Knockout of the Placenta Specific 8 Gene Affects the Proliferation and Migration of Human Embryonic Kidney 293T Cell
- 65 Downloads
Candidate oncogene placenta specific 8 (PLAC8) has been identified to participate in different cellular process and human diseases. However, the effects of PLAC8 on cell proliferation and migration in human kidney cancer (KC) remained unclear. In current study, physiological effects of PLAC8 in immortalized human embryonic kidney cell line (HEK293T) were investigated in vitro. Two PLAC8 knockout (KO) cell lines were established via CRISPR/Cas9-mediated methods combined with fluorescence activated single cell sorting. To classify the characteristic of PLAC8 during cell proliferation and migration in HEK293T, cellular proliferative activity was analyzed by cell counting and colony formation assay. Cell cycle distribution was analyzed by flow cytometry. Cellular motile activity was analyzed by wound-healing and migration assay. Further underlying molecular mechanism was explored via western blot. With the KO cell lines, it was found that PLAC8 KO could decrease cell proliferation. Moreover, the inhibitory effects of PLAC8 KO on cell proliferation were associated with a G2/M arrest in cell cycle progression concomitant with a remarkable inhibition of Cyclin B1 and elevation of Cyclin A. The alteration of cell cycle proteins and E-cadherin might further associate with the enhancement of cell motility. Our study revealed a novel role for PLAC8 in cell proliferation and migration of HEK293T cells, which might shed light on further study of PLAC8 on human KC.
KeywordsPLAC8 Cell proliferation Cell motility Cell cycle-related proteins E-cadherin
clear cell renal cell carcinoma
cell division cycle
fluorescence activated cell sorting
embryonic kidney cell line
Placenta specific 8
The authors thank all the colleagues in Institute for Medical Biology for their technical support.
This project was supported by Fund for Key Laboratory Construction of Hubei Province (Grant No. 2018BFC360), the Natural Science Foundation of Hubei Province, China (Grant No. 2018CFB594 to LX), “the Fundamental Research Funds for the Central Universities”, South-Central University for Nationalities (Grant No. CZD19003 to LX) and the China Scholarship Council (Grant No. 201808420069 to LX). The funding body had no role in the design of the study and collection, analysis, and interpretation of data or in writing the manuscript.
J.S., Q.H.L. and L.X. conceived and designed the experiments. X.H.Q., H.X.W. and L.M. performed the experiments. X.H.Q. and L.X. analyzed the data and generated the figures. L.X. wrote the manuscript. All authors gave final approval for the submitted version.
Compliance with Ethical Standards
Conflict of Interest
The authors declare that they have no conflict of interest.
- 5.Balakrishnan, V., et al. (2015). Plac8 links oncogenic mutations to regulation of autophagy and is critical to pancreatic cancer progression. Cancer Research, 75, 1143–1155.Google Scholar
- 14.Ouyang, C., et al. (2018). Placenta-specific 9, a putative secretory protein, induces G2/M arrest and inhibits the proliferation of human embryonic hepatic cells. Bioscience Reports, 38, BSR20180820.Google Scholar
- 24.Agarwal, M. L., Agarwal, A., Taylor, W. R., & Stark, G. R. (1995). p53 controls both the G2/M and the G1 cell cycle checkpoints and mediates reversible growth arrest in human fibroblasts. Proceedings of the National Academy of Sciences of the United States of America, 92, 8493–8497.CrossRefGoogle Scholar
- 37.Dai, M., et al. (2017). Erratum to: Cyclin D1 cooperates with p21 to regulate TGFbeta-mediated breast cancer cell migration and tumor local invasion. Breast Cancer Research: BCR, 19, 43.Google Scholar