CDC37 Human
Description
Molecular Structure and Domains
CDC37 consists of three structural domains:
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N-terminal domain: Binds directly to client protein kinases, facilitating their recognition and recruitment to Hsp90 .
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Central domain: Mediates interaction with Hsp90, forming a chaperone-client complex .
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C-terminal domain: Functionally uncharacterized but speculated to modulate kinase-Hsp90 interactions .
A cryoEM study revealed that CDC37 mimics interactions between kinase N- and C-lobes, stabilizing kinases like Cdk4 in an unfolded state for Hsp90-dependent regulation .
Kinase Stabilization and Activation
CDC37 targets over 60% of the human kinome, including:
| Kinase Clients | Biological Roles | References |
|---|---|---|
| CDK4, CDK6 | Cell cycle regulation | |
| RAF-1, SRC | Signal transduction | |
| Aurora B | Chromosome segregation, cytokinesis | |
| LKB1 | Energy metabolism regulation |
CDC37 stabilizes kinase folding intermediates, enabling their activation by Hsp90 . Loss of CDC37 disrupts kinase stability, leading to cell cycle defects and apoptosis .
Hsp90 Coordination
CDC37 suppresses Hsp90 ATPase activity while recruiting kinases to the chaperone complex . This partnership is critical for maintaining oncogenic kinases like BRAF and Akt in a functional state .
Cancer
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Oncogenic Role: CDC37 overexpression promotes tumorigenesis in prostate, breast, and salivary gland cancers by stabilizing kinases like CDK4 and RAF-1 . Transgenic mice overexpressing CDC37 developed mammary adenosquamous carcinomas and lymphomas .
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Therapeutic Target: Inhibiting CDC37 disrupts Hsp90-dependent kinase stabilization, sensitizing cancer cells to proteasome inhibitors like Bortezomib (BTZ) .
Multiple Myeloma (MM)
CDC37 downregulation correlates with BTZ resistance and disease relapse. Suppression of CDC37 in MM cells reduces Xbp1s signaling, impairing plasma cell maturation and promoting drug resistance .
CryoEM Structure of Hsp90-CDC37-Cdk4 Complex
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Kinase Conformation: Cdk4 adopts an unfolded state with separated N- and C-lobes .
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CDC37 Mechanism: Mimics kinase αC-β4 loop interactions, stabilizing the open conformation .
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Hsp90 Interaction: Clamps around the kinase β5 strand, preventing refolding until activation .
Post-Translational Modifications
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Phosphorylation of CDC37 Ser13 by CK2 is essential for kinase recruitment .
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Hsp90 phosphorylation at specific sites modulates client binding and ATPase activity .
Biomarker Potential
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CDC37 is upregulated in early-stage prostate and breast cancers, serving as a prognostic marker .
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Reduced CDC37 expression in relapsed MM predicts BTZ resistance .
Therapeutic Strategies
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Small-Molecule Inhibitors: Targeting CDC37-Hsp90 interactions disrupts oncogenic kinases (e.g., BRAF, Akt) .
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Combination Therapy: Co-targeting CDC37 and proteasome pathways enhances efficacy in BTZ-resistant MM .
Recombinant CDC37 Production
Product Specs
Q&A
What is CDC37 and what is its primary function in human cells?
CDC37 is a 50 kDa molecular chaperone that physically stabilizes the catalytic domains of protein kinases, functioning as a wide-spectrum regulator of protein phosphorylation. It acts as a specialized co-chaperone that mediates the interaction between client kinases and HSP90. CDC37 was first discovered in a yeast screen for cell division cycle proteins, reflecting its essential role in promoting cell proliferation . In normal human tissues, CDC37 is typically expressed at low levels but becomes upregulated during periods of rapid cellular proliferation .
What is the structure of CDC37 and its functional domains?
