ANXA2 KO cell lines help research on the development of tumors

Annexin A2(ANXA2) gene is located on chromosome 15 and the encoded protein consists of 339 amino acids with a size of 36kDa. It is an important member of Annexin family, which is a calcium-dependent phospholipid binding protein widely existing in plants and animals. ANXA2 protein can be produced by a variety of cells, including vascular endothelial cells, monocytes, macrophages, dendritic cells, trophoblast cells, epithelial cells, and tumor cells. It can be present as free monomers in the cytoplasm, bind to the inner membrane of cells or attach to the outer surface of the plasma membrane. It is mainly involved in membrane transport and a series of calmodulin-dependent activities on the membrane surface, including membrane fusion in exocytosis, vesicle transport, cell adhesion, cell proliferation, apoptosis, replication, signal transduction and ion channel formation.

 In recent years,  numbers of studies have confirmed that expression dysregulation of ANXA2 is closely related to the occurrence and development of a variety of tumors, and its expression is up-regulated in most human tumors. Such overexpression is closely related to the occurrence, development, invasion, metastasis and poor prognosis of tumors.

Regulatory effects of ANXA2 on the behavior of human colorectal cancer cells.

It has been reported that ANXA2 interacts with cytoskeleton proteins (such as Actin F-actin) and kinds of cytoskeleton allosteric and motion-related regulatory factors to influence malignant behavior of tumor cells. In order to elucidate the role of ANXA2 in the development of colorectal cancer, the researchers construct ANXA2^-/-Caco2 cell line by knockouting ANXA2 gene in human colorectal cancer cell line Caco2. Using wild type Caco2 cells (ANXA2^+/+Caco2) as control, they studied the effect of ANXA2 gene expression on the growth, motonation and apoptosis of cancer cells. The results showed that the growth of filamentous pseudopodia on Caco2 KO cell surface was slow, and the microvilli became thinner and less; Tem results showed that more chromatin agglutination, less mitochondria, and nuclear membrane depolymerization were observed in KO cells; In wild-type Caco2 cells, f-actin was densely distributed along the inner surface of plasma membrane and cytoplasm, while in KO cells, it was significantly decreased. Besides,β-tubulin was decreased in cytoplasm, but the difference was not significant; LaminB was densely distributed in the medial nuclear membrane of wild type Caco2 cells, while was significantly reduced in KO cells. Ubigene can provide the ANXA2 gRNA KO plasmid for you to efficiently construct  ANXA2 KO cell line. In addition, Ubigene has over 10000 gRNA plasmids in stock. if you are interested, please click to view >>

 In conclusion, ANXA2 is closely related to the growth, proliferation, apoptosis and motility of human colorectal cancer Caco2 cells, and the inhibition of ANXA2's expression can inhibit the proliferation and migration ability of Caco2 cells in vitro, and promote cell apoptosis.

ANXA2 KO CHO cell line was constructed to evaluate the effect of gene knockout on HCP reduction.

Chinese hamster ovary (CHO) cells are often used as host cells to produce a range of recombinant therapeutic proteins, including monoclonal antibodies and FC fusion proteins.  During this process, host cellular proteins (HCP) must be removed from formulations due to the potential risk of immunogenicity. While most HCP impurities can be effectively removed during typical downstream purification processes, removing small amounts of HCP can still be challenging.

Researchers successfully constructed ANXA2 and ctsd KO CHO cell lines with CRISPR/Cas9 to evaluate the feasibility of knockout methods to reduce the risk of HCP contamination. The results confirmed that the corresponding HCP was completely eliminated in cell lysates. Which means knockouting the unnecessary genes can reduce the contamination of HCP to the production of recombinant therapeutic proteins. However, further studies are needed to determine the effect of gene knockout on the yield of these cell lines.

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ANXA2 KO MDCK cell lines were constructed to verify εtoxin receptor on MDCK cell surface.

PALentiCRISPR V2 recombinant plasmid was constructed according to CRISPR/Cas9 target design principle. The constructed plasmid was transfected into MDCK cells, and the MDCK cell lines with ANXA2 protein knockout were screened by purinomycin. The cytotoxicity test was performed to verify whether ANXA2 protein on the surface of the MDCK cell membrane was the receptor protein of ε toxin. The results of cytotoxicity assay showed that knockout ANXA2 gene did not reduce the toxicity ofεtoxin to MDCK cells, which indicate that ANXA2 protein is not the receptor ofεtoxin on MDCK cell surface. The results of this study provide a test method for the knockout of other proteins in MDCK cells in the future, and the constructed MDCK cells with ANXA2 gene knockout also lay a foundation for the study of other diseases caused by ANXA2 protein. If you are interested in this cell, Ubigene can provide a variety of MDCK cells, including KO/wild-type /Cas9/Luc cell lines. Please click here to learn more >>

"Make gene editing easier!"has always been Ubigene aspiration. Ubigene developed the CRISPR-U™ technology, which is greatly improve the efficiency of homologous recombination. With CRISPR-U™ , Ubigene has successfully generated over 5000 KO cell lines for the researchers all over the world. In addition, Ubigene now has nearly 2,000 types of KO cells in stock. For more information about our services,  please inquire >>

Reference：

[1] Lin Zhiyun, Song Ying, He Wei, DAI Haibin. Structure, function and research progress of Annexin A2 in disease [J]. Chinese journal of modern applied pharmacy,2016,33(10):1350-1354.

[2] Cui T T. Study on motion-related morphology of Caco2 cells after ANXA2 knockout in colon cancer [D]. Shaanxi Normal University,2012.

[3] Fukuda Nobuo,Senga Yukako,Honda Shinya. Anxa2- and Ctsd-knockout CHO cell lines to diminish the risk of contamination with host cell proteins.[J]. Biotechnology progress,2019,35(4):

[4] Huang Jing, GAO Jie, XIA Susu, JIN Zhiying, KANG Lin, GAO Shan, Yang Hao, Xin Wenwen, ZHAO Baohua, WANG Jinglin. Construction of Annexin A2 gene knockout MDCK cell line [J]. Advances in veterinary medicine,2019,40 (05):13-17.DOI:10.16437 /j.cnki.1007-5038.2019.05.003.