Recombinant Human Hsp90 co-chaperone Cdc37-like 1 (CDC37L1)

What is CDC37L1 and how does it differ from CDC37?

CDC37L1 (Cell division cycle 37 like 1) is an analogue of CDC37 that shares 31% homology with the CDC37 protein . Unlike CDC37, which is well-documented to promote cancer development by regulating multiple protein kinases, CDC37L1 appears to function primarily as a tumor suppressor in various cancers, including gastric cancer . While CDC37 enhances the activity of cancer-promoting kinases such as CDK2, Erk, Akt, and mTOR, CDC37L1 exhibits opposite effects, particularly through its regulation of CDK6 expression .

What are the expression patterns of CDC37L1 in normal versus cancer tissues?

Immunohistochemical studies have revealed that CDC37L1 shows weak expression in high-grade gastric cancer tissues compared to low-grade tissues . Online database analyses further confirm that CDC37L1 levels are decreased in stage 4 gastric cancer patients . Similar expression patterns have been observed in other cancer types - CDC37L1 is downregulated in HBV-related hepatocellular carcinoma tissues compared to non-cancerous liver tissues, and specifically expressed in non-tumor nasopharyngeal epithelial tissues while showing reduced expression in nasopharyngeal carcinoma . These consistent findings across multiple cancer types suggest CDC37L1 downregulation may be a common feature in advanced malignancies.

What are effective methods for modulating CDC37L1 expression in experimental models?

Transient transfection of CDC37L1 plasmid or siRNA has proven effective for overexpression or knockdown experiments in gastric cancer cell lines . For researchers designing similar experiments, Western blot validation is essential to confirm successful modulation of CDC37L1 expression. CRISPR-Cas9 approaches have also been developed for CDC37L1 targeting, with guide RNA sequences designed to efficiently target the CDC37L1 gene with minimal off-target effects . The laboratory of Feng Zhang at the Broad Institute has developed optimized CDC37L1 CRISPR guide RNA sequences that researchers should consider for gene editing experiments .

Which assays are most appropriate for studying CDC37L1 function in cancer cells?

Based on published research, a comprehensive functional assessment of CDC37L1 should include multiple complementary assays:

• Proliferation assays: CCK8 assays for short-term effects and colony formation assays for long-term effects

• DNA synthesis measurement: EdU incorporation assays to quantify actively proliferating cells

• Migration capacity: Transwell chamber assays to assess cell migration potential

• Cell cycle analysis: Flow cytometry to determine cell cycle distribution changes

• In vivo tumor growth: Xenograft models in nude mice to validate in vitro findings

This multi-assay approach ensures robust characterization of CDC37L1's tumor-suppressive functions by examining different aspects of cancer cell behavior.

What protein interactions should be investigated when studying CDC37L1?

Studies indicate that CDC37L1, like CDC37, interacts with heat shock protein 90 (Hsp90) . Researchers should consider co-immunoprecipitation experiments to investigate CDC37L1-Hsp90 interactions, as well as potential interactions with tetratricopeptide repeat (TPR) proteins like Sti1, which has been shown to interact with CDC37 in yeast . Additionally, CDC37L1's effects on CDK6 expression suggest a functional relationship that warrants further investigation through protein-protein interaction studies .

How does CDC37L1 regulate cell cycle progression in cancer cells?

CDC37L1 appears to regulate the cell cycle primarily through its effects on CDK6 expression . In gastric cancer cells, CDC37L1 knockdown leads to increased CDK6 protein levels and an accumulation of cells in S phase, indicating enhanced cell cycle progression . Conversely, CDC37L1 overexpression results in reduced CDK6 levels. Importantly, treatment with Palbociclib, a specific CDK4/6 inhibitor, blocks the increased proliferation and S phase accumulation induced by CDC37L1 silencing . This suggests that CDC37L1's tumor-suppressive effects are mediated, at least in part, through CDK6 inhibition.

Table 1: CDC37L1's effect on cell cycle regulators in gastric cancer cells

What is the potential significance of CDC37L1-Hsp90 interactions in cancer?