CDC37 possesses two major structural domains with distinct functions:
| Domain | Location (human) | Function | Conservation |
|---|---|---|---|
| Kinase binding domain | Residues 1-126 | Client kinase interface | Most highly conserved region |
| HSP90-binding domain | Residues ~127-378 | Mediates association with HSP90 | Less conserved than N-terminal domain |
The N-terminal domain serves as the "client interface" and is essential for CDC37's growth-promoting functions. Within the HSP90-CDC37-BRAF^V600E complex, CDC37-Trp31 makes important contacts with the kinase that affect the conformation of catalytically important motifs such as the DFG motif . CDC37 also contains a C-terminal region that forms a loosely bound globular domain in complex with client proteins .
How is CDC37 regulated through post-translational modifications?
CDC37 undergoes several phosphorylation events that regulate its function, with phosphorylation of Ser13 by casein kinase 2 (CK2) being particularly critical. This phosphorylation permits stable interaction of CDC37 with client kinases and is essential for its chaperoning function .
Interestingly, CK2 itself is a CDC37 client, creating a positive feedback loop between the client kinase and CDC37 chaperoning . The dephosphorylation of Ser13 is mediated by protein phosphatase 5 (PP5), but only occurs when PP5 and CDC37 are bound simultaneously to the same HSP90 dimer . This creates a complex regulatory network sometimes referred to as the "Chaperone Code" .
Which experimental models are most suitable for studying CDC37 function?
| Experimental Model | Advantages | Limitations | Applications |
|---|---|---|---|
| Yeast systems (S. cerevisiae and S. pombe) | Simple genetic manipulation | Low sequence conservation (~20%) | Basic function studies |
| Mammalian cancer cell lines | Relevant to human disease | Complex signaling networks | Cancer-related studies |
| Transgenic mouse models | In vivo tumorigenesis | Time-consuming | Oncogenic cooperation studies |
| Insect cell expression systems | Production of protein complexes | Not mammalian | Structural studies |
| In vitro reconstitution | Biochemical precision | Lacks cellular context | Interaction studies |
For cancer-specific studies, human cancer cell lines with elevated CDC37 expression (such as prostate cancer cells) are particularly valuable . The HSP90-CDC37-BRAF^V600E complex has been successfully co-expressed and purified from insect cells for structural studies .
What is the role of CDC37 in oncogenic kinase stabilization and cancer progression?
CDC37 plays a crucial role in cancer as an overexpressed oncoprotein that mediates carcinogenesis by stabilizing mutant and/or overexpressed oncogenic kinases . It is particularly significant in prostate cancer, where immunohistological studies found CDC37 overexpression in 100% of neoplastic prostate tissue compared to normal prostate .
The oncogenic function of CDC37 extends beyond kinase stabilization to interaction with the androgen receptor (AR), a rare non-kinase CDC37 client critical for prostatic cell growth . This interaction may explain CDC37's particular significance in prostate cancer. When CDC37 is overexpressed in transgenic mice, cells become transformed, proliferate, and tumors arise . This effect is amplified when CDC37 is co-expressed with c-Myc, resulting in more and larger tumors .
The degree of dependency of the cancer cell kinome on CDC37 expression appears to be extensive, with at least 90% of protein kinases examined in a yeast screen requiring CDC37 for function .
How does extracellular/surface CDC37 differ functionally from intracellular CDC37?
CDC37 is not restricted intracellularly but is also present on the surface of cancer cells such as MDA-MB-453 and MDA-MB-231 human breast cancer cells . This surface pool of CDC37 participates in cancer cell motility processes, distinguishing it functionally from its intracellular counterpart .
Similar to intracellular CDC37, the surface pool specifically interacts with HSP90 as well as kinase receptors such as HER2 and EGFR on the cell surface, likely acting as a co-factor in HSP90's extracellular chaperoning activities . Functional inhibition of surface HSP90 using cell-impermeable antibodies (such as mAb 4C5) leads to disruption of the CDC37/HSP90 complex and inhibition of the CDC37/ErbB receptor complexes .
This extracellular function of CDC37 represents a potential target for therapeutic intervention using cell-impermeable agents that would not affect intracellular CDC37 functions required for normal cellular processes .
What is the structural basis of the HSP90-CDC37-client kinase complex?
