Mitochondrial antiviral-signalling protein is a client of the BAG6 protein quality control complex.

The heterotrimeric BAG6 complex coordinates the direct handover of newly synthesised tail-anchored (TA) membrane proteins from an SGTA-bound preloading complex to the endoplasmic reticulum (ER) delivery component TRC40. In contrast, defective precursors, including aberrant TA proteins, form a stable complex with this cytosolic protein quality control factor, enabling such clients to be either productively re-routed or selectively degraded. We identify the mitochondrial antiviral-signalling protein (MAVS) as an endogenous TA client of both SGTA and the BAG6 complex. Our data suggest that the BAG6 complex binds to a cytosolic pool of MAVS before its misinsertion into the ER membrane, from where it can subsequently be removed via ATP13A1-mediated dislocation. This BAG6-associated fraction of MAVS is dynamic and responds to the activation of an innate immune response, suggesting that BAG6 may modulate the pool of MAVS that is available for coordinating the cellular response to viral infection.

Canine REIC/Dkk-3 interacts with SGTA and restores androgen receptor signalling in androgen-independent prostate cancer cell lines.

BACKGROUND: The pathological condition of canine prostate cancer resembles that of human androgen-independent prostate cancer. Both canine and human androgen receptor (AR) signalling are inhibited by overexpression of the dimerized co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA), which is considered to cause the development of androgen-independency. Reduced expression in immortalised cells (REIC/Dkk-3) interferes with SGTA dimerization and rescues AR signalling. This study aimed to assess the effects of REIC/Dkk-3 and SGTA interactions on AR signalling in the canine androgen-independent prostate cancer cell line CHP-1. RESULTS: Mammalian two-hybrid and Halo-tagged pull-down assays showed that canine REIC/Dkk-3 interacted with SGTA and interfered with SGTA dimerization. Additionally, reporter assays revealed that canine REIC/Dkk-3 restored AR signalling in both human and canine androgen-independent prostate cancer cells. Therefore, we confirmed the interaction between canine SGTA and REIC/Dkk-3, as well as their role in AR signalling. CONCLUSIONS: Our results suggest that this interaction might contribute to the development of a novel strategy for androgen-independent prostate cancer treatment. Moreover, we established the canine androgen-independent prostate cancer model as a suitable animal model for the study of this type of treatment-refractory human cancer.

The cochaperone SGTA (small glutamine-rich tetratricopeptide repeat-containing protein alpha) demonstrates regulatory specificity for the androgen, glucocorticoid, and progesterone receptors.

Steroid hormone receptors are ligand-dependent transcription factors that require the ordered assembly of multichaperone complexes for transcriptional activity. Although heat shock protein (Hsp) 90 and Hsp70 are key players in this process, multiple Hsp70- and Hsp90-associated cochaperones associate with receptor-chaperone complexes to regulate receptor folding and activation. Small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA) was recently characterized as an Hsp70 and Hsp90-associated cochaperone that specifically regulates androgen receptor activity. However, the specificity of SGTA for additional members of the steroid hormone receptor superfamily and the mechanism by which SGTA regulates receptor activity remain unclear. Here we report that SGTA associates with and specifically regulates the androgen, glucocorticoid, and progesterone receptors and has no effect on the mineralocorticoid and estrogen receptors in both yeast and mammalian cell-based reporter assays. In both systems, SGTA knockdown/deletion enhances receptor activity, whereas SGTA overexpression suppresses receptor activity. We demonstrate that SGTA binds directly to Hsp70 and Hsp90 in vitro with similar affinities yet predominately precipitates with Hsp70 from cell lysates, suggesting a role for SGTA in early, Hsp70-mediated folding. Furthermore, SGTA expression completely abrogates the regulation of receptor function by FKBP52 (52-kDa FK506-binding protein), which acts at a later stage of the chaperone cycle. Taken together, our data suggest a role for SGTA at distinct steps in the chaperone-dependent modulation of androgen, glucocorticoid, and progesterone receptor activity.

High expression of SGTA in esophageal squamous cell carcinoma correlates with proliferation and poor prognosis.

