Nlp Deficiency Promotes Tumorigenesis In Mice And Acts As A Regulator In Vesicle Trafficking

Centrosome associated protein Nip is an essential regulator in cell cycle and mitosis.It is essential for centrosome maturation,microtubule development,and cytokinesis.During mitosis,Nip plays a critical role in spindle assembling,metaphase/anaphase and anaphase/telophase transition.Aberrant expression of Nlp often leads to mitosis abnormality and cell cycle disorders,resulting in abnormal separation of chromosomes and aneuploidy,leading to chromosomal instability and tumorigenesis.Overexpression of Nlp is closely associated with cancers.It is reported that Nip is overexpressed in numerous carcinomas,including lung cancer,breast cancer,oophoroma,head and neck neoplasm,renal carcinoma and thyroid cancer.It was also found to be gene amplified in lung cancer significantly.On the other hand,low expression of Nip may also result in aneuploidy and Nip is lowly expressed in human lymphomas.Transgenic mice overexpressing human-derived Nlp spontaneously developed tumors in breast,ovary and testis.It was also more sensitive to chemical DMBA and physical IR induced tumorigenesis.Moreover,MEFs of Nlp transgenic mice exhibited accelerated cell growth and apoptosis resistence,and showed centrosome amplification.The function of Nip in mitosis and its oncogenic role when overexpressed have been illuminated well.However,tumorigenesis of Nlp deficiency and the role of Nip in processes beyond cell cycle regulation still need more exploration.To further understand the causality between physiological functions of Nip and tumorigenesis,we constructed Nlp deficient mice.In this study,we found that the Nlp deficient mice were more prone to lymphoma spontaneously.The lymphoma cases were examined by IHC following paraffin embedding and section,and results showed the lymphomas were B-cell derived.Further,the hematological study of Nlp-deficient mice showed a reduced reconstitution ability especially for B-cells,and a decrease in the proportion of pre-B cells in the bone marrow and proportion of B-cells in peripheral blood.But no interference on HSCs was found.Then we examined multiple cytokine secretion in bone marrow and peripheral blood of Nlp deficient mice using ELISA kits.We found that IL-13,IL-17 and IL-21 secretion in bone marrow of Nlp-/-deficient mice were significantly decreased compare with control wild type mice,suggesting that apart from causing chromosome instability,Nlp might also have an effect on B-cell development via regulating immune cytokine secretion,which contributed to development of specific B cell lymphoma spontaneously.On the other hand,to further understand chemically induced tumorigenesis of Nlp deficient mice,we constructed DMBA-induced tumorigenesis model by intraperitoneal injection of a classical chemical carcinogen-DMBA.Based on data from a total of 234 DMBA-injection and control mice,we found that DMBA-induced mice exhibited various carcinomas including hepatoma and oophoroma.Nlp deficient mice showed higher incidence and malignancy of hepatoma.The liver mass,liver ratio,node No.of hepatoma and max node volume of hepatoma of Nlp deficient mice were all higher than those of wild type mice.Conclusively,the study indicates that Nlp deficient mice are more sensitive to DMBA carcinogen.In order to find new functions of Nlp,according to published evidences and previous research of our lab,we studied function of Nlp in intracellular vesicle traffic system.Vesicle traffic system is a network that is used to transport intracellular molecule cargo.Endosomes contain proteins and other molecules are sorted and transported by vesicle transport system to accomplish protein molecules processing,degradation,endocytosis and exocytosis and other biological processes.It is not only necessary for intracellular material and information exchange network,but also essential for various processes including autophagy,immune response and cytokine secretion.In the research,we found that Nlp was colocalized with late-endosome marker LAMP2 via immunofluorescence,suggesting that Nlp might be correlated with late-endosome.We emphasized on interaction of Nlp and late-endosome sorting protein Rab7 to explore function of Nlp in regulating late-endosome.Results suggested that Nlp promotes Rab7 interacting with one of its downstream effector,FYCO1,thereby endosome labeled by Rab7 is driven to move along microtubule towards plus end.Co-immunoprecipitation had confirmed that Nlp interacted with Rab7 and FYCO1,more than that,overexpression of Nlp facilitated interaction of Rab7 and FYCO1.Results showed above claimed that Nlp is a newly defined regulator of late-endosome sorting.Nlp is found to interact with Rab7 and regulate its functions to effect late-endosome sorting for the first time.This research have found Nlp deficiency promotes tumorigenesis and discussed the potential mechanisms,which helps to understand animal models of centrosome protein.On the other hand,Nlp is found to be a newly defined vesicle traffic regulator,which brings new sight to understand centrosome and vesicle traffic.The discovery is essential to complete the blueprint of intracellular exchange network.