The structural basis of the HSP90-CDC37-client kinase complex has been revealed through cryo-electron microscopy studies of the HSP90-CDC37-BRAF^V600E complex . This complex consists of two molecules of human HSP90β arranged in the ATP-bound closed conformation, with CDC37 and the BRAF^V600E kinase domain positioned between them .
Key structural features include:
| Component Interaction | Structural Details | Functional Significance |
|---|---|---|
| CDC37-Kinase | CDC37-Trp31 interacts with BRAF^V600E | Forces the DFG motif into a different conformation |
| HSP90-Kinase | Limited direct contacts with BRAF^V600E C-lobe | Peripheral support for kinase positioning |
| HSP90-BRAF^V600E | Involves residues 521-533 of BRAF^V600E | These would be part of the N-terminal lobe in folded kinase |
| Threading | BRAF^V600E chain threads between HSP90 monomers | Creates structural platform for kinase stabilization |
This structural arrangement explains why oncogenic mutations like BRAF^V600E show stronger dependence on HSP90-CDC37 for stability and activation . The V600E mutation destabilizes the activation segment's ordered inhibitory conformation, facilitating the conformational switch required for interaction with CDC37 .
What is the role of PP5 phosphatase in regulating CDC37 function?
Protein phosphatase 5 (PP5) plays a crucial role in regulating CDC37 function through dephosphorylation . PP5 has a tetratricopeptide (TPR) domain that confers high affinity for the MEEVD motif at the C-terminus of HSP90, allowing it to associate with HSP90-CDC37 complexes .
PP5 specifically dephosphorylates the critical pSer13 in CDC37, which is essential for regulating CDC37's interaction with client kinases . This dephosphorylation only occurs when both proteins are bound simultaneously to the same HSP90 dimer, ensuring spatial and temporal regulation .
Research has demonstrated that PP5 can dephosphorylate CDC37-pSer13 even when CDC37 is engaged in a complex with a client protein and HSP90, as shown with the HSP90-CDC37-BRAF^V600E complex . This activity has been implicated in the maturation/activation of HSP90-dependent client proteins .
The interplay between CK2 (which phosphorylates CDC37-Ser13) and PP5 (which dephosphorylates it) represents a key regulatory mechanism controlling the chaperoning function of CDC37 and the activation status of its client kinases .
How can CDC37 be effectively targeted for cancer therapy?
CDC37 represents a promising target for cancer therapy due to its overexpression in malignant cells and role in stabilizing oncogenic kinases . Several potential targeting approaches include:
| Targeting Strategy | Mechanism | Potential Advantage |
|---|---|---|
| Disrupting CDC37-client interactions | Targeting the kinase binding domain | May affect specific client subsets |
| Inhibiting CDC37-HSP90 interaction | Preventing ternary complex formation | Destabilizes multiple oncogenic kinases |
| Targeting post-translational modifications | Inhibiting CK2-mediated phosphorylation | Disrupts regulatory mechanisms |
| RNA interference approaches | Reducing CDC37 expression | Direct reduction of chaperone availability |
| Surface CDC37 targeting | Cell-impermeable antibodies | Spares intracellular CDC37 functions |
The therapeutic potential is supported by several observations: CDC37 is increased in proliferating tissues and heavily expressed in certain cancers while most normal tissues do not proliferate or require high CDC37 levels . This creates a potential therapeutic window not as obvious with HSP90 inhibition .
One agent, celastrol, has been reported to disrupt the CDC37-HSP90 complex, although further studies are needed to develop agents with more novel mechanisms of action .
What methodologies are effective for studying CDC37-client kinase interactions?
| Methodology | Application | Technical Considerations |
|---|---|---|
| Co-immunoprecipitation (Co-IP) | Detecting physical interactions | Requires specific antibodies |
| Cell-impermeable antibodies | Studying surface CDC37 | Doesn't affect intracellular pools |
| Recombinant protein expression | In vitro binding studies | Can use insect cell systems |
| Size exclusion chromatography | Purifying stable complexes | Useful for structural studies |
| Cryo-electron microscopy | Structural analysis | Revealed HSP90-CDC37-BRAF^V600E complex |
| Mutagenesis studies | Identifying key residues | Ser13 and Trp31 are critical |
| Phospho-specific antibodies | Detecting phosphorylation states | Used to monitor Ser13 phosphorylation |
| Functional kinase assays | Assessing kinase activity | Measures client functionality |
The combination of these approaches has revealed that CDC37 interacts with the N-lobe of client kinases and forces conformational changes that are important for kinase regulation and stabilization . The HSP90-CDC37-BRAF^V600E-PP5 complex has been successfully purified using size exclusion chromatography and analyzed by cryo-EM to understand the structural basis of these interactions .
