Serine-threonine Kinase Receptor-associated Protein Research Articles

STRAP upregulates antiviral innate immunity against PRV by targeting TBK1

Serine/threonine kinase receptor-associated protein (STRAP) serves as a scaffold protein and is engaged in a variety of cellular activities, although its importance in antiviral innate immunity is unknown. We discovered that STRAP works as an interferon (IFN)-inducible positive regulator, facilitating type I IFN signaling during pseudorabies virus infection. Mechanistically, STRAP interacts with TBK1 to activate type I IFN signaling. Both the CT and WD40 7 − 6 domains contribute to the function of STRAP. Furthermore, TBK1 competes with PRV-UL50 for binding to STRAP, and STRAP impedes the degradation of TBK1 mediated by PRV-UL50, thereby increasing the interaction between STRAP and TBK1. Overall, these findings reveal a previously unrecognized role for STRAP in innate antiviral immune responses during PRV infection. STRAP could be a potential therapeutic target for viral infectious diseases.

CircPCNXL2 promotes tumor growth and metastasis by interacting with STRAP to regulate ERK signaling in intrahepatic cholangiocarcinoma

Backgroundcircular RNAs (circRNAs) have been reported to exert important effects in the progression of numerous cancers. However, the functions of circRNAs in intrahepatic cholangiocarcinoma (ICC) are still unclear.MethodscircPCNXL2 (has_circ_0016956) were identified in paired ICC by circRNA microarray. Then, we assessed the biological functions of circPCNXL2 by CCK8, EdU, clone formation, transwell, wound healing assays, and xenograft models. RNA pull-down, mass spectrometry, and RNA immunoprecipitation (RIP) were applied to explore the interaction between cirrcPCNXL2 and serine-threonine kinase receptor-associated protein (STRAP). RNA pull-down, RIP and luciferase reporter assays were used to investigate the sponge functions of circPCNXL2. In the end, we explore the effects of circPCNXL2 and trametinib (a MEK1/2 inhibitor) in vivo.ResultscircPCNXL2 was upregulated in ICC tissues and cell lines, which promoted the proliferation and metastasis of ICC in vitro and in vivo. In terms of the mechanisms, circPCNXL2 could directly bind to STRAP and induce the interaction between STRAP and MEK1/2, resulting in the tumor promotion in ICC by activation of ERK/MAPK pathways. Besides, circPCNXL2 could regulate the expression of SRSF1 by sponging miR-766-3p and subsequently facilitated the growth of ICC. Finally, circPCNXL2 could partially inhibit the anti-tumor activity of trametinib in vivo.ConclusioncircPCNXL2 played a crucial role in the progression of ICC by interacting with STRAP to activate the ERK signaling pathway, as well as by modulating the miR-766-3p/SRSF1 axis. These findings suggest that circPCNXL2 may be a promising biomarker and therapeutic target for ICC.

A pan-cancer analysis of the oncogenic role of Golgi transport 1B in human tumors.

Owing to the aggressiveness and treatment-refractory nature of cancer, ideal candidates for early diagnosis and treatment are needed. Golgi transport 1B (GOLT1B) has been associated with cellular malignant behaviors and immune responses in colorectal and lung cancer, but a systematic pan-cancer analysis on GOLT1B has not been conducted. The expression status and clinical association of GOLT1B in The Cancer Genome Atlas (TCGA) were analyzed. Genetic and methylation alterations in GOLT1B were explored. The relationship between GOLT1B and immune cell infiltration was also investigated. Genes related to GOLT1B expression were selected and analyzed. GOLT1B was highly expressed in most tumors, and there was a positive correlation between GOLT1B expression and clinical pathological parameters. High expression levels of GOLT1B have been associated with poor prognosis of most cancers. Copy number amplification was the primary type of GOLT1B genetic alterations, which was related to the prognosis of pan-cancer cases. There were different levels of GOLT1B promoter methylation across cancer types. The methylation level of the probe cg07371838 and cg25816357 was closely associated with prognosis in diverse cancers. There was also a positive correlation between GOLT1B genetic alterations and CD4+ T lymphocytes, especially the Th2 subset, as well as between GOLT1B expression and the estimated infiltration value of cancer-associated fibroblasts. Serine/threonine kinase receptor-associated protein (STRAP), integrator complex subunit 13 (INTS13), and ethanolamine kinase 1 (ETNK1) were the most relevant genes for GOLT1B expression, and their interactions with GOLT1B were involved in regulating the transforming growth factor (TGF)-β receptor signaling pathway and epithelial-mesenchymal transition (EMT). This pan-cancer analysis provided a comprehensive understanding of the oncogenic role of GOLT1B, highlighting a potential mechanism whereby GOLT1B influences the tumor microenvironment, as well as cancer immunotherapy.

