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Effects of VACM‐1/Cul5 Gene Knockout on Cellular Proliferation Using CRISPR‐Cas9 Approach
Author(s) -
Gager Christopher,
Rios Santiago,
Cunningham Mark,
BurnatowskaHledin Maria
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.916.9
Subject(s) - crispr , knockout mouse , ubiquitin ligase , gene knockout , microbiology and biotechnology , biology , gene , cancer research , ubiquitin , genetics
The VACM‐1 gene codes for the VACM‐1/Cul5 protein, which is a part of the ubiquitin E3 ligase system. This system is utilized to degrade cellular proteins. VACM‐1/Cul5 expression is shown to decrease cellular proliferation. Lack of regulation in this pathway can lead to cancer. We utilized CRISPR‐Cas9 to knockout the VACM‐1 gene. The CRISPR system is a bacterial immune system that functions by targeting specific sequences of DNA. It can be programmed to target genes of interest, enabling specific gene editing in eukaryotic cells. We have used this system to knockout VACM‐1/Cul5 in a human umbilical vein endothelial cell line (HUVEC). Growth assays indicate that VACM‐1/Cul5 knockout allows cells to proliferate at an increased rate. We also explored whether CRISPR‐Cas9 knockout of VACM‐1/Cul5 compromises the antiproliferative effects of resveratrol. Our results indicate that VACM‐1/Cul5 knockout cells do not respond to these drugs in the same manner as normal HUVEC cells, and experienced more rapid growth. We also aim to prove gene knockout via genomic analysis, as well as to use microarrays to explore possible pathways impacted by the knockout.