The endoplasmic reticulum (ER) chaperone Grp94 is required for the correct folding and secretion of insulin-like growth factors (IGF), but the underlying mechanism is not understood. Our goal is to discover how Grp94 assists the folding of IGF proteins. Grp94 is a member of the Hsp90 chaperone family, which require ATP- driven conformational cycling to chaperone their ?client proteins?. Many oncogenic proteins depend on Hsp90 for their function. As a result, ATP-competitive inhibitors of Hsp90 have been extensively investigated as anti- cancer drugs. Although cytosolic Hsp90s were the first intended targets of inhibitors in clinical trials, Grp94 is receiving increasing attention as a drug target. Grp94 can receive clients from BiP, the Hsp70-family chaperone in the ER. Through a chance discovery, we have found that the combination of BiP and Hsp90 inhibitors locks Grp94 in a novel conformation, a state apparently poised for client transfer from BiP to Grp94. Client transfer from BiP to Grp94 requires two sequential steps: handover of the client from BiP to Grp94, followed by BiP dissociation from the ternary complex. Our first hypothesis is that Hsp90 inhibitors block both steps: Hsp90 inhibitors stall Grp94 and BiP in a client transfer state because ATP-dependent conformational changes of Grp94 are required to handover the client from BiP and then displace BiP from Grp94. We will test this hypothesis with IGF2, as we have discovered that this Grp94 client also has a well-defined BiP binding site.
Aim1 : Determine whether ATP-dependent conformational changes of Grp94 cause BiP to handover a bound client protein, and whether Hsp90 inhibitors disrupt this client-transfer function.
Aim2 : Determine whether ATP- dependent conformational changes of Grp94 are required to actively displace BiP from Grp94.
Aims 1 &2 will test a unified mechanism that can explain how Hsp90 inhibitors disrupt Grp94 function, but does not explain what essential chaperoning function is provided by BiP and Grp94 for IGF proteins. We have discovered that IGF2 forms dynamic oligomers. Similar to other peptide hormones that are known to oligomerize, this self-association may enable IGF2 to be effectively concentrated and packaged prior to its export to the Golgi. Despite a plausible biological role for IGF2 oligomerization, we have also observed that IGF2 can transition from dynamic reversible oligomers to irreversible aggregates. Thus, our second hypothesis is that the essential chaperoning function of BiP and Grp94 is to maintain a dynamic oligomerization state of IGF2. This idea is tested in Aim3: Determine the influence of Grp94 and BiP on the oligomerization properties of ProIGF2.
The endoplasmic reticulum (ER) chaperone Grp94 is required for the correct folding and secretion of insulin-like growth factors (IGFs), but the underlying mechanism is not understood. Our goal is to discover how Grp94 assists the folding of IGF proteins.
| Halpin, Jackson C; Street, Timothy O (2017) Hsp90 Sensitivity to ADP Reveals Hidden Regulation Mechanisms. J Mol Biol 429:2918-2930 |
| Jin, Yi; Hoxie, Reyal S; Street, Timothy O (2017) Molecular mechanism of bacterial Hsp90 pH-dependent ATPase activity. Protein Sci 26:1206-1213 |
| Halpin, Jackson C; Huang, Bin; Sun, Ming et al. (2016) Crowding Activates Heat Shock Protein 90. J Biol Chem 291:6447-55 |
| Liu, Shanshan; Street, Timothy O (2016) 5'-N-ethylcarboxamidoadenosine is not a paralog-specific Hsp90 inhibitor. Protein Sci 25:2209-2215 |