STRAP binds to and promotes the repair of N1-methyldeoxyadenosine in DNA

N1-methyladenosine (m1A) is an important RNA modification that functions in various biological processes by interacting with cellular proteins. However, the binding proteins of N1-methyldeoxyadenosine (1mdA) in DNA remain largely unknown. Herein, we employed a quantitative proteomics strategy to identify the potential binding proteins of 1mdA in human cells. Our results revealed that serine‑threonine kinase receptor-associated protein (STRAP) can bind to 1mdA-carrying DNA. We further demonstrated that STRAP participates in alkylating agent-induced DNA damage response and can promote the repair of 1mdA embedded in DNA. Moreover, we investigated the effects of STRAP on 1mdA-induced perturbation in transcription using a shuttle vector- and next-generation sequencing-based assay, and found that STRAP is involved in the transcriptional bypass of 1mdA in human cells. Together, our study revealed STRAP as a novel 1mdA-binding protein in human cells and provided new insight into the biological implications of STRAP and 1mdA modification in human diseases.

Serine-threonine Kinase Receptor-Associated Protein is a Critical Mediator of APC Mutation–Induced Intestinal Tumorigenesis Through a Feed-Forward Mechanism

Inactivation of the Apc gene is a critical early event in the development of sporadic colorectal cancer (CRC). Expression of serine-threonine kinase receptor-associated protein (STRAP) is elevated in CRCs and is associated with poor outcomes. We investigated the role of STRAP in Apc mutation-induced intestinal tumor initiation and progression. We generated Strap intestinal epithelial knockout mice (StrapΔIEC) by crossing mice containing floxed alleles of Strap (Strapfl/fl) with Villin-Cre mice. Then we generated ApcMin/+;Strapfl/fl;Vill-Cre (ApcMin/+;StrapΔIEC) mice for RNA-sequencing analyses to determine the mechanism of function of STRAP. We used human colon cancer cell lines (DLD1, SW480, and HT29) and human and mouse colon tumor-derived organoids for STRAP knockdown and knockout and overexpression experiments. Strap deficiency extended the average survival of ApcMin/+ mice by 80 days and decreased the formation of intestinal adenomas. Expression profiling revealed that the intestinal stem cell signature, the Wnt/β-catenin signaling, and the MEK/ERK pathway are down-regulated in Strap-deficient adenomas and intestinal organoids. Correlation studies suggest that these STRAP-associated oncogenic signatures are conserved across murine and human colon cancer. STRAP associates with MEK1/2, promotes binding between MEK1/2 and ERK1/2, and subsequently induces the phosphorylation of ERK1/2. STRAP activated Wnt/β-catenin signaling through MEK/ERK-induced phosphorylation of LRP6. STRAP was identified as a target of mutated Apc and Wnt/β-catenin signaling as chromatin immunoprecipitation and luciferase assays revealed putative binding sites of the β-catenin/TCF4 complex on the Strap promoter. STRAP is a target of, and is required in, Apc mutation/deletion-induced intestinal tumorigenesis through a novel feed-forward STRAP/MEK-ERK/Wnt-β-catenin/STRAP regulatory axis.

