HSPA13 Human

What is HSPA13 and how does it differ from other HSP70 family members?

HSPA13 (Heat Shock Protein Family A Member 13) is a unique 60 kDa protein belonging to the HSP70 family. Unlike classic heat shock proteins such as HSPA1A and HSPA1B that are induced by heat stress, HSPA13 expression is triggered specifically by calcium stress, not heat shock . This fundamental difference in stress response makes HSPA13 distinct within the HSP70 family.

While most HSP70 proteins play general roles in protein folding and refolding of denatured proteins, HSPA13 appears to have specialized functions related to the endoplasmic reticulum (ER) and protein import machinery. It localizes primarily to the ER and interacts with the Sec61 translocon complex and its associated factors .

The molecular structure of HSPA13 includes a conserved ATPase structural domain that is characteristic of the HSP70 family, enabling its chaperone functions through ATP-dependent mechanisms . This structural similarity with other HSP70s is maintained despite its functional specialization.

Where is HSPA13 expressed in human tissues and cells?

HSPA13 exhibits a relatively broad expression pattern across human tissues, though its expression levels vary significantly. Unlike some HSP70 family members such as HSPA2 and HSPA1L that show testis-specific expression patterns , HSPA13 is not confined to a single tissue type.

Research has demonstrated elevated HSPA13 expression in various cancer tissues compared to corresponding non-cancerous tissues. For instance, HSPA13 is highly expressed in hepatocellular carcinoma (HCC) tissues, where its expression correlates with poor clinical prognosis and vascular invasion .

At the subcellular level, HSPA13 primarily localizes to the endoplasmic reticulum membrane, where it associates with the protein translocation machinery. Proximity labeling experiments have confirmed its interaction with the Sec61 translocon complex, positioning it as an important factor in ER protein import processes .

What are the primary cellular functions of HSPA13?

HSPA13 serves several critical functions in cellular physiology:

• Regulation of protein translocation: HSPA13 regulates the import of secretory and membrane proteins through the Sec61 translocon into the endoplasmic reticulum. Both overexpression and knockout studies have demonstrated that proper HSPA13 levels are essential for efficient protein translocation, as either condition disrupts normal protein import .

• Maintenance of proteostasis: HSPA13 helps maintain both ER and cytosolic protein homeostasis. Research shows that HSPA13 knockout destabilizes proteostasis and increases sensitivity to ER disruption, highlighting its role in ensuring proper protein folding and processing .

• Regulation of secretory protein maturation: HSPA13 appears to influence the maturation of secretory proteins. Studies using transthyretin (TTR) as a model secretory protein demonstrated that HSPA13 overexpression inhibits TTR import into the ER and affects its maturation process .

• Cell signaling regulation: HSPA13 participates in signaling pathways that control cellular processes such as epithelial-mesenchymal transition (EMT). Evidence shows that HSPA13 knockdown inhibits TGFβ1-induced EMT and migration in retinal pigment epithelial cells, suggesting a role in regulating cell plasticity .

How does HSPA13 regulate protein import through the ER translocon?

HSPA13 regulates protein import through the ER translocon via several mechanisms:

• Translocon interaction: Mass spectrometry analysis has revealed that HSPA13 interacts with multiple components of the Sec61 translocon complex, including Sec61a1, Sec61b, NCLN, Sec62, and Sec63. It also associates with the signal recognition particle receptor component SRPRB and the translocon unclogger ZMPSTE24 .

• Signal peptidase complex interaction: HSPA13 interacts with four of the five components of the translocon-associated signal peptidase complex (SEC11C, SEC11A, SPCS2, and SPCS3), suggesting a role in signal peptide processing during protein import .

• Oligosaccharyltransferase (OST) complex interaction: HSPA13 associates with seven of the twelve components of the OST complexes (RPN1, RPN2, DAD1, DDOST, MAGT1, STT3A, and STT3B), indicating potential involvement in N-linked glycosylation of nascent proteins .

• ATPase-dependent regulation: HSPA13's ATPase activity appears crucial for proper protein translocation. ATPase-inactive mutants of HSPA13 exacerbate the inhibition of translocation and maturation of secretory proteins, leading to accumulation and aggregation of immature proteins in the cytosol .

