Abstract
Heat shock protein 90 (HSP90) is a molecular chaperone that supervises folding of cellular signaling proteins such as steroid receptors and many protein kinases. HSP90 relies on ATP hydrolysis for powering a conformational circuit that helps fold the client protein. To that end, HSP90 binds to co-chaperone proteins that regulate ATP hydrolysis rate or interaction with client proteins. Co-chaperones such as P23, cell division cycle 37 (CDC37), or activator of HSP90 ATPase activity 1 (AHA1) interact with the N-terminal or middle domain of HSP90, whereas others, such as HSP70/HSP90-organizing protein (HOP), use tetratricopeptide repeat (TPR) domains to bind the EEVD motif at the very C-terminal end of HSP90. Recently, the lysine methyltransferase SET and MYND domain-containing 2 (SMYD2) has been proposed as an HSP90-binding partner, and interaction analyses indicate that SMYD2 binding to HSP90 is independent of the EEVD motif. Using the amplified luminescence proximity homogeneous assay (Alpha) technique, I identified a new (M/I/L/V)PXL motif at the C termini of HSP90 and P23 that mediates an interaction with SMYD2, and synthetic peptides harboring this motif dissociated this complex. Of note, the HSP90- and P23-dependent client estrogen receptor α (ERα), was a major methylation target of SMYD2. In a reconstituted system in bacteria, I analyzed HSP90/P23-associated, SMYD2-mediated ERα methylation and found that when SMYD2 binds to the molecular chaperones, it considerably increases methylation of Lys-266 in ERα. Because methylation represses ERα activity, the observed complex formation between SMYD2 and HSP90/P23 may contribute to ERα regulation.
Highlights
Heat shock protein 90 (HSP90) is a molecular chaperone that supervises folding of cellular signaling proteins such as steroid receptors and many protein kinases
SET and MYND domain– containing 2 (SMYD2) and HSP90-organizing protein (HOP) were mixed with HSP90␣, and the interaction was analyzed by gel filtration chromatography (19 –21)
The present study shows that the lysine methyltransferase SMYD2 binds to the C-terminal domain of HSP90, yet independent of the EEVD motif that is used by tetratricopeptide repeat (TPR) domain cochaperones such as HOP for interaction (7, 8)
Summary
A motif in HSP90 and P23 that links molecular chaperones to efficient estrogen receptor ␣ methylation by the lysine methyltransferase SMYD2. HSP90 binds to co-chaperone proteins that regulate ATP hydrolysis rate or interaction with client proteins. HSP90 is assisted by a multitude of cochaperone proteins that modulate its ATP hydrolysis rate or mediate the interaction with client proteins Some cochaperones, such as P23, CDC37, or AHA1, interact with the N-terminal domain or the middle domain of the molecular chaperone (6). Others, such as HOP, CHIP, DNAJC7, PP5 (protein phosphatase 5), and the immunophilins, use tetratricopeptide repeat (TPR)[2] domains to clamp the C-terminal EEVD motif of HSP90 for interaction (7–9). HSP90/P23 contribute to another layer of ER␣ regulation by SMYD2-dependent methylation
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