FTO downregulation mediated by hypoxia facilitates colorectal cancer metastasis

Fat mass and obesity-associated protein (FTO), an N6-methyladenosine (m6A) demethylase, participates in tumor progression and metastasis in many malignancies, but its role in colorectal cancer (CRC) is still unclear. Here, we found that FTO protein levels, but not RNA levels, were downregulated in CRC tissues. Reduced FTO protein expression was correlated with a high recurrence rate and poor prognosis in resectable CRC patients. Moreover, we demonstrated that hypoxia restrained FTO protein expression, mainly due to an increase in ubiquitin-mediated protein degradation. The serine/threonine kinase receptor associated protein (STRAP) might served as the E3 ligase and K216 was the major ubiquitination site responsible for hypoxia-induced FTO degradation. FTO inhibited CRC metastasis both in vitro and in vivo. Mechanistically, FTO exerted a tumor suppressive role by inhibiting metastasis-associated protein 1 (MTA1) expression in an m6A-dependent manner. Methylated MTA1 transcripts were recognized by an m6A “reader”, insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2), which then stabilized its mRNA. Together, our findings highlight the critical role of FTO in CRC metastasis and reveal a novel epigenetic mechanism by which the hypoxic tumor microenvironment promotes CRC metastasis.

Serine-Threonine Kinase Receptor-Associated Protein (STRAP) Knockout Decreases the Malignant Phenotype in Neuroblastoma Cell Lines.

Simple SummaryNeuroblastoma is the most common extra-cranial tumor in children and despite medical advancements in cancer treatment, five-year survival for high-risk neuroblastoma remains less than 50%. Investigation of the mechanisms responsible for aggressive disease is necessary to identify novel therapeutic targets and improve survival. Serine-threonine kinase receptor associated protein (STRAP) is upregulated in several malignancies and plays an important role in tumor growth and metastasis. The role of STRAP in pediatric malignancies and specifically in neuroblastoma has not been explored. We sought to determine whether STRAP functions assisted to promote the malignant phenotype in neuroblastoma and could provide a potential target for future therapies.Background: Serine-threonine kinase receptor-associated protein (STRAP) plays an important role in neural development but also in tumor growth. Neuroblastoma, a tumor of neural crest origin, is the most common extracranial solid malignancy of childhood and it continues to carry a poor prognosis. The recent discovery of the role of STRAP in another pediatric solid tumor, osteosarcoma, and the known function of STRAP in neural development, led us to investigate the role of STRAP in neuroblastoma tumorigenesis. Methods: STRAP protein expression was abrogated in two human neuroblastoma cell lines, SK-N-AS and SK-N-BE(2), using transient knockdown with siRNA, stable knockdown with shRNA lentiviral transfection, and CRISPR-Cas9 genetic knockout. STRAP knockdown and knockout cells were examined for phenotypic alterations in vitro and tumor growth in vivo. Results: Cell proliferation, motility, and growth were significantly decreased in STRAP knockout compared to wild-type cells. Indicators of stemness, including mRNA abundance of common stem cell markers Oct4, Nanog, and Nestin, the percentage of cells expressing CD133 on their surface, and the ability to form tumorspheres were significantly decreased in the STRAP KO cells. In vivo, STRAP knockout cells formed tumors less readily than wild-type tumor cells. Conclusion: These novel findings demonstrated that STRAP plays a role in tumorigenesis and maintenance of neuroblastoma stemness.

STRAP regulates alternative splicing fidelity during lineage commitment of mouse embryonic stem cells

Alternative splicing (AS) is involved in cell fate decisions and embryonic development. However, regulation of these processes is poorly understood. Here, we have identified the serine threonine kinase receptor-associated protein (STRAP) as a putative spliceosome-associated factor. Upon Strap deletion, there are numerous AS events observed in mouse embryoid bodies (EBs) undergoing a neuroectoderm-like state. Global mapping of STRAP-RNA binding in mouse embryos by enhanced-CLIP sequencing (eCLIP-seq) reveals that STRAP preferably targets transcripts for nervous system development and regulates AS through preferred binding positions, as demonstrated for two neuronal-specific genes, Nnat and Mark3. We have found that STRAP involves in the assembly of 17S U2 snRNP proteins. Moreover, in Xenopus, loss of Strap leads to impeded lineage differentiation in embryos, delayed neural tube closure, and altered exon skipping. Collectively, our findings reveal a previously unknown function of STRAP in mediating the splicing networks of lineage commitment, alteration of which may be involved in early embryonic lethality in mice.