Receptor tyrosine kinases (RTKs) expression and the growth factor such as platelet-derived growth factor (PDGF) and their receptors have been considered relevant in the process of angiogenesis and dissemination in esophageal squamous cell carcinoma (ESCC). Small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA) downstream of RTK signaling was a critical regulator of PDGF receptors (PDGFR) stability. The aim of the present study was to examine the expression of SGTA and to elucidate its clinicopathologic significance in ESCC. Immunohistochemistry and western blot analysis were performed for SGTA in ESCC samples. SGTA was up-regulated in ESCC as compared with the adjacent normal tissue. High expression of SGTA was associated with tumor grade (P < 0.01), and SGTA was positively correlated with proliferation marker Ki-67 (P < 0.05). Univariate analysis showed that SGTA expression did has a remarkable prediction for poor prognosis (P = 0.016). Knockdown or overexpression of SGTA affected ESCC cells proliferation and cell cycle. Additionally, after ESCC cells silenced for SGTA were treated with cisplatin (an anti-ESCC agent), the cell growth was down-regulated. These findings suggested that SGTA was involved in the pathogenesis of ESCC and might indicate a poor prognosis for ESCC patients.

SGTA associates with intracellular aggregates in neurodegenerative diseases.

Intracellular aggregates are a common pathological hallmark of neurodegenerative diseases such as polyglutamine (polyQ) diseases, amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and multiple system atrophy (MSA). Aggregates are mainly formed by aberrant disease-specific proteins and are accompanied by accumulation of other aggregate-interacting proteins. Although aggregate-interacting proteins have been considered to modulate the formation of aggregates and to be involved in molecular mechanisms of disease progression, the components of aggregate-interacting proteins remain unknown. In this study, we showed that small glutamine-rich tetratricopeptide repeat-containing protein alfa (SGTA) is an aggregate-interacting protein in neurodegenerative diseases. Immunohistochemistry showed that SGTA interacted with intracellular aggregates in Huntington disease (HD) cell models and neurons of HD model mice. We also revealed that SGTA colocalized with intracellular aggregates in postmortem brains of patients with polyQ diseases including spinocerebellar ataxia (SCA)1, SCA2, SCA3, and dentatorubral-pallidoluysian atrophy. In addition, SGTA colocalized with glial cytoplasmic inclusions in the brains of MSA patients, whereas no accumulation of SGTA was observed in neurons of PD and ALS patients. In vitro study showed that SGTA bound to polyQ aggregates through its C-terminal domain and SGTA overexpression reduced intracellular aggregates. These results suggest that SGTA may play a role in the formation of aggregates and may act as potential modifier of molecular pathological mechanisms of polyQ diseases and MSA.

The STI1-domain is a flexible alpha-helical fold with a hydrophobic groove.

STI1-domains are present in a variety of co-chaperone proteins and are required for the transfer of hydrophobic clients in various cellular processes. The domains were first identified in the yeast Sti1 protein where they were referred to as DP1 and DP2. Based on hidden Markov model searches, this domain had previously been found in other proteins including the mammalian co-chaperone SGTA, the DNA damage response protein Rad23, and the chloroplast import protein Tic40. Here, we refine the domain definition and carry out structure-based sequence alignment of STI1-domains showing conservation of five amphipathic helices. Upon examinations of these identified domains, we identify a preceding helix 0 and unifying sequence properties, determine new molecular models, and recognize that STI1-domains nearly always occur in pairs. The similarity at the sequence, structure, and molecular levels likely supports a unified functional role.

Tumor suppressor REIC/Dkk-3 and its interacting protein SGTA inhibit glucocorticoid receptor to nuclear transport.

REIC/Dkk-3 is a tumor suppressor, and its expression is significantly downregulated in a variety of human cancer types. A previous study performed yeast two-hybrid screening and identified the small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA), known as a negative modulator of cytoplasmic androgen receptor (AR) signaling, which is a novel interacting partner of REIC/Dkk-3. The previous study also indicated that the REIC/Dkk-3 protein interferes with the dimerization of SGTA and then upregulates the AR transport and signaling in human prostate cancer PC3 cells. Since the transport of some steroid receptors to nucleus is conducted similarly by dynein motor-dependent way, the current study aimed to investigate the role of SGTA and REIC/Dkk-3 in the transport of other glucocorticoid receptors (GR). In vitro reporter assays for the cytoplasmic GR transport were performed in human prostate cancer PC3 cells and 293T cells. As for the SGTA protein, a suppressive effect on the GR transport to the nucleus was observed in the cells. As for the REIC/Dkk-3 protein, an inhibitory effect was observed for the GR transport in PC3 cells. Under the depleted condition of SGTA by short-hairpin (sh)RNA, the downregulation of GR transport by REIC/Dkk-3 was significantly enhanced compared with the non-depleted condition in PC3 cells, suggesting a compensatory role of REIC/Dkk-3 in the SGTA mediated inhibition of GR transport. The current study therefore demonstrated that SGTA inhibited the cytoplasmic transport of GR in 293T and PC3 cells, and REIC/Dkk-3 also inhibited the cytoplasmic transport of GR in PC3 cells. These results may be used to gain novel insight into the GR transport and signaling in normal and cancer cells.

