| Colon cancer is one of the most common malignant tumors in the world.The latest data for 2018 show that the incidence of colon cancer ranks third in the world and the mortality rate of colon cancer ranks second.The traditional treatment of colon cancer is surgery combined with post-operative chemotherapy,but it is prone to recurrence and toxic side effects of chemotherapy drugs,which is also the main cause of death of patients with colon cancer.With the research of colon cancer treatment,targeted therapy has become a new trend for colon cancer treatment because it does not damage normal cells.In recent years,effective targets for the treatment of colon cancer have been found continuously.Therefore,screening non-toxic and side-effects targeted drugs from natural products has become a research hotspot of scientists at home and abroad.Nostoc commune Vauch.is a nitrogen-fixing blue-green algae.Nostoc commune Vauch.is rich in nutrients,protein,lipids,vitamins and phosphorus,zinc,calcium and other minerals.As a delicacy,it has the functions of removing fire and supplementing nutrition.As a kind of medicine,it has the functions of lowering blood pressure and lipid,nourishing liver and kidney,and has curative effect on senile dementia.Previous studies showed that WSP1,a water stress protein extracted from Nostoc commune Vauch.,had a targeted anti-colon cancer effect in vivo and in vitro.Further,recombinant expression of Re-WSP1 by genetic engineering technology also had a significant anti-colon cancer effect.However,the molecular target of anti-colon cancer effect of Re-WSP1 and its active fragments are still unclear.Therefore,this study mainly explored the active fragments and molecular target of Re-WSP1 against colon cancer,and further elucidated the molecular mechanism of its anti-tumor effect,including the following three aspects:1.Effects of Re-WSP1 on Wnt/?-catenin signaling pathway in colon cancer cells.The expression of Wnt/?-catenin signaling pathway-relatedproteins in colon cancer cells were treated with Re-WSP1 were detected by Western blot.The results showed that the expression of ?-catenin,Dvl and cyclin D1 in colon cancer cells were decreased significantly after treatment with Re-WSP1.This suggests that Re-WSP1 can inhibit Wnt/?-catenin which is abnormally activated in colon cancer cells.2.Molecular targets of anti-colon cancer effects of Re-WSP1.Immunofluorescence showed that Re-WSP1 acted on the surface of colon cancer cells.Pull down and Western blot showed that there was an interaction between Re-WSP1 and FZD1.Antibody blocking showed that Re-WSP1 could significantly inhibit the growth of colon cancer cells,and the inhibition of colon cancer cells was reversed with FZD1 antibody treatment.These results suggests that FZD1 is the target protein of anti-colon cancer effects induced by Re-WSP1.3.Screening of anti-tumor activity fragments of Re-WSP1.The sequence and functional domains of Re-WSP1 were analyzed and predicted.The truncated WSP1-1,WSP1-2 and WSP1-3 of Re-WSP1 were constructed according to the predicted results for exogenous expression.The effects of three truncated fragments on the proliferation of colon cancer cells were detected by MTT.The results showed that the activity of WSP1-1 was significantly higher than other two truncated fragments.The apoptotic effects of WSP1-1 on colon cancer cells were further detected by flow cytometry.The results showed that there is the apoptotic effects of WSP1-1 on colon cancer cells.These results suggest that WSP1-1 is an active fragment of Re-WSP1 which plays an anti-tumor role.These studies suggest that Re-WSP1 exerts its anti-colon cancer effect by inhibiting Wnt/?-catenin signal pathway.Further research on the target,the study indicates that Re-WSP1 has the direct interaction with FZD1,a seven-time transmembrane protein receptor in Wnt/?-catenin signaling pathway.Re-WSP1 inhibits Wnt/?-catenin signaling pathway through targeting FZD1 and exerts its anti-colonic effect.Meantime,the study shows that truncated WSP1-1 is the active fragment of Re-WSP1 which plays ananti-colon cancer effect.This study provided a solid theoretical support for the treatment of colon cancer and effective biological targets of the water stress protein of Nostoc commune Vauch.. |
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