RESUMO
As an important second messenger, calcium (Ca2+) regulates a wide variety of physiological processes. Disturbance of intracellular calcium homeostasis implicated in the occurrence of multiple types of diseases. Orai1 is the major player in mediating store-operated calcium entry (SOCE) and regulates calcium homeostasis in non-excitable cells. Over-expression and activation of Orai1 have been reported in breast cancer. However, its molecular mechanisms are still not very clear. Here, we demonstrated that Nucleolin (NCL) was a novel interacting partner of Orai1. NCL is a multifunctional nucleocytoplasmic protein and is upregulated in human breast tumors. The binding of C-termini of NCL (NCL-CT) to N-termini of Orai1 (Orai1-NT) is critical for mediating calcium influx and proliferation of breast cancer cells. Blocking the NCL-Orai1 interaction by synthesized Orai1 peptide can effectively reduce the intracellular calcium influx and suppress the proliferation of breast cancer cells in vitro and in vivo. Our findings reveal a novel activation mechanism of Orai1 via direct interaction with NCL, which may lead to calcium homeostasis imbalance and promote the proliferation of breast cancer cells. Blocking NCL-Orai1 interaction might be an effective treatment of breast cancer.
RESUMO
Recent studies have suggested the potency of berberine (BBR) for multiple cancer treatments, including multiple myeloma (MM). However, the direct target and underlying mechanism of BBR remain largely understood in MM. Here, we demonstrated that BBR inhibited cell proliferation and acted synergistically with bortezomib in MM.1S cells. BBR treatment induced MM cell cycle arrest by downregulating several cell cycle-related proteins. Murine double minute 2 (MDM2) as a BBR-binding protein was identified by surface plasmon resonance image (SPRi) analysis and molecular docking. Overexpression of MDM2 is associated with MM progression and a poor prognosis. Knockdown MDM2 by siRNA transfection can repress MM malignant progression and attenuate the BBR sensitivity to MM.1S cells. BBR treatment induced the degradation of MDM2 through the ubiquitin-proteasome system and reactivated P53/P21 in MM cells. Overall, our data has illustrated that MDM2, as a binding protein of BBR for the first time, may serve as a potential therapeutic option for MM.
Assuntos
Berberina , Mieloma Múltiplo , Animais , Apoptose , Berberina/farmacologia , Berberina/uso terapêutico , Bortezomib/metabolismo , Carcinogênese , Linhagem Celular Tumoral , Humanos , Camundongos , Simulação de Acoplamento Molecular , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , RNA Interferente Pequeno , Proteína Supressora de Tumor p53/genética , UbiquitinaRESUMO
BACKGROUND: Current therapies for multiple myeloma (MM) are associated with toxicity and resistance, highlighting the need for novel effective therapeutics. Berberine (BBR), a botanical alkaloid derived from several Berberis medicinal plants, has exhibited anti-tumor effects, including against multiple myeloma (MM); however, the molecular mechanism underlying the anti-MM effect has not been previously described. This study aimed to identify the target of berberine and related mechanisms involved in its therapeutic activity against MM. RESULTS: Here, we demonstrated that BBR treatment killed MM cells in vitro and prolonged the survival of mice bearing MM xenografts in vivo. A screening approach integrating surface plasmon resonance (SPR) with liquid chromatography-tandem mass spectrometry (LC-MS/MS) identified UHRF1 (ubiquitin-like with PHD and RING Finger domains 1) as a potential target of BBR. Combining molecular docking and SPR analysis, we confirmed UHRF1 as a BBR-binding protein and discovered that BBR binds UHRF1 in the tandem tudor domain and plant homeodomain (TTD-PHD domain). BBR treatment induced UHRF1 degradation via the ubiquitin-dependent proteasome system and reactivated p16INK4A and p73 in MM cells. Overexpression of UHRF1 promoted the MM cell proliferation and rendered MM cells more resistant to BBR, while silencing of UHRF1 with siRNA attenuated BBR-induced cytotoxicity. CONCLUSIONS: In summary, our study has identified UHRF1 as a direct target of BBR and uncovered molecular mechanisms involved in the anti-MM activity of BBR. Targeting UHRF1 through BBR may be a novel therapeutic strategy against MM.
Assuntos
Anticarcinógenos/farmacologia , Berberina/farmacologia , Mieloma Múltiplo/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
Berberine (BBR), a traditional Chinese herbal medicine compound, has emerged as a novel class of anti-tumor agent. Our previous microRNA (miRNA) microarray demonstrated that miR-106b/25 was significantly down-regulated in BBR-treated multiple myeloma (MM) cells. Here, systematic integration showed that miR-106b/25 cluster is involved in multiple cancer-related signaling pathways and tumorigenesis. MiREnvironment database revealed that multiple environmental factors (drug, ionizing radiation, hypoxia) affected the miR-106b/25 cluster expression. By targeting the seed region in the miRNA, tiny anti-mir106b/25 cluster (t-anti-mir106b/25 cluster) significantly induced suppression in cell viability and colony formation. Western blot validated that t-anti-miR-106b/25 cluster effectively inhibited the expression of P38 MAPK and phospho-P38 MAPK in MM cells. These findings indicated the miR-106b/25 cluster functioned as oncogene and might provide a novel molecular insight into MM.
Assuntos
Berberina/farmacologia , MicroRNAs/genética , Mieloma Múltiplo/patologia , Transdução de Sinais , Linhagem Celular Tumoral , HumanosRESUMO
BACKGROUND: Berberine (BBR) is a natural alkaloid derived from a traditional Chinese herbal medicine. However, the exact mechanisms underlying the different effects of berberine on MM cells have not been fully elucidated. METHODS: A systematic analysis assay integrated common signaling pathways modulated by the 3 miRNA clusters and mRNAs in MM cells after BBR treatment. The role of the mir-99a â¼ 125b cluster, an important oncomir in MM, was identified by comparing the effects of t-anti-mirs with complete complementary antisense locked nucleic acids (LNAs) against mature mir-125b (anti-mir-125b). RESULTS: Three miRNAs clusters (miR-99a â¼ 125b, miR-17 â¼ 92 and miR-106 â¼ 25) were significantly down-regulated in BBR-treated MM cells and are involved in multiple cancer-related signaling pathways. Furthermore, the top 5 differentially regulated genes, RAC1, NFκB1, MYC, JUN and CCND1 might play key roles in the progression of MM. Systematic integration revealed that 3 common signaling pathways (TP53, Erb and MAPK) link the 3 miRNA clusters and the 5 key mRNAs. Meanwhile, both BBR and seed-targeting t-anti-mir-99a â¼ 125b cluster LNAs significantly induced apoptosis, G2-phase cell cycle arrest and colony inhibition. CONCLUSIONS: our results suggest that BBR suppresses multiple myeloma cells, partly by down-regulating the 3 miRNA clusters and many mRNAs, possibly through TP53, Erb and MAPK signaling pathways. The mir-99a â¼ 125b cluster might be a novel target for MM treatment. These findings provide new mechanistic insight into the anticancer effects of certain traditional Chinese herbal medicine compounds.