| Pten?phosphate and tension homology deleted on chromsome ten?is located in the chromosome 10 and consists of 403 amino acids.It encodes a tumor suppressor protein PTEN with lipid phosphatase activity and protein phosphatase activity.Since cloned in1997 and demonstrated as a tumor suppressor gene,it has been a star molecule in the tumor research field.Mutations and deletions of Pten are found in a variety of tumor patient tissues.The importance of PTEN in tumor suppression is confirmed by the existence of germline mutations of PTEN in the PTEN hereditary tumor syndromes?PHTS?,including Cowden disease,Bannayan–Riley–Ruvalcaba syndrome.PTEN is a dual-specificity protein and lipid phosphatase,and its primary cellular substrate is the second messenger phosphatidylinositol?3,4,5?-trisphosphate?PIP3?,which hydrolyzes to phosphatidylinositol?4,5?-bisphosphate?PIP2?.PTEN blocks PI3K signaling by inhibiting PIP3-dependent processes,playing a tumor suppressor function.As a result,changes of the protein level and activity of PTEN could show significant on its tumor suppress function.In addition,PTEN also function to maimtain genomic stability,inhibit cell cycle progression and rerulate gene expression through a PI3K-independent way.PTEN is an important tumor suppressor,which has complicated regulation model.In order to study the effect of PTEN dose changes on its tumor suppressor function,a mouse model expressing different PTEN protein level has been constructed by Pier Paolo Pandolfi laboratory.The found when the protein level of PTEN was reduced to80%,the incidence of breast cancer was as high as 40%,and when the protein level of PTEN droped to 30%,the incidence of breast cancer climbed to 75%.In addition,their laboratory also constructed mice with elevated levels of PTEN protein?Super-PTEN mice?and the research showed the body weight and size of mice were dose-dependent on the PTEN protein level.The higher level of PTEN,the smaller the body size of mice and the stronger the anti-cancer ability.There are three levels of protein regulation:transcriptional regulation,post-transcriptional regulation,and post-translational regulation.Among them,post-translational regulation of proteins is an important way to maintain protein stability.Post-translational modification of PTEN can affect its phosphatase activity,protein conformation,subcellular localization,and protein stability,such as phosphorylation,acetylation,and ubiquitination.Our research focuses on the regulation of PTEN ubiquitilation and deubiquitilation.Ubiquitinated PTEN undergoes degradation via the ubiquitin-proteasome pathway,whereas deubiquitination antagonizes this process.The ubiquitin ligase and deubiquitinating enzyme are function as the two ends of the balance,involved in these two important processes can regulate the stability of PTEN protein.There are five ubiquitin ligases?WWP2,CHIP,XIAP,Nedd4-1 and TRIM27?and three deubiquitinating enzymes?USP7,USP13 and OTUD3?have been discovered to involve in PTEN regulation,suggesting the regulation of PTEN's protein stability is complex.Studies have shown that ubiquitination of PTEN promoted by TRIM27 only regulates its phosphatase activity,whereas does not affect its protein level and localization.Nedd4-1 could promote both mono-ubiquitination and poly-ubiquitination of PTEN,which promotes PTEN nuclear transport and proteasome degradation respectively.However,the ability of Nedd4-1 ubiquitinating PTEN was challenged by Nedd4-1 knockout mouse models.XIAP can regulate PTEN protein levels through poly-ubiquitination and promote the transfer of cytoplasmic PTEN to the nucleus through mono-ubiquitination in vivo.However,whether WWP2 and CHIP can remove PTEN polyubiquitination and regulate PTEN protein stability lacks systematic studies under physiological validation.In order to solve this problem,we used WWP2 and CHIP single knockout mouse model to conduct research.Our main findings are as follows:?1?By comparing and analyzing the phenotypes of WWP2 knockout mice and wild type mice,we found WWP2 knockout mice have phenotypes including elevation of PTEN level and smaller body size.?2?Analysis of WWP2 knockout mouse embryonic fibroblasts and various tissues revealed that ubiquitination level of PTEN was decreased,the stability of the protein was enhanced,and the AKT pathway was inhibited.?3?Analysis of CHIP knockout mice found that the birth rate of CHIP-deficient mice was very low,and the size of mice was significantly reduced.However,the level of ubiquitination and protein stability of PTEN in CHIP-deficient mice did not change significantly,we suggested that CHIP plays an important role in growth and development but does not modulate PTEN protein stability.Currently,only three deubiquitilases of PTEN have been reported.USP7 can remove mono-ubiquitination of PTEN and regulate the subcellular localization of PTEN protein.USP13 and OTUD3 can remove polyubiquitination of PTEN and improve the level of PTEN protein,thereby inhibiting the development of tumors.According to previous research in our laboratory,USP10 is another deubiquitinating enzyme of PTEN.Previous studies have confirmed that USP10 can remove ubiquitination of PTEN and stabilize PTEN,but this conclusion requires more evidence.Through further research,we found that depletion of USP10 through CRASPR/CAS9 technology in H1299 cells and MCF7 cells demonstrated that USP10 inhibit tumor cell proliferation and migration.Furthermore,through further texting the protein stability of PTEN and the sensitivity of AKT signaling in USP10+/+and USP10-/-MEFs,revealed the stability of PTEN by removing ubiquitination of PTEN in vivo.Our research involved the ubiquitination and deubiquitination of PTEN,an important tumor suppressor,and demonstrated that WWP2 is a major E3 ubiquitin ligase for PTEN in vivo.Therefore,WWP2 is more suitable than CHIP to target for tumor treatment.The current studies also revealed that USP10 is associated with the development of pancreatic cancer,esophageal cancer,and colorectal cancer.Our research might contribute to development of drugs for cancer treatment. |
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