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Knockout of the Placenta Specific 8 Gene Affects the Proliferation and Migration of Human Embryonic Kidney 293T Cell

  • Xu-Hui Qin
  • Hai-Xia Wang
  • Liqun Ma
  • Jinhua Shen
  • Qing-Hua Liu
  • Lu XueEmail author
Original Paper
  • 65 Downloads

Abstract

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.

Keywords

PLAC8 Cell proliferation Cell motility Cell cycle-related proteins E-cadherin 

Abbreviations

ccRCC

clear cell renal cell carcinoma

CDC

cell division cycle

CDK

cyclin-dependent kinase

EMT

epithelial-mesenchymal transition

FACS

fluorescence activated cell sorting

FITC

fluorescein isothiocyanate

gRNA

guide RNA

HEK293T

embryonic kidney cell line

KC

kidney cancer

KO

knockout

NC

nitrocellulose

PBS

phosphate-buffered saline

PI

propidium iodide

PLAC8

Placenta specific 8

WT

wildtype

Notes

Acknowledgements

The authors thank all the colleagues in Institute for Medical Biology for their technical support.

Funding

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.

Authors’ Contributions

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.

Supplementary material

12013_2019_893_MOESM1_ESM.pdf (202 kb)
Supplementary Figure S1
12013_2019_893_MOESM2_ESM.pdf (248 kb)
Supplementary Figure S2
12013_2019_893_MOESM3_ESM.docx (13 kb)
Supplementary Figures Legends

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Institute for Medical Biology and Hubei Provincial Key Laboratory for Protection and Application of Special Plants in Wuling Area of China, College of Life SciencesSouth-Central University for NationalitiesWuhanChina

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