This regulatory function appears to be finely balanced, as both overexpression and deficiency of HSPA13 can disrupt normal protein import processes, indicating that proper HSPA13 levels are essential for maintaining ER and cytosolic proteostasis.

What is the role of HSPA13 in cancer progression?

HSPA13 has been implicated in cancer progression, particularly in hepatocellular carcinoma (HCC):

• Elevated expression: HSPA13 is highly expressed in HCC tissues compared to non-tumor tissues, and this elevated expression correlates with poor clinical prognosis . The expression of HSPA13 is significantly higher in tumor tissues compared to corresponding non-tumor tissues across multiple cancer types .

• Promotion of tumor aggressiveness: Upregulation of HSPA13 is significantly associated with vascular invasion in HCC patients, suggesting a role in promoting tumor invasiveness and metastatic potential .

• Functional impact: Knockdown experiments have demonstrated that HSPA13 promotes HCC proliferation, migration, and invasion, confirming its oncogenic function in hepatocellular carcinogenesis .

• Molecular mechanism: HSPA13 exerts its oncogenic effects in HCC by interacting with TANK (TRAF family member-associated NF-κB activator) to inhibit its ubiquitination and degradation. The expression of HSPA13 and TANK are positively correlated in HCC tissues, suggesting a cooperative role in promoting cancer progression .

• Therapeutic potential: The identification of HSPA13's oncogenic function suggests that it may serve as a promising target for the diagnosis and treatment of HCC and potentially other cancers where it is overexpressed .

How is HSPA13 involved in epithelial-mesenchymal transition (EMT)?

HSPA13 plays a significant role in the process of epithelial-mesenchymal transition (EMT), a critical cellular transformation process involved in both development and pathological conditions:

• TGFβ1-induced EMT: In retinal pigment epithelial (RPE) cells, TGFβ1 treatment leads to increased intracellular Ca²⁺ levels, which subsequently upregulates HSPA13 expression. This cascade suggests that HSPA13 is a downstream mediator of TGFβ1-induced EMT .

• Functional significance: Knockdown of HSPA13 inhibits TGFβ1-induced EMT and migration in RPE cells, demonstrating its essential role in this cellular transformation process .

• In vivo relevance: In a rat model of proliferative vitreoretinopathy (PVR), HSPA13 is expressed in epiretinal membranes (ERMs), and its knockdown in RPE cells reduced the development of PVR, suggesting clinical relevance for this function .

• Molecular pathway: Mechanistically, downregulation of HSPA13 hinders the phosphorylation of PI3K/Akt in TGFβ1-induced RPE cells, indicating that HSPA13 may promote EMT through activation of the PI3K/Akt signaling pathway .

• Shared mechanisms with cancer: The involvement of HSPA13 in EMT represents a mechanistic parallel between fibrogenesis and cancer progression, as both processes involve similar cellular transformation events .

What experimental models are most effective for studying HSPA13 function?

Several experimental models have proven effective for investigating HSPA13 function:

• Cell culture models:

  • Human embryonic kidney 293T (HEK293T) cells have been successfully used to study HSPA13's role in protein translocation and proteostasis .

  • Human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cells serve as an excellent model for studying HSPA13's involvement in EMT processes .

  • HCT116 human cells have been used to investigate HSPA13 knockout effects on sensitivity to signal peptidase inhibitors .

• Gene manipulation approaches:

  • CRISPR/Cas9 system: This has been effectively used to generate knockout models for HSP family genes, similar to the approach used for Hspa1l knockout mice . This approach could be adapted for HSPA13 studies.

  • shRNA-mediated knockdown: This approach has been successfully employed to downregulate HSPA13 expression in both in vitro and in vivo studies, particularly in investigating its role in EMT and cancer progression .

• In vivo models:

  • Rat model of proliferative vitreoretinopathy (PVR): This model has been used to study HSPA13's role in EMT and fibrotic membrane formation in retinal disease .