While CDC37L1 is known to be an Hsp90 co-chaperone, the specific functional consequences of this interaction in cancer remain incompletely understood . Research in yeast has shown that CDC37 interacts with other Hsp90 co-chaperones such as Sti1 and Cpr7, suggesting that CDC37L1 may similarly form varied protein complexes that expand its functional diversity . The lethal effect of combined mutations in yeast CDC37 and Sti1 suggests these interactions may be essential for viability . Future research should investigate whether CDC37L1's tumor-suppressive functions are dependent on or independent of its interactions with Hsp90 and other co-chaperones.

What are the challenges in developing CDC37L1-targeted therapeutic approaches?

As CDC37L1 functions as a tumor suppressor, therapeutic strategies would likely aim to restore or enhance its expression/function rather than inhibit it. This presents several challenges:

• Delivery systems: Developing effective methods to deliver CDC37L1 or activators to tumor cells

• Specificity: Ensuring that interventions specifically target CDC37L1 without affecting CDC37, which has opposing functions

• Patient selection: Identifying biomarkers to select patients most likely to benefit from CDC37L1-targeted approaches

• Combination strategies: Determining optimal combinations with existing therapies, particularly CDK4/6 inhibitors like Palbociclib

Given CDC37L1's apparent regulation of CDK6, researchers might explore combining CDC37L1-enhancing approaches with existing CDK4/6 inhibitors to achieve synergistic effects.

What additional signaling pathways might be influenced by CDC37L1?

While CDC37L1's regulation of CDK6 has been established in gastric cancer , comprehensive pathway analyses are needed to identify other potential targets. CDC37L1 may influence additional kinases or signaling molecules through its co-chaperone functions. Phosphoproteomic and interactome analyses would be valuable approaches to identify the full spectrum of CDC37L1-regulated pathways. Research should also investigate whether CDC37L1 affects post-translational modifications of target proteins, as suggested by its co-chaperone function.

How might epigenetic regulation contribute to CDC37L1 expression patterns in cancer?

The consistent downregulation of CDC37L1 in advanced cancers raises questions about the regulatory mechanisms controlling its expression. Future studies should investigate potential epigenetic modifications (DNA methylation, histone modifications) at the CDC37L1 gene locus across cancer types. Understanding these regulatory mechanisms might reveal approaches to restore CDC37L1 expression in tumors. Additionally, analysis of transcription factor binding sites in the CDC37L1 promoter could identify key regulators that might be targeted therapeutically.

What are the important quality control measures for recombinant CDC37L1 protein production?

When producing recombinant human CDC37L1 for experimental use, researchers should implement several quality control measures:

• Purity assessment: SDS-PAGE and Western blotting to confirm protein identity and purity

• Functional validation: Co-immunoprecipitation with known binding partners like Hsp90

• Structural integrity: Circular dichroism to assess proper protein folding

• Batch consistency: Lot-to-lot comparison of activity in standardized assays

• Endotoxin testing: Especially important for in vivo applications

Maintaining proper storage conditions (-80°C for long-term storage, minimal freeze-thaw cycles) is crucial to preserve protein activity.

What cell line models are most appropriate for CDC37L1 functional studies?

Successful CDC37L1 research has been conducted using gastric cancer cell lines including MGC-803 and BGC-823 . These cell lines show detectable baseline expression of CDC37L1 and respond to both overexpression and knockdown approaches. When selecting cell models for CDC37L1 studies in other cancer types, researchers should first screen for baseline CDC37L1 expression and consider using paired cell lines with different levels of malignancy to better understand CDC37L1's role in cancer progression. Additionally, comparing results across multiple cell lines helps ensure findings are not cell line-specific artifacts.

How can researchers address the challenge of CDC37L1 antibody specificity?

Given the 31% homology between CDC37L1 and CDC37 , antibody cross-reactivity is a potential concern in CDC37L1 research. Researchers should:

• Validate antibodies using positive controls (CDC37L1-overexpressing cells) and negative controls (CDC37L1-knockdown cells)

• Perform parallel detection of both CDC37L1 and CDC37 to confirm specificity

• Consider using epitope-tagged CDC37L1 constructs for overexpression studies

• When possible, verify key findings using multiple antibodies targeting different epitopes of CDC37L1

These precautions help ensure experimental observations are truly attributable to CDC37L1 rather than cross-reactivity with CDC37.