Mixed Lineage Leukemia 1 Promoted Neuron Apoptosis in Ischemic Penumbra via Regulating ASK-1/TNF-α Complex.

Neuron apoptosis in ischemic penumbra was proved to be involved in ischemic stroke (IS) development and contributed to the poor prognosis of IS. Recent studies showed that aberrant trimethylation of histone H3 lysine 4 (H3K4me3) level was associated with cell apoptosis. This study aimed to explore the underlying mechanism of neuron apoptosis in ischemic penumbra via histone methyltransferase (HMT) mixed lineage leukemia 1 (MLL1) mediated epigenetic pathway. Mouse IS model was established by middle cerebral artery occlusion (MCAO). Mouse primary cortical mixed cells were cultured and treated with oxygen–glucose deprivation (OGD) to simulate IS process. The expressions of apoptosis signal regulating kinase-1 (ASK-1), pASK-1, cleaved caspase-3, ASK-1/serine–threonine kinase receptor-associated protein (STRAP)/14-3-3 complex, ASK-1/tumor necrosis factor-α (TNF-α) complex, and MLL1 in mouse brain tissue and mouse primary cortical mixed cells were analyzed. The function of MLL1 was investigated using small interfering RNA (siRNA) targeting MLL1 and vector overexpressing MLL1. In vivo inhibition of MLL1 was conducted to explore its value as a therapeutic target. The prognostic value of MLL1 was investigated in IS patients. Results showed that the expressions of ASK-1, pASK-1, cleaved caspase-3, ASK-1/TNF-α complex, and MLL1 increased significantly in ischemic penumbra compared to brain tissue from the control group (P < 0.05). MCAO and OGD significantly upregulated the H3K4me3 level in ASK-1 promoter region and promoted the recruitment of MLL1 to this region (P < 0.05). siMLL1 significantly reversed the proapoptosis effects of OGD in primary cortical mixed cells, while MLL1 overexpression induced apoptosis of cells (P < 0.05). In vivo inhibition of MLL1 significantly reduced the infarct volume and the neurological score of MCAO mice (P < 0.05). Serum MLL1 level had a positive association with that in ischemic core and penumbra in mouse model and was positively correlated with the infarct volume and neurological score (P < 0.05). Besides, serum MLL1 level was also significantly correlated with the severity of IS (P < 0.05), and high serum MLL1 level indicated poor prognosis of IS patients (P < 0.05). These results revealed that MLL1 contributed to neuron cell apoptosis in ischemic penumbra after IS onset by promoting the formation of ASK-1/TNF-α complex, and its serum level was associated with poor prognosis of IS.

MicroRNA-494 represses osteosarcoma development by modulating ASK-1 related apoptosis complexes.

BackgroundMicroRNA (miR)-494 had been proven to be involved in tumor pathogenesis, while studies on miR-494 and osteosarcoma (OS) are still limited. In the current study, we aimed to explore the expression pattern and the roles of miR-494 in the development of OS.MethodsMouse models and osteoblastic cells were used to investigate the functions of miR-494. A case-control study including 87 OS patients and 100 controls was also conducted to investigate the prognostic value of miR-494.ResultsAnimal researches and human studies showed that miR-494 expression level in OS tissue (OST) was down-regulated compared to paracancerous tissue (PT), and had a significant positive association with its level in serum (P<0.05). Gain of miR-494 function suppressed OS tumor growth and promoted apoptosis of OS cells, while loss of miR-494 inhibited apoptosis and promoted invasion of osteoblastic cells. MiR-494 expressions in both OST and serum were correlated with the T stage, lymph node metastasis and distant metastasis of OS (P<0.05). Results of survival analysis showed that compared to patients with high miR-494 level, the overall survival rate in 24 months was significantly lower in the low miR-494 group (P<0.05). In vitro study revealed that miR-494 interacted with apoptosis signal regulating kinase 1 (ASK-1)/serine-threonine kinase receptor-associated protein (STRAP)/14-3-3 complex to promote the activation of tumor necrosis factor-α (TNF-α)/ASK-1-mediated apoptosis.ConclusionsThese results revealed that miR-494 functioned as a tumor suppressor and improved the prognosis of OS via modulating apoptosis complexes.

