACT The 90-kDa heat shock protein (HSP90) of eukaryotes is a highly abundant and essential chaperone required for the maturation of regulatory and signal proteins. In the protozoan parasite Leishmania donovani, causative agent of the fatal visceral leishmaniasis, HSP90 activity is essential for cell proliferation and survival. Even more importantly, its inhibition causes life cycle progression from the insect stage to the pathogenic, mammalian stage. To unravel the molecular impact of HSP90 activity on the parasites’ gene expression, we performed a ribosome profiling analysis of L. donovani, comparing genome-wide protein synthesis patterns in the presence and absence of the HSP90-specific inhibitor radicicol and an ectopically expressed radicicol-resistant HSP90 variant. We find that ribosome-protected RNA faithfully maps open reading frames and represents 97% of the annotated protein-coding genes of L. donovani. Protein synthesis was found to correlate poorly with RNA steady-state levels, indicating a regulated translation as primary mechanism for HSP90-dependent gene expression. The results confirm inhibitory effects of HSP90 on the synthesis of Leishmania proteins that are associated with the pathogenic, intracellular stage of the parasite. Those include heat shock proteins, redox enzymes, virulence-enhancing surface proteins, proteolytic pathways, and a complete set of histones. Conversely, HSP90 promotes fatty acid synthesis enzymes. Complementing radicicol treatment with the radicicol-resistant HSP90rr variant revealed important off-target radicicol effects that control a large number of the above-listed proteins. Leishmania lacks gene-specific transcription regulation and relies on regulated translation instead. Our ribosome footprinting analysis demonstrates a controlling function of HSP90 in stage-specific protein synthesis but also significant, HSP90-independent effects of the inhibitor radicicol. IMPORTANCE Leishmania parasites cause severe illness in humans and animals. They exist in two developmental stages, insect form and mammalian form, which differ in shape and gene expression. By mapping and quantifying RNA fragments protected by protein synthesis complexes, we determined the rates of protein synthesis for >90% of all Leishmania proteins in response to the inhibition of a key regulatory protein, the 90-kDa heat shock protein. We find that Leishmania depends on a regulation of protein synthesis for controlling its gene expression and that heat shock protein 90 inhibition can trigger the developmental program from insect form to mammalian form of the pathogen. |
| first_indexed | 2024-12-14T08:51:00Z |
| format | Article |
| id | doaj.art-5dbeae5ae8294a028155e3a33ec51390 |
| institution | Directory Open Access Journal |
| issn | 2379-5077 |
| language | English |
| last_indexed | 2024-12-14T08:51:00Z |
| publishDate | 2018-10-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mSystems |
| spelling | doaj.art-5dbeae5ae8294a028155e3a33ec513902022-12-21T23:09:02ZengAmerican Society for MicrobiologymSystems2379-50772018-10-013610.1128/mSystems.00214-18Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic>Eugenia Bifeld0Stephan Lorenzen1Katharina Bartsch2Juan-José Vasquez3T. Nicolai Siegel4Joachim Clos5Leishmaniasis Group, Bernhard Nocht Institute for Tropical Medicine, Hamburg, GermanyBioinformatics Unit, Bernhard Nocht Institute for Tropical Medicine, Hamburg, GermanyLeishmaniasis Group, Bernhard Nocht Institute for Tropical Medicine, Hamburg, GermanyResearch Center for Infectious Diseases, University of Würzburg, Würzburg, GermanyResearch Center for Infectious Diseases, University of Würzburg, Würzburg, GermanyLeishmaniasis Group, Bernhard Nocht Institute for Tropical Medicine, Hamburg, GermanyABSTRACT The 90-kDa heat shock protein (HSP90) of eukaryotes is a highly abundant and essential chaperone required for the maturation of regulatory and signal proteins. In the protozoan parasite Leishmania donovani, causative agent of the fatal visceral leishmaniasis, HSP90 activity is essential for cell proliferation and survival. Even more importantly, its inhibition causes life cycle progression from the insect stage to the pathogenic, mammalian stage. To unravel the molecular impact of HSP90 activity on the parasites’ gene expression, we performed a ribosome profiling analysis of L. donovani, comparing genome-wide protein synthesis patterns in the presence and absence of the HSP90-specific inhibitor radicicol and an ectopically expressed radicicol-resistant HSP90 variant. We find that ribosome-protected RNA faithfully maps open reading frames and represents 97% of the annotated protein-coding genes of L. donovani. Protein synthesis was found to correlate poorly with RNA steady-state levels, indicating a regulated translation as primary mechanism for HSP90-dependent gene expression. The results confirm inhibitory effects of HSP90 on the synthesis of Leishmania proteins that are associated with the pathogenic, intracellular stage of the parasite. Those include heat shock proteins, redox enzymes, virulence-enhancing surface proteins, proteolytic pathways, and a complete set of histones. Conversely, HSP90 promotes fatty acid synthesis enzymes. Complementing radicicol treatment with the radicicol-resistant HSP90rr variant revealed important off-target radicicol effects that control a large number of the above-listed proteins. Leishmania lacks gene-specific transcription regulation and relies on regulated translation instead. Our ribosome footprinting analysis demonstrates a controlling function of HSP90 in stage-specific protein synthesis but also significant, HSP90-independent effects of the inhibitor radicicol. IMPORTANCE Leishmania parasites cause severe illness in humans and animals. They exist in two developmental stages, insect form and mammalian form, which differ in shape and gene expression. By mapping and quantifying RNA fragments protected by protein synthesis complexes, we determined the rates of protein synthesis for >90% of all Leishmania proteins in response to the inhibition of a key regulatory protein, the 90-kDa heat shock protein. We find that Leishmania depends on a regulation of protein synthesis for controlling its gene expression and that heat shock protein 90 inhibition can trigger the developmental program from insect form to mammalian form of the pathogen.https://journals.asm.org/doi/10.1128/mSystems.00214-18HSP90Leishmania donovaniradicicolribosome profiling |
| spellingShingle | Eugenia Bifeld Stephan Lorenzen Katharina Bartsch Juan-José Vasquez T. Nicolai Siegel Joachim Clos Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> mSystems HSP90 Leishmania donovani radicicol ribosome profiling |
| title | Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> |
| title_full | Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> |
| title_fullStr | Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> |
| title_full_unstemmed | Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> |
| title_short | Ribosome Profiling Reveals HSP90 Inhibitor Effects on Stage-Specific Protein Synthesis in <italic toggle="yes">Leishmania donovani</italic> |
| title_sort | ribosome profiling reveals hsp90 inhibitor effects on stage specific protein synthesis in italic toggle yes leishmania donovani italic |
| topic | HSP90 Leishmania donovani radicicol ribosome profiling |
| url | https://journals.asm.org/doi/10.1128/mSystems.00214-18 |
| work_keys_str_mv | AT eugeniabifeld ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic AT stephanlorenzen ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic AT katharinabartsch ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic AT juanjosevasquez ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic AT tnicolaisiegel ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic AT joachimclos ribosomeprofilingrevealshsp90inhibitoreffectsonstagespecificproteinsynthesisinitalictoggleyesleishmaniadonovaniitalic |