Construction and Functional Analysis of beclin1 promoter luciferase reporter plasmid

DOI

10.2991/isaeece-17.2017.51How to use a DOI?

Keywords

autophagy, beclin1, E2F1, promoter activity, luciferase assay

Abstract

Autophagy is a conserved process in eukaryotic cells, which is induced by extracellular and intracellular signals, degrading biological macromolecules and damaged organelles through fusion with lysosomes. Beclin1, which is homologous to the yeast autophagy related gene Atg6, acts as a key gene for autophagy. It combines with the class III phosphatidylinositol 3-kinase (PtdIns3K) and Vps34 to form the key complex in the induction of autophagy. E2Fs transcription factors are known to be involved in the regulation of cell cycle progression. By analyzing the promoter sequence of beclin1, we found that there were two E2F1 binding sites within the promoter. In the present study, the gene fragment of human beclin1 gene promoter containing E2F1 was cloned into empty vector pGL3-Basic to construct beclin1 promoter luciferase reporter plasmid. E2F1 expression plasmids, together with beclin1 promoter-luc plasmids were transfected into COS-7 cells, and then luciferase assay was performed to analyze the effects of E2F1 on regulating the promoter activity of beclin1. The results showed that E2F1 can activate the transcription of beclin1. Constuction of beclin1 promoter luciferase reporter plasmid will provide the theory basis for investigating the function of beclin1 in autophagy.

Copyright

© 2017, the Authors. Published by Atlantis Press.

Open Access

This is an open access article distributed under the CC BY-NC license (http://creativecommons.org/licenses/by-nc/4.0/).

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TY  - CONF
AU  - Doudou Li
AU  - Na Huang
AU  - Xiaomei Li
AU  - Hao Zhou
AU  - Nan Wang
PY  - 2017/03
DA  - 2017/03
TI  - Construction and Functional Analysis of beclin1 promoter luciferase reporter plasmid
BT  - Proceedings of the 2017 2nd International Symposium on Advances in Electrical, Electronics and Computer Engineering (ISAEECE 2017)
PB  - Atlantis Press
SP  - 266
EP  - 269
SN  - 2352-5401
UR  - https://doi.org/10.2991/isaeece-17.2017.51
DO  - 10.2991/isaeece-17.2017.51
ID  - Li2017/03
ER  -