Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition.
BACKGROUND: The 14-3-3 proteins are structurally conserved throughout eukaryotes and participate in protein kinase signaling. All 14-3-3 proteins are known to bind to evolutionally conserved phosphoserine-containing motifs (modes 1 and/or 2) with high affinity. In Trypanosoma brucei, 14-3-3I and II...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2010-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3006207?pdf=render |
| _version_ | 1819037714335399936 |
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| author | Masahiro Inoue Kouichi Yasuda Haruki Uemura Natsumi Yasaka Hiroshi Inoue Yoshitatsu Sei Nobuo Horikoshi Toshihide Fukuma |
| author_facet | Masahiro Inoue Kouichi Yasuda Haruki Uemura Natsumi Yasaka Hiroshi Inoue Yoshitatsu Sei Nobuo Horikoshi Toshihide Fukuma |
| author_sort | Masahiro Inoue |
| collection | DOAJ |
| description | BACKGROUND: The 14-3-3 proteins are structurally conserved throughout eukaryotes and participate in protein kinase signaling. All 14-3-3 proteins are known to bind to evolutionally conserved phosphoserine-containing motifs (modes 1 and/or 2) with high affinity. In Trypanosoma brucei, 14-3-3I and II play pivotal roles in motility, cytokinesis and the cell cycle. However, none of the T. brucei 14-3-3 binding proteins have previously been documented. METHODOLOGY/PRINCIPAL FINDINGS: Initially we showed that T. brucei 14-3-3 proteins exhibit far lower affinity to those peptides containing RSxpSxP (mode 1) and RxY/FxpSxP (mode 2) (where x is any amino acid residue and pS is phosphoserine) than human 14-3-3 proteins, demonstrating the atypical target recognition by T. brucei 14-3-3 proteins. We found that the putative T. brucei protein phosphatase 2C (PP2c) binds to T. brucei 14-3-3 proteins utilizing its mode 3 motif (-pS/pTx(1-2)-COOH, where x is not Pro). We constructed eight chimeric PP2c proteins replacing its authentic mode 3 motif with potential mode 3 sequences found in Trypanosoma brucei genome database, and tested their binding. As a result, T. brucei 14-3-3 proteins interacted with three out of eight chimeric proteins including two with high affinity. Importantly, T. brucei 14-3-3 proteins co-immunoprecipitated with an uncharacterized full-length protein containing identified high-affinity mode 3 motif, suggesting that both proteins form a complex in vivo. In addition, a synthetic peptide derived from this mode 3 motif binds to T. brucei 14-3-3 proteins with high affinity. CONCLUSION/SIGNIFICANCE: Because of the atypical target recognition of T. brucei 14-3-3 proteins, no 14-3-3-binding proteins have been successfully identified in T. brucei until now whereas over 200 human 14-3-3-binding proteins have been identified. This report describes the first discovery of the T. brucei 14-3-3-binding proteins and their binding motifs. The high-affinity phosphopeptide will be a powerful tool to identify novel T. brucei 14-3-3-binding proteins. |
| first_indexed | 2024-12-21T08:25:48Z |
| format | Article |
| id | doaj.art-f5dee102d4684b4cb7a72fc58466fe4c |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-21T08:25:48Z |
| publishDate | 2010-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-f5dee102d4684b4cb7a72fc58466fe4c2022-12-21T19:10:19ZengPublic Library of Science (PLoS)PLoS ONE1932-62032010-01-01512e1556610.1371/journal.pone.0015566Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition.Masahiro InoueKouichi YasudaHaruki UemuraNatsumi YasakaHiroshi InoueYoshitatsu SeiNobuo HorikoshiToshihide FukumaBACKGROUND: The 14-3-3 proteins are structurally conserved throughout eukaryotes and participate in protein kinase signaling. All 14-3-3 proteins are known to bind to evolutionally conserved phosphoserine-containing motifs (modes 1 and/or 2) with high affinity. In Trypanosoma brucei, 14-3-3I and II play pivotal roles in motility, cytokinesis and the cell cycle. However, none of the T. brucei 14-3-3 binding proteins have previously been documented. METHODOLOGY/PRINCIPAL FINDINGS: Initially we showed that T. brucei 14-3-3 proteins exhibit far lower affinity to those peptides containing RSxpSxP (mode 1) and RxY/FxpSxP (mode 2) (where x is any amino acid residue and pS is phosphoserine) than human 14-3-3 proteins, demonstrating the atypical target recognition by T. brucei 14-3-3 proteins. We found that the putative T. brucei protein phosphatase 2C (PP2c) binds to T. brucei 14-3-3 proteins utilizing its mode 3 motif (-pS/pTx(1-2)-COOH, where x is not Pro). We constructed eight chimeric PP2c proteins replacing its authentic mode 3 motif with potential mode 3 sequences found in Trypanosoma brucei genome database, and tested their binding. As a result, T. brucei 14-3-3 proteins interacted with three out of eight chimeric proteins including two with high affinity. Importantly, T. brucei 14-3-3 proteins co-immunoprecipitated with an uncharacterized full-length protein containing identified high-affinity mode 3 motif, suggesting that both proteins form a complex in vivo. In addition, a synthetic peptide derived from this mode 3 motif binds to T. brucei 14-3-3 proteins with high affinity. CONCLUSION/SIGNIFICANCE: Because of the atypical target recognition of T. brucei 14-3-3 proteins, no 14-3-3-binding proteins have been successfully identified in T. brucei until now whereas over 200 human 14-3-3-binding proteins have been identified. This report describes the first discovery of the T. brucei 14-3-3-binding proteins and their binding motifs. The high-affinity phosphopeptide will be a powerful tool to identify novel T. brucei 14-3-3-binding proteins.http://europepmc.org/articles/PMC3006207?pdf=render |
| spellingShingle | Masahiro Inoue Kouichi Yasuda Haruki Uemura Natsumi Yasaka Hiroshi Inoue Yoshitatsu Sei Nobuo Horikoshi Toshihide Fukuma Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. PLoS ONE |
| title | Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. |
| title_full | Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. |
| title_fullStr | Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. |
| title_full_unstemmed | Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. |
| title_short | Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition. |
| title_sort | phosphorylation dependent protein interaction with trypanosoma brucei 14 3 3 proteins that display atypical target recognition |
| url | http://europepmc.org/articles/PMC3006207?pdf=render |
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