SIRT7 Deacetylates STRAP to Regulate p53 Activity and Stability.

Serine-threonine kinase receptor-associated protein (STRAP) functions as a regulator of both TGF-β and p53 signaling that participates in the regulation of cell proliferation and cell death in response to various stresses. Here, we demonstrate that STRAP acetylation plays an important role in p53-mediated cell cycle arrest and apoptosis. STRAP is acetylated at lysines 147, 148, and 156 by the acetyltransferases CREB-binding protein (CBP) and that the acetylation is reversed by the deacetylase sirtuin7 (SIRT7). Hypo- or hyperacetylation mutations of STRAP at lysines 147, 148, and 156 (3KR or 3KQ) influence its activation and stabilization of p53. Moreover, following 5-fluorouracil (5-FU) treatment, STRAP is mobilized from the cytoplasm to the nucleus and promotes STRAP acetylation. Our finding on the regulation of STRAP links p53 with SIRT7 influencing p53 activity and stability.

Integrated Analysis of the Gene Expression Changes During Colorectal Cancer Progression by Bioinformatic Methods.

We attempted to analyze the aberrant pathways and genes underlying the successive stages of colorectal cancer (CRC). The CRC related microarray data (GSE77953) were retrieved from Gene Expression Omnibus database, which included 17 colonic adenoma, 17 carcinoma, 11 CRC metastases, and 13 normal colonic epithelium samples. The differential expression patterns in colonic adenoma, carcinoma, and metastases were analyzed. Gene functional interaction (FI) and coexpressed network were constructed. Pathway enrichment analysis was performed to investigate the perturbed pathways, and disease-related genes were explored based on the Comparative Toxicogenomics Database. Total 438 genes were identified to be differentially expressed in colonic adenoma, 885 in carcinoma and 736 in metastases. The upregulated genes in adenoma were significantly related with ribosome, oxidative phosphorylation, and protein export related pathways. The downregulated genes in carcinoma and metastases were enriched in the same pathways, such as nitrogen metabolism, mineral absorption, and steroid hormone biosynthesis. FI network was constructed with 219 and 3914 edges, which were further divided to 12 modules. The genes in module 0 were closely related with ribosome, protein export, and RNA transport. Coexpressed genes were enriched in ribosome, protein export, and mineral absorption pathways. Total eight common upregulated genes were found to be the CRC-related genes such as RNF43 (ring finger protein 43), EIF3H (eukaryotic translation initiation factor 3 subunit H), and STRAP (serine/threonine kinase receptor associated protein). The common downregulated genes included ABCG2 (ATP binding cassette subfamily G member 2), GCG (glucagon), and SULT1A1 (sulfotransferase family 1A member 1). Oxidative phosphorylation, nitrogen metabolism, mineral absorption, and protein synthesis may significantly be perturbed in the progression of CRC. The overexpression of EIF3H may be the predictor for CRC formation.

Oncogenic STRAP Supports Hepatocellular Carcinoma Growth by Enhancing Wnt/β-Catenin Signaling.

Aberrant activation of Wnt/β-catenin signaling plays a key role in the onset and development of hepatocellular carcinomas (HCC), with about half of them acquiring mutations in either CTNNB1 or AXIN1. The serine/threonine kinase receptor-associated protein (STRAP), a scaffold protein, was recently shown to facilitate the aberrant activation of Wnt/β-catenin signaling in colorectal cancers. However, the function of STRAP in HCC remains completely unknown. Here, increased levels of STRAP were observed in human and mouse HCCs. RNA sequencing of STRAP knockout clones generated by gene editing of Huh6 and Huh7 HCC cells revealed a significant reduction in expression of various metabolic and cell-cycle-related transcripts, in line with their general slower growth observed during culture. Importantly, Wnt/β-catenin signaling was impaired in all STRAP knockout/down cell lines tested, regardless of the underlying CTNNB1 or AXIN1 mutation. In accordance with β-catenin's role in (cancer) stem cell maintenance, the expressions of various stem cell markers, such as AXIN2 and LGR5, were reduced and concomitantly differentiation-associated genes were increased. Together, these results show that the increased STRAP protein levels observed in HCC provide growth advantage among others by enhancing Wnt/β-catenin signaling. These observations also identify STRAP as a new player in regulating β-catenin signaling in hepatocellular cancers. IMPLICATIONS: Elevated STRAP levels in hepatocellular cancers provide a growth advantage by enhancing Wnt/β-catenin signaling.