How does phosphorylation of CDC37 at Ser13 affect its chaperoning activity?
Phosphorylation of CDC37 at Ser13 by casein kinase 2 (CK2) is critical for its chaperoning function . This phosphorylation permits stable interaction of CDC37 with client kinases, which is essential for the recruitment of kinases to the HSP90 chaperone system .
The phosphorylation creates a positive feedback mechanism, as CK2 itself is a CDC37 client, thus mediating a loop between the client kinase and CDC37 chaperoning . When CDC37 is engaged in a complex with a client protein and HSP90, PP5 can dephosphorylate CDC37-pSer13 in a time-dependent manner, which has been demonstrated using phospho-specific antibodies .
This phosphorylation-dephosphorylation cycle represents a molecular switch that regulates the chaperoning activity of CDC37 and influences the maturation and activation of HSP90-dependent client kinases . The timing of Ser13 dephosphorylation appears crucial for the client protein activation cycle .
What are the methodological challenges in studying CDC37-dependent signaling networks?
Studying CDC37-dependent signaling networks presents several methodological challenges:
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Client protein diversity: CDC37 interacts with approximately 90% of kinases in yeast screens, making it difficult to isolate specific client relationships .
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Structural flexibility: The CDC37-HSP90-client complexes show conformational flexibility, making structural studies challenging .
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Dynamic interactions: The interactions between CDC37, HSP90, and client kinases are regulated by ATP hydrolysis and post-translational modifications .
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Surface vs. intracellular pools: Studying the distinct functions of surface CDC37 requires specialized techniques to avoid affecting the intracellular pool .
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Species differences: The low sequence conservation between yeast and mammalian CDC37 (less than 20%) makes it challenging to extrapolate findings from yeast models to human systems .
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Context-dependent function: The function of CDC37 may vary depending on cell type or disease state, requiring multiple experimental models .
Addressing these challenges requires integrated approaches combining biochemical, structural, genetic, and systems biology methods to comprehensively understand CDC37's role in signaling networks.
Product Science Overview
CDC37 is known to form complexes with Heat Shock Protein 90 (Hsp90) and a variety of protein kinases, including CDK4, CDK6, SRC, RAF-1, MOK, and eIF2 alpha kinases . This interaction is critical for the proper functioning of these kinases, as CDC37 directs Hsp90 to its target kinases, thereby stabilizing them and promoting their activity . Additionally, CDC37 inhibits the ATPase activity of HSP90AA1, further contributing to the stabilization of the kinase-Hsp90 complex .
CDC37 is expressed in proliferative zones during embryonic development and in adult tissues, indicating its positive role in cell proliferation . It is required for cell division in budding yeast and is thought to play an important role in the establishment of signaling pathways that control cell growth and division . The protein’s ability to bind to numerous kinases and promote their interaction with the Hsp90 complex is essential for the regulation of cell cycle progression and signal transduction .
Mutations or dysregulation of CDC37 have been associated with various diseases, including Fanconi Anemia and Parkinson’s Disease . The protein’s involvement in multiple signaling pathways makes it a potential target for therapeutic interventions in diseases characterized by abnormal cell proliferation and signal transduction.
Recombinant human CDC37 protein is produced using recombinant DNA technology, which involves inserting the gene encoding CDC37 into a suitable expression system, such as bacteria or mammalian cells. This allows for the large-scale production of the protein, which can be used for research and therapeutic purposes. Recombinant CDC37 is used in various studies to understand its role in cell cycle regulation and to develop potential therapeutic strategies targeting its function .