The roles of cytosolic quality control proteins, SGTA and the BAG6 complex, in disease.

SGTA is a co-chaperone that, in collaboration with the complex of BAG6/UBL4A/TRC35, facilitates the biogenesis and quality control of hydrophobic proteins, protecting them from the aqueous cytosolic environment. This work includes targeting tail-anchored proteins to their resident membranes, sorting of membrane and secretory proteins that mislocalize to the cytoplasm and endoplasmic reticulum-associated degradation of misfolded proteins. Since these functions are all vital for the cell's continued proteostasis, their disruption poses a threat to the cell, with a particular risk of protein aggregation, a phenomenon that underpins many diseases. Although the specific disease implications of machinery involved in quality control of hydrophobic substrates are poorly understood, here we summarize much of the available information on this topic.

[MicroRNA-138 regulates cell adhesion-mediated drug resistance phenotype by targeting SGTA in non-Hodgkin's lymphoma].

Objective: To analyze the effects of miR-138 on the expression of small glutamine-rich TPR-containing protein A (SGTA) and cell adhesion-mediated drug resistance (CAM-DR) phenotype in non-Hodgkin's lymphoma (NHL). Methods: The adhesion model was constructed using fibronectin (FN) or bone marrow stromal cells HS-5. The effect of miR-138 on the expression of SGTA was analyzed by Western blotting and RQ-PCR. Dual-luciferase assays were performed to probe the effects of miR-138 on SGTA 3' UTR activities. Subsequently, we investigated the effect of miR-138 on cell cycle, adhesion ability and CAM-DR. Moreover, the correlation between miR-138 expression and therapeutic response was analyzed in 35 paraffin-embedded diffuse large B cell lymphoma samples. Results: Our data showed that adhesion of NHL cells to FN or HS-5 cells significantly increased miR-138 expression (P<0.05). Knockdown of miR-138 markedly increased the protein (all P<0.05) but not for mRNA (all P>0.05) levels of SGTA in NHL cell. The luciferase activity of SGTA 3' UTR was significantly suppressed by miR-138 transfected cells (0.73±0.03 vs 1.00±0.02, t=0.914, P=0.002). No change in terms of reporter activity was observed in SGTA 3'UTR mutant transfected cells (0.93±0.04 vs 1.00±0.02, t=1.375, P=0.241). Also we found that ectopic expression of miR-138 significantly induced cell cycle arrest at G(1) phase in both suspension and adherent cells (all P<0.05). Knockdown of miR-138 had no effect on cell adhesion ability (all P>0.05). More importantly, in suspension cells, knockdown of miR-138 significantly decreased the percentage of doxorubicin-induced cell death. However, knockdown of miR-138 dramatically increased the percentage of doxorubicin-induced cell death in FN/HS-5-adherent cells. Furthermore, the miR-138 expression was significantly higher in patients with progression of disease/stable disease than those experiencing complete response/partial response (9.72±1.11 vs 3.06±0.22, t=9.144, P<0.001). Conclusion: MiR-138 promoted CAM-DR phenotype through cell adhesion-mediated SGTA down-regulation and cell cycle arrest.

Tumor suppressor REIC/DKK-3 and co-chaperone SGTA: Their interaction and roles in the androgen sensitivity.