Oncogenic roles of serine-threonine kinase receptor-associated protein (STRAP) in osteosarcoma.

To validate the presence of serine-threonine kinase receptor-associated Protein (STRAP) in osteosarcoma tissue and to investigate the oncological role of STRAP in osteosarcoma. Expression of STRAP protein in osteosarcoma tissue compared to soft callus (hyperactive bone healing tissue) and in multiple cell lines was examined using western blot analysis. Effects of STRAP silencing on cell proliferation, invasion, migration and re-implantability in chick chorioallantoic membrane (CAM) were observed in osteosarcoma cell lines (MNNG-HOS, 143B, and U2OS). The result demonstrated that STRAP was highly up-regulated in osteosarcoma tissues compared with the normal physiological bone healing tissue (soft callus). Expression level of STRAP was markedly high in osteosarcoma cell lines with aggressive phenotype. Upon STRAP silencing, invasion and migration, but not proliferative activity, were selectively modulated in high-expression-STRAP cell lines. In addition, STRAP silencing reduced the success rate of tumor implantation and growth of MNNG-HOS cells in CAM model. Serine-threonine kinase receptor-associated protein is up-regulated during osteosarcoma progression. The presence of STRAP enhances osteosarcoma cell invasion, migration and re-implantation ability, factors which play a critical role in metastasis. Serine-threonine kinase receptor-associated protein and its related pathway are worthy for further exploration as a novel target for anti-metastasis agents.

Serine Threonine Kinase Receptor-Associated Protein Deficiency Impairs Mouse Embryonic Stem Cells Lineage Commitment Through CYP26A1-Mediated Retinoic Acid Homeostasis.

Retinoic acid (RA) signaling is essential for the differentiation of embryonic stem cells (ESCs) and vertebrate development. RA biosynthesis and metabolism are controlled by a series of enzymes, but the molecular regulators of these enzymes remain largely obscure. In this study, we investigated the functional role of the WD-domain protein STRAP (serine threonine kinase receptor-associated protein) in the pluripotency and lineage commitment of murine ESCs. We generated Strap knockout (KO) mouse ESCs and subjected them to spontaneous differentiation. We observed that, despite the unchanged characteristics of ESCs, Strap KO ESCs exhibited defects for lineage differentiation. Signature gene expression analyses revealed that Strap deletion attenuated intracellular RA signaling in embryoid bodies (EBs), and exogenous RA significantly rescued this deficiency. Moreover, loss of Strap selectively induced Cyp26A1 expression in mouse EBs, suggesting a potential role of STRAP in RA signaling. Mechanistically, we identified putative Krüppel-like factor 9 (KLF9) binding motifs to be critical in the enhancement of non-canonical RA-induced transactivation of Cyp26A1. Increased KLF9 expression in the absence of STRAP is partially responsible for Cyp26A1 induction. Interestingly, STRAP knockdown in Xenopus embryos influenced anterior-posterior neural patterning and impaired the body axis and eye development during early Xenopus embryogenesis. Taken together, our study reveals an intrinsic role for STRAP in the regulation of RA signaling and provides new molecular insights for ESC fate determination. Stem Cells 2018;36:1368-1379.

