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InterAKTions with FKBPs--mutational and pharmacological exploration.

Fabian AK, März A, Neimanis S, Biondi RM, Kozany C, Hausch F - PLoS ONE (2013)

Bottom Line: The FK506-binding protein 51 (FKBP51) is an Hsp90-associated co-chaperone which regulates steroid receptors and kinases.The FKBP51-Akt interaction is sensitive to the conformation of Akt1, but does not depend on the FK506-binding pocket of FKBP51.Therefore, FKBP inhibitors are unlikely to inhibit the Akt-FKBP-PHLPP network.

View Article: PubMed Central - PubMed

Affiliation: Research Group Chemical Genomics, Max Planck Institute of Psychiatry, Munich, Germany.

ABSTRACT
The FK506-binding protein 51 (FKBP51) is an Hsp90-associated co-chaperone which regulates steroid receptors and kinases. In pancreatic cancer cell lines, FKBP51 was shown to recruit the phosphatase PHLPP to facilitate dephosphorylation of the kinase Akt, which was associated with reduced chemoresistance. Here we show that in addition to FKBP51 several other members of the FKBP family bind directly to Akt. FKBP51 can also form complexes with other AGC kinases and mapping studies revealed that FKBP51 interacts with Akt via multiple domains independent of their activation or phosphorylation status. The FKBP51-Akt1 interaction was not affected by FK506 analogs or Akt active site inhibitors, but was abolished by the allosteric Akt inhibitor VIII. None of the FKBP51 inhibitors affected AktS473 phosphorylation or downstream targets of Akt. In summary, we show that FKBP51 binds to Akt directly as well as via Hsp90. The FKBP51-Akt interaction is sensitive to the conformation of Akt1, but does not depend on the FK506-binding pocket of FKBP51. Therefore, FKBP inhibitors are unlikely to inhibit the Akt-FKBP-PHLPP network.

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Other AGC kinases can also bind to FKBP51.A HEK293T cells were co-transfected in duplicates with GST-tagged Akt1 or ΔN_SGKS422D and FLAG-tagged FKBP51. After 48 h, the lysates were immunoprecipitated and analyzed in duplicates by Western blotting. B HeLa cells were co-transfected with FLAG-tagged FKBPs and HA-tagged S6K, treated with rapamycin (25 nM) or DMSO for 60 min and lysed. Lysates were immunoprecipitated and analyzed by Western blotting.
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pone-0057508-g002: Other AGC kinases can also bind to FKBP51.A HEK293T cells were co-transfected in duplicates with GST-tagged Akt1 or ΔN_SGKS422D and FLAG-tagged FKBP51. After 48 h, the lysates were immunoprecipitated and analyzed in duplicates by Western blotting. B HeLa cells were co-transfected with FLAG-tagged FKBPs and HA-tagged S6K, treated with rapamycin (25 nM) or DMSO for 60 min and lysed. Lysates were immunoprecipitated and analyzed by Western blotting.

Mentions: It was shown that FKBP51 binds to Akt1 and Akt2 but not to Akt3 [23]. To test whether the interaction of FKBP51 is specific to Akt or whether other AGC kinases could also interact with FKBP51 we performed co-immunoprecipitation experiments with SGK and p70S6K. Both wildtype SGK and SGK harboring an activating S422D mutation, clearly co-immunoprecipitated with FKBP51 to a similar extent as GST-tagged Akt1 (Figure 2A). FKBP51 and FKBP52 co-immunoprecipitated also with p70S6K overexpressed in HeLa cells (Figure 2B) while FKBP12 only marginally bound to p70S6K.


InterAKTions with FKBPs--mutational and pharmacological exploration.

Fabian AK, März A, Neimanis S, Biondi RM, Kozany C, Hausch F - PLoS ONE (2013)

Other AGC kinases can also bind to FKBP51.A HEK293T cells were co-transfected in duplicates with GST-tagged Akt1 or ΔN_SGKS422D and FLAG-tagged FKBP51. After 48 h, the lysates were immunoprecipitated and analyzed in duplicates by Western blotting. B HeLa cells were co-transfected with FLAG-tagged FKBPs and HA-tagged S6K, treated with rapamycin (25 nM) or DMSO for 60 min and lysed. Lysates were immunoprecipitated and analyzed by Western blotting.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585324&req=5

pone-0057508-g002: Other AGC kinases can also bind to FKBP51.A HEK293T cells were co-transfected in duplicates with GST-tagged Akt1 or ΔN_SGKS422D and FLAG-tagged FKBP51. After 48 h, the lysates were immunoprecipitated and analyzed in duplicates by Western blotting. B HeLa cells were co-transfected with FLAG-tagged FKBPs and HA-tagged S6K, treated with rapamycin (25 nM) or DMSO for 60 min and lysed. Lysates were immunoprecipitated and analyzed by Western blotting.
Mentions: It was shown that FKBP51 binds to Akt1 and Akt2 but not to Akt3 [23]. To test whether the interaction of FKBP51 is specific to Akt or whether other AGC kinases could also interact with FKBP51 we performed co-immunoprecipitation experiments with SGK and p70S6K. Both wildtype SGK and SGK harboring an activating S422D mutation, clearly co-immunoprecipitated with FKBP51 to a similar extent as GST-tagged Akt1 (Figure 2A). FKBP51 and FKBP52 co-immunoprecipitated also with p70S6K overexpressed in HeLa cells (Figure 2B) while FKBP12 only marginally bound to p70S6K.

Bottom Line: The FK506-binding protein 51 (FKBP51) is an Hsp90-associated co-chaperone which regulates steroid receptors and kinases.The FKBP51-Akt interaction is sensitive to the conformation of Akt1, but does not depend on the FK506-binding pocket of FKBP51.Therefore, FKBP inhibitors are unlikely to inhibit the Akt-FKBP-PHLPP network.

View Article: PubMed Central - PubMed

Affiliation: Research Group Chemical Genomics, Max Planck Institute of Psychiatry, Munich, Germany.

ABSTRACT
The FK506-binding protein 51 (FKBP51) is an Hsp90-associated co-chaperone which regulates steroid receptors and kinases. In pancreatic cancer cell lines, FKBP51 was shown to recruit the phosphatase PHLPP to facilitate dephosphorylation of the kinase Akt, which was associated with reduced chemoresistance. Here we show that in addition to FKBP51 several other members of the FKBP family bind directly to Akt. FKBP51 can also form complexes with other AGC kinases and mapping studies revealed that FKBP51 interacts with Akt via multiple domains independent of their activation or phosphorylation status. The FKBP51-Akt1 interaction was not affected by FK506 analogs or Akt active site inhibitors, but was abolished by the allosteric Akt inhibitor VIII. None of the FKBP51 inhibitors affected AktS473 phosphorylation or downstream targets of Akt. In summary, we show that FKBP51 binds to Akt directly as well as via Hsp90. The FKBP51-Akt interaction is sensitive to the conformation of Akt1, but does not depend on the FK506-binding pocket of FKBP51. Therefore, FKBP inhibitors are unlikely to inhibit the Akt-FKBP-PHLPP network.

Show MeSH
Related in: MedlinePlus