REIC/DKK-3 is a tumor suppressor, however, its intracellular physiological functions and interacting molecules have not been fully clarified. Using yeast two-hybrid screening, we found that small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA), known as a negative modulator of cytoplasmic androgen receptor (AR) signaling, is a novel interacting partner of REIC/DKK-3. Mammalian two-hybrid and pull-down assay results indicated that the SGTA-REIC/DKK-3 interaction involved the N-terminal regions of both REIC/DKK-3 and SGTA and that REIC/DKK-3 interfered with the dimerization of SGTA, which is a component of the AR complex and a suppressor of dynein motor-dependent AR transport and signaling. A reporter assay in human prostate cancer cells that displayed suppressed AR signaling by SGTA showed recovery of AR signaling by REIC/DKK-3 expression. Considering these results and our previous data that REIC/DKK-3 interacts with the dynein light chain TCTEX-1, we propose that the REIC/DKK-3 protein interferes with SGTA dimerization, promotes dynein-dependent AR transport and then upregulates AR signaling.

Molecular cloning of canine co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA) and investigation of its ability to suppress androgen receptor signalling in androgen-independent prostate cancer.

Although the morbidity of canine prostate cancer is low, the majority of cases present with resistance to androgen therapy and poor clinical outcomes. These pathological conditions are similar to the signs of the terminal stage of human androgen-independent prostate cancer. The co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA) is known to be overexpressed in human androgen-independent prostate cancer. However, there is little information about the structure and function of canine SGTA. In this study, canine SGTA was cloned and analysed for its ability to suppress androgen receptor signalling. The full-length open reading frame (ORF) of the canine SGTA gene was amplified by RT-PCR using primers designed from canine-expressed sequence tags that were homologous to human SGTA. The canine SGTA ORF has high homology with the corresponding human (89%) and mouse (81%) sequences. SGTA dimerisation region and tetratricopeptide repeat (TPR) domains are conserved across the three species. The ability of canine SGTA to undergo homodimerisation was demonstrated by a mammalian two-hybrid system and a pull-down assay. The negative impact of canine SGTA on androgen receptor (AR) signalling was demonstrated using a reporter assay in androgen-independent human prostate cancer cell lines. Pathological analysis showed overexpression of SGTA in canine prostate cancer, but not in hyperplasia. A reporter assay in prostate cells demonstrated suppression of AR signalling by canine SGTA. Altogether, these results suggest that canine SGTA may play an important role in the acquisition of androgen independence by canine prostate cancer cells.

Structural and Functional Insights into Small, Glutamine-Rich, Tetratricopeptide Repeat Protein Alpha.

The small glutamine-rich, tetratricopeptide repeat-containing protein alpha (SGTA) is an emerging player in the quality control of secretory and membrane proteins mislocalized to the cytosol, with established roles in tail-anchored (TA) membrane protein biogenesis. SGTA consists of three structural domains with individual functions, an N-terminal dimerization domain that assists protein sorting pathways, a central tetratricopeptide repeat (TPR) domain that mediates interactions with heat-shock proteins, proteasomal, and hormonal receptors, and viral proteins, and a C-terminal glutamine rich region that binds hydrophobic substrates. SGTA has been linked to viral lifecycles and hormone receptor signaling, with implications in the pathogenesis of various disease states. Thus far, a range of biophysical techniques have been employed to characterize SGTA structure in some detail, and to investigate its interactions with binding partners in different biological contexts. A complete description of SGTA structure, together with further investigation into its function as a co-chaperone involved quality control, could provide us with useful insights into its role in maintaining cellular proteostasis, and broaden our understanding of mechanisms underlying associated pathologies. This review describes how some structural features of SGTA have been elucidated, and what this has uncovered about its cellular functions. A brief background on the structure and function of SGTA is given, highlighting its importance to biomedicine and related fields. The current level of knowledge and what remains to be understood about the structure and function of SGTA is summarized, discussing the potential direction of future research.

Expression of small glutamine-rich TPR-containing protein A (SGTA) in Non-Hodgkin's Lymphomas promotes tumor proliferation and reverses cell adhesion-mediated drug resistance (CAM-DR).

The expression and biologic function of SGTA in Non-Hodgkin's Lymphomas (NHL) was investigated in this study. Clinically, by immunohistochemistry analysis we detected SGTA expression in both reactive lymphoid tissues and NHL tissues. In addition, we also correlated high expression of SGTA with poor prognosis. Functionally, SGTA expression was positively related with cell proliferation and negative related with cell adhesion. Finally, SGTA knockdown induced adhesion-mediated drug resistance. Our finding supports a role of SGTA in NHL cell proliferation, adhesion and drug resistance, and it may pave the way for a novel therapeutic approach for CAM-DR in NHL.