Abstract 5058: STRAP-induced Wnt/ß-catenin signaling is important in inactivated Apc-mediated tumorigenicity and stemness of colorectal cancer

Abstract Aberrant Inactivation of the Apc gene and activation of Wnt/β-catenin signaling in colorectal cancer (CRC) has been implicated in tumorigenesis and homeostasis of cancer stem-like cells (CSCs), albeit the mechanism remains mostly obscure. We have previously reported the identification of a WD-domain protein, STRAP (Serine-Threonine Kinase Receptor-Associated Protein) that inhibits TGF-ß signaling, interacts with a variety of proteins, and influences a wide range of cellular processes. Upregulation of STRAP in CRC patients had a worse survival with adjuvant therapy. In contrast, patients carrying tumors with normal or low STRAP expression benefited from the treatment. Our preliminary data suggest that homozygous deletion of STRAP causes embryonic lethality at embryonic day E9.5. Therefore, to study the role of STRAP in vivo in CRC, we have generated Strap-conditional knockout mice Villin-Cre;Strapfl/fl(Strap-cko) and used two mouse models of CRC: Apc+/min and AOM/DSS. Conditional deletion of Strap in both mouse models resulted in a dramatic decrease in tumor formation. Our results demonstrate that STRAP is required for tumor formation through activation of Wnt/β-catenin signaling pathway in both models. Immunoblotting and immunohistochemistry analyses of tumors derived from the Strap-cko;Apc+/min mice showed decreased levels of β-catenin and of its nuclear staining. Furthermore, mRNA levels of several Wnt target genes are also decreased in tumors derived from Strap-cko;Apc+/min mice compared to those from Apc+/min mice. Formation and sustained propagation of intestinal enteroids require cells from the stem cell niche. APC mutation accompanied by elevated Wnt signaling promotes spheroid-shaped enteroids as indicated by the absence of crypt buds. We found that enteroids prepared from Strap-cko;Apc+/min mice exhibited more budding compared with the Apc+/min enteroids, indicating the role of STRAP in stemness. Strap deletion in enteroids decrease stem cell markers such as CD133, CD44, and LGR5. By using chromatin immunoprecipitation (ChIP) assay, we have found that STRAP knockdown can affect the recruitment of β-catenin/TCF4 complex on target promoter regions. Interestingly, STRAP protein and mRNA levels were found to be upregulated in Apc+/min induced tumors. STRAP is upregulated by Wnt/β-catenin signaling pathway, as evidenced by downregulation of STRAP in β-catenin/TCF4 inhibitor iCRT3 treated cells. In addition, ChIP assay has identified TCF DNA binding sites in the Strap promoter through which Wnt signaling regulates Strap gene expression. Altogether, our findings reveal that STRAP is a novel target of Wnt/β-catenin signaling and it is required for inactivated Apc- and Wnt/β-catenin-induced stemness and tumorigenesis through a complex feedforward loop. Citation Format: Trung Vu, Arunima Datta, Lin Jin, Pran K. Datta. STRAP-induced Wnt/ß-catenin signaling is important in inactivated Apc-mediated tumorigenicity and stemness of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5058.

Structure, molecular dynamics simulation, and docking studies of Dictyostelium discoideum and human STRAPs.

The Serine Threonine kinase Receptor Associated Protein (STRAP) is a WD40 containing protein that provides a platform for protein interactions during cell proliferation and development. Overexpression and misregulation of STRAP contributes to various carcinomas that are now recognized as therapeutic targets especially for colorectal and lung cancers. The present study was undertaken to find an effective drug against this molecule using a simple system like Dictyostelium discoideum; which shares close homology to humans. Using techniques like structural modeling, molecular dynamics (MD) simulation and molecular docking, we found similar structure and dynamic behaviors in both, except for the presence of dissimilar numbers of β-sheets and loop segments. We identified a novel and potential drug targeted to STRAP. The results obtained allow us to use Dictyostelium as a model system for further in vivo studies. Finally, the results of protein-protein interactions using molecular docking and essential dynamics studies show STRAP to participate in TGF-β signaling in humans. Further, we show some structural units that govern the interaction of TGFβ-RI with STRAP and Smad7 proteins in TGF-β signaling pathway. In conclusion, we propose that D. discoideum can be used for enhancing our knowledge about STRAP protein.

