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Biotechnol J. 2018 Feb;13(2). doi: 10.1002/biot.201700342. Epub 2017 Nov 14.

Identifying HIPK1 as Target of miR-22-3p Enhancing Recombinant Protein Production From HEK 293 Cell by Using Microarray and HTP siRNA Screen.

Author information

1
Biotechnology Core Laboratory NIDDK, NIH, Bethesda, Maryland 20892, USA.
2
Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland 21218, USA.
3
Chemical Genomics Center, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850.

Abstract

Protein expression from human embryonic kidney cells (HEK 293) is an important tool for structural and clinical studies. It is previously shown that microRNAs (small, noncoding RNAs) are effective means for improved protein expression from these cells, and by conducting a high-throughput screening of the human microRNA library, several microRNAs are identified as potential candidates for improving expression. From these, miR-22-3p is chosen for further study since it increased the expression of luciferase, two membrane proteins and a secreted fusion protein with minimal effect on the cells' growth and viability. Since each microRNA can interact with several gene targets, it is of interest to identify the repressed genes for understanding and exploring the improved expression mechanism for further implementation. Here, the authors describe a novel approach for identification of the target genes by integrating the differential gene expression analysis with information obtained from our previously conducted high-throughput siRNA screening. The identified genes were validated as being involved in improving luciferase expression by using siRNA and qRT-PCR. Repressing the target gene, HIPK1, is found to increase luciferase and GPC3 expression 3.3- and 2.2-fold, respectively.

KEYWORDS:

HIPK1; miR-22-3p; microRNA; microarray; protein expression; siRNA

PMID:
28987030
PMCID:
PMC5823020
DOI:
10.1002/biot.201700342
[Indexed for MEDLINE]
Free PMC Article

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