The Role Of P53^N236S Mutation In Regulating Senescence Induced By Telomere Dysfunction

Aging and cancer are important problems in the study of human diseases.Both of them are all associated with the cumulative damage of cells and regulation of DNA damage response genes.p53 as an important tumor suppressor gene,its mutantion not only lead to loss of function of the inhibition of cancer,some types of p53 mutations may also be related to the occurrence of certain cancers,such as the acute myelogenous leukemia.In addition,the mutant p53 may acquired the function of the inhibition of cancer,which can be described as gain-of-function(GOF)of mutp53.Many studies suggested that missense mutation of TP53 genes were found in about 50%human cancers.Same as p53,telomerase is also play an important role in the development of cancer and the aging process.The mutp53 and re-activated telomerase are also detected in many human tumors.When telomere dysfunction,the normal p53 can activate the emergency DNA damage response,then cells enter senescence or apoptosis program.However,the loss of function of mutp53 may cause cells escape from senescence or apoptosis.The survival rate and probability of occurrence of chromosomal rearrangements and other abnormal events of cells will be increased,which can lead to tumorigenesis.Therefore,the research of the p53’s regulatory role of telomere dysfunction is very important.It is an important significance that uncover the regulatory role and mechanism of mutp53 in aning and tumor process induced by telomere dysfunction.Then we can shed light on the artificial modulation of aging and finding safe anti-aging strategies.Werner Syndrome mouse model is produced by wrn and telomerase gene double knockout.It faithfully displays huaman WS symptoms.In our preliminary work,we found an p53 mutation(p53N239S)in three independent ALT tumorigenic cell lines derived from senesced Werner Syndrome MEFs,which escaped from senscence in long time culturing and become immortalized cell colonies.During the study of the characteristic of p53S,we have observed that p53S lost transcriptional regulatory function in both cell cycle arrest and apoptotic pathways,and gained new function in promoting tumorigenesis in vivo when cooperates with oncogenic Ras.Then we successfully established the p53N236S gene knock-in mouse model,in which lymphomas(56%)and sarcomas(26%)can be seen.And sarcomas that developed in p53S knock-in mice is frequently metastasized.It suggests that,p53S is an mutation which has potential of driving the cancer.Existing research shows that the p53 played a negative regulation role in cell cycle of hematopoietic stem cells.The amount of hematopoietic stem cells in p53 null mice remarkablely increased.And p53 mutations are more likely to cause of the instability of genome of hematopoietic stem cells,which causing the occurrence of leukemia and other tumors.p53 mutations are detected in about 13%human cancers.The abnomal of p53 is closely related to the development and prognosis of leukemia.Firstly,we study the hematopoietic phenotype of p53S mice.To explore whether it would cause hematopoietic system disease under the condition of the lymphatic System has affected and research on the regulation of p535 in WS background.We want to know the relationship between p53S mutation and telomere abnormality.We are also seeking a new theory of cancer prevention and control or anti-aging in science.First of all,We analysis the community distribution of p53s/s in mice bone marrow hematopoietic stem cells by mouse bone marrow cells LSK(Lin-sca+c kit+)analysis,and compared the results with wild type mice.We found that the ratio of p53S mice bone marrow hematopoietic stem cells was increased sharply compared with the wild type mice.It suggests that p53S might have lost the negative regulation of hematopoietic stem cells cell cycle from GO to G1 phase.In order to further study the regulation function of p53S in WS mice,we crossed breeding the WS mice and p53S mice,and gained the mTR-/-Wrn-/-p53S mice.And then to research the regulation function of p53S in premature aging phenotype and DNA damage stress related pathways and metabolism regulation related pathways in WS mice.We use the immunofluorescence,Real-time PCR and Western Blot to detect the DNA damage,gene and protein expression level of p53,p21 and puma in mTR-/－Wrn-/-p53S MEF cells.We found that DNA damage in G1mTR-/-Wrn-/-p53s/s is more serious than wild type,G1mTR-/-Wrn-/-,G3mTR-/-Wrn-/-.Real-time PCR and western blot results show that the classic tumor suppressor gene p53 in DNA damage pathway is activated,but almost lost its regulation function of p21 and puma expression.Cell proliferation data also show that the proliferation rate of GlmTR-/-Wrn-/-p53s/s MEF cells is significantly faster than G1mTR-/-Wrn-/-MEF cells.This suggests that p53S has lost the regulation function of cell cycle arrest and apoptosis,and disturbed the original cell cycle arrest,reversed the aging of WS cells.We also found that p53S play an important role on SIRT-P53-PGC1-a pathway.The expression of sirtl was decreased,while the expression of PGC1-α was increased.The mTR-/-Wrn-/-p53S MEF cells have lower levels of ROS than wild type cells and WS cells.This shows that the model might be a reversal of the aging of cells through the regulation of sirt1-PGC1-α pathway,and maintain a good mitochondrial function.In summary,our study found that the p53S could make mice hematopoietic stem cells increased.It provided a theoretical basis to further study whether p53S mice will suffer from hematopoietic system disease.On the mTR-/-Wrn-/-p53S MEF cells level,we preliminary study the related molecular mechanism regulation of p53S in WS MEF cells.We found that p53S can reverse the aging of WS cells,perhaps even cause the phenomenon of immortalized in the cell.This conclusion provided a preliminary theory to study the relationship between aging and tumor.