Sanguinarine triggers intrinsic apoptosis to suppress colorectal cancer growth through disassociation between STRAP and MELK

BackgroundPrevious studies showed sanguinarine induced apoptosis in CRC cells but did not define the underlying mechanisms. The purpose of this work was to determine the in vivo and in vitro effects of sanguinarine on CRC tumors and to elucidate the mechanism in regulating the intrinsic apoptosis.MethodsCell viability of CRC cell lines treated with sanguinarine was measured by MTT assay. Apoptotic cells stained with Annexin V and 7-AAD were detected by flow cytometry. Mitochondrial membrane potential and reactive oxygen species (ROS) were analyzed by JC-1 and DCFH-DA staining, respectively. The in vitro kinase activity of MELK was analyzed by using HTRF® KinEASE™-STK kit. The expression of proteins were determined using Western blotting and immunohistochemistry. Co-immunoprecipitation and immunofluorecence were used to study the interaction between STRAP and MELK. The anti-neoplastic effect of sanguinarine was observed in vivo in an orthotopic CRC model.ResultsSanguinarine decreased the tumor size in a dose-dependent manner in orthotopical colorectal carcinomas through intrinsic apoptosis pathway in BALB/c-nu mice. It significantly increased cleavage of caspase 3 and PARP in implanted colorectal tissues. Sanguinarine increased mitochondrial ROS and triggered mitochondrial outer membrane permeabilization in multiple colorectal cancer (CRC) cell lines. NAC pretreatment lowered ROS level and downregulated apoptosis induced by sanguinarine. The intrinsic apoptosis induced by sanguinarine was Bax-dependent. The elevated expression and association between serine-threonine kinase receptor-associated protein (STRAP) and maternal embryonic leucine zipper kinase (MELK) were observed in Bax positive cells but not in Bax negative cells. Sanguinarine dephosphorylated STRAP and MELK and disrupted the association between them in HCT116 and SW480 cells. The expression and association between STRAP and MELK were also attenuated by sanguinarine in the tumor tissues. Importantly, we found that STRAP and MELK were overexpressed and highly phosphorylated in colorectal adenocarcinomas and their expression were significantly correlated with tumor stages. Furthermore, the expression of MELK, but not STRAP, was associated with lymph node metastasis.ConclusionsSanguinarine dephosphorelates STRAP and MELK and disassociates the interaction between them to trigger intrinsic apoptosis. Overexpression of STRAP and MELK may be markers of CRC and their disassociation may be a determinant of therapeutic efficacy.

Progress of the role of serine-threonine kinase receptor-associated protein in tumorigenesis

Serine-threonine kinase receptor-associated protein (STRAP) is initially identified as a putative inhibitor of the canonical transforming growth factor-β (TGF-β) signaling pathway by Dr. Datta. Because the Smad-dependent TGF-β pathway negatively regulates cellular growth, and STRAP has been found overexpressed in various cancers, like colorectal, lung and breast carcinoma, early functional studies suggested that STRAP behaves as an oncogene. As the progress of new studies on the biological function of STRAP, it is becoming clear that STRAP has diversity function to regulate multiple signaling pathways. Due to the absence of enzymatic activity, it appears that STRAP influences multiple biological processes through direct associations with other cellular proteins within the β propeller of its WD40 domain. In this review, we will comprehensively describe the progress of the role of STRAP in tumorigenesis. Key words: Serine-threonine kinase receptor-associated protein; Transforming growth factor-β; β-catenin; Nonmetastatic gene 23-H1; Apoptosis signal-regulating kinase 1; Tumorigenesis

Dual Roles of Serine-Threonine Kinase Receptor-Associated Protein (STRAP) in Redox-Sensitive Signaling Pathways Related to Cancer Development.

Serine-threonine kinase receptor-associated protein (STRAP) is a transforming growth factor β (TGF-β) receptor-interacting protein that has been implicated in both cell proliferation and cell death in response to various stresses. However, the precise roles of STRAP in these cellular processes are still unclear. The mechanisms by which STRAP controls both cell proliferation and cell death are now beginning to be unraveled. In addition to its biological roles, this review also focuses on the dual functions of STRAP in cancers displaying redox dysregulation, where it can behave as a tumor suppressor or an oncogene (i.e., it can either inhibit or promote tumor formation), depending on the cellular context. Further studies are needed to define the functions of STRAP and the redox-sensitive intracellular signaling pathways that enhance either cell proliferation or cell death in human cancer tissues, which may help in the development of effective treatments for cancer.