  • Xenograft models: These would be appropriate for studying HSPA13's role in cancer progression, particularly for validating findings from in vitro studies of hepatocellular carcinoma .

• Protein translocation assays:

  • Transthyretin (TTR) serves as an excellent model substrate for studying ER protein import processes due to its well-characterized biophysical parameters and the clear distinction between its mature and immature forms on SDS-PAGE .

What methodological approaches are most effective for studying HSPA13 interactions?

Researchers investigating HSPA13 protein interactions have successfully employed several complementary methodological approaches:

• Proximity labeling methods:

  • BioID or TurboID: These proximity-dependent biotinylation approaches have been used to identify proteins that interact with HSPA13 in its native cellular environment. These methods are particularly valuable for detecting transient or weak interactions in the ER membrane .

• Mass spectrometry-based techniques:

  • Tandem Mass Tag (TMT)-MS2 experiments: This approach has been used to quantitatively assess HSPA13 interactors, although researchers should be aware of ratio compression issues often seen in TMT-MS2 experiments .

  • The raw mass spectrometry data associated with HSPA13 interaction studies have been deposited in the PRIDE archive (accession number PXD033498), providing a valuable resource for comparative analyses .

• Co-immunoprecipitation assays:

  • These traditional protein-protein interaction assays remain valuable for validating interactions identified through high-throughput approaches and for studying the dynamics of specific interactions, such as that between HSPA13 and TANK in hepatocellular carcinoma .

• Functional validation approaches:

  • ATPase-inactive mutants: Generation of ATPase-deficient HSPA13 variants has proven useful for understanding the functional significance of its enzymatic activity in protein translocation .

  • Domain mapping: Systematic analysis of protein domains can help identify critical regions required for specific protein-protein interactions.

What are the current limitations in understanding HSPA13 function?

Despite significant progress, several limitations persist in our understanding of HSPA13 function:

• Incomplete characterization of tissue-specific roles: While HSPA13 has been studied in the context of hepatocellular carcinoma and retinal pigment epithelial cells, its function in many other tissues remains poorly characterized. This limits our understanding of how HSPA13 dysfunction might contribute to various tissue-specific pathologies.

• Unclear stress response specificity: While HSPA13 is known to respond to calcium stress rather than heat shock , the precise mechanisms governing this specificity and the full spectrum of stress conditions that modulate HSPA13 expression remain incompletely defined.

• Limited understanding of regulation: The transcriptional, post-transcriptional, and post-translational mechanisms that regulate HSPA13 expression and activity under normal and pathological conditions are not fully elucidated.

• Incomplete characterization of interactome: While several HSPA13 interactors have been identified, particularly in the context of the ER translocon , a comprehensive understanding of its full interactome across different cellular conditions is lacking.

• Unclear relationship with other HSP family members: The functional redundancy or cooperation between HSPA13 and other HSP family members, particularly in protein quality control and stress response pathways, requires further investigation.

How does HSPA13 research contribute to understanding broader cellular stress responses?

HSPA13 research provides valuable insights into broader cellular stress response mechanisms:

• ER stress responses: HSPA13's role in protein translocation and ER proteostasis offers a window into how cells maintain protein homeostasis during ER stress conditions. HSPA13 knockout has been shown to destabilize proteostasis and increase sensitivity to ER disruption .

• Calcium signaling integration: HSPA13 represents an important link between calcium signaling and protein quality control mechanisms, as its expression is triggered by calcium stress rather than heat shock . This provides insights into how different stress signals are integrated and responded to at the cellular level.

• EMT regulation: Research on HSPA13's role in TGFβ1-induced epithelial-mesenchymal transition reveals connections between cellular stress responses and tissue remodeling processes. This has implications for understanding fibrotic diseases and cancer progression .

• Cancer stress adaptation: HSPA13's oncogenic function in hepatocellular carcinoma highlights how cancer cells can co-opt stress response proteins to promote their proliferation, survival, and metastatic potential .

• Proteostasis network integration: HSPA13 research contributes to our understanding of how different components of the cellular proteostasis network cooperate to maintain protein homeostasis under various stress conditions, particularly in the context of the secretory pathway.