HEK cell
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Human Embryonic Kidney cells, also known as HEK cells, HEK 293 or just 293 cells, are a cell line originally derived, as their name indicates, from an embryonic human kidney. HEK cells have been grown in tissue culture for many years and have become very widely used. They are very easy to grow and transfect very readily and so are widely-used in cell biology research. They are also used by the biotechnology industry to produce therapeutic proteins and viruses for gene therapy.
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[edit] Origins of HEK Cells
HEK 293 cells were generated by transformation of cultures of normal human embryonic kidney cells with sheared adenovirus 5 DNA in the laboratory of Alex Van der Eb in Leiden, Holland in the early 70s. The human embryonic kidney cells were obtained from a healthy aborted fetus and originally cultured by Van der Eb himself, and the transformation by adenovirus was performed by Frank Graham who published his findings in the late 1970s after he left Leiden for McMaster University in Canada.[1] They are called HEK for human embryonic kidney, while the number 293 comes from Graham's habit of numbering his experiments; the original HEK 293 cell clone was simply the product of his 293rd experiment.
Subsequent analysis has shown that the transformation was brought about by an insert consisting of ~4.5 kilobases from the left arm of the viral genome, which became incorporated into human chromosome 19.[2]
For many years it was assumed that HEK 293 cells were generated by transformation of either a fibroblastic, endothelial or epithelial cell all of which are abundant in kidney. However the fact that the cells originated from cultured kidney cells does not say much about the exact cellular origin of the HEK 293, as embryonic kidney cultures may contain small numbers of almost all cell types of the body. In fact Graham and coworkers more recently provided evidence that HEK 293 cells and several other human cell lines generated by adenovirus transformation of human embryonic kidney cells have many properties of immature neurons, suggesting that the adenovirus was taken up and transformed a neuronal lineage cell in the original kidney culture.[3]
[edit] Uses of HEK 293 Cells
As an experimentally transformed cell line, HEK cells are not a particularly good model for normal cells, cancer cells, or any other kind of cell that is a fundamental object of research. However, they are extremely easy to work with, being straightforward to culture and to transfect, and so can be used in experiments in which the behaviour of the cell itself is not of interest. Typically, these experiments involve transfecting in a gene (or combination of genes) of interest, and then analysing the expressed protein; essentially, the cell is used simply as a test tube with a membrane. The widespread use of this cell line is due to its extreme transfectability by the calcium phosphate method, achieving efficiencies approaching 100% as determined by FACS using a 2XPBS buffer. A lower efficiency might be achievable with an HBS buffer.
An important variant of this cell line is the 293T cell line that contains, in addition, the SV40 large T antigen, that allows for episomal replication of transfected plasmids containing the SV40 origin of replication. This allows for amplification of transfected plasmids and extended temporal expression of the desired gene products. Note that any similarly domesticated cell line can be used for this sort of work; HeLa, COS and Chinese Hamster Ovary cell are common alternatives.
Examples of such experiments include:
- A study of the effects of a drug on sodium channels[4]
- Testing of an inducible RNA interference system[5]
- Testing of an isoform-selective protein kinase C agonist[6]
- Investigation of the interaction between two proteins[7]
- Analysis of a nuclear export signal in a protein[8]
A more specific use of HEK cells is in the propagation of adenoviral vectors. Viruses offer an extremely efficient means of delivering genes into cells, since this is what they have evolved to do, and are thus of great use as experimental tools. However, as pathogens, they also present a degree of danger to the experimenter. This danger can be avoided by the use of viruses which lack key genes, and which are thus unable to replicate after entering a cell. In order to propagate such viral vectors, a cell line that expresses the missing genes is required. Since HEK cells express a number of adenoviral genes, they can be used to propagate adenoviral vectors in which these genes (typically, E1 and E3) are deleted, such as AdEasy.[9]
293, and especially 293T, cells are commonly used for the production of lentiviral and retroviral vectors. Various retroviral and lentiviral packaging cell lines are based on these cells.
[edit] Native proteins of interest
Depending on various conditions gene expression of HEK cells may vary. The following proteins of interest (among many others) are commonly found in untreated HEK cells:
- Corticotrophin releasing factor type 1 receptor[10]
- Sphingosine-1-phosphate receptors EDG1, EDG3 and EDG5[11]
- Muscarinic acetylcholine receptor M3[12]
- Transient receptor potential TRPC1, TRPC3, TRPC4, TRPC6[13]
[edit] External links
- HEK 293 Database
- 293 Cells (CRL-1573) in the ATCC database
- Transcript of FDA meeting, in which, starting page 77, Dr. Alex Van der Eb describes in detail the origin of HEK 293 cell
[edit] References
- ^ Graham FL, Smiley J, Russell WC, Nairn R (July 1977). "Characteristics of a human cell line transformed by DNA from human adenovirus type 5". J. Gen. Virol. 36 (1): 59–74. PMID 886304.
- ^ Louis N, Evelegh C, Graham FL (July 1997). "Cloning and sequencing of the cellular-viral junctions from the human adenovirus type 5 transformed 293 cell line". Virology 233 (2): 423–9. doi:. PMID 9217065.
- ^ Shaw G, Morse S, Ararat M, Graham FL (June 2002). "Preferential transformation of human neuronal cells by human adenoviruses and the origin of HEK 293 cells". FASEB J. 16 (8): 869–71. doi:. PMID 11967234.
- ^ Fredj S, Sampson KJ, Liu H, Kass RS (May 2006). "Molecular basis of ranolazine block of LQT-3 mutant sodium channels: evidence for site of action". Br. J. Pharmacol. 148 (1): 16–24. doi:. PMID 16520744.
- ^ Amar L, Desclaux M, Faucon-Biguet N, Mallet J, Vogel R (2006). "Control of small inhibitory RNA levels and RNA interference by doxycycline induced activation of a minimal RNA polymerase III promoter". Nucleic Acids Res. 34 (5): e37. doi:. PMID 16522642.
- ^ Kanno T, Yamamoto H, Yaguchi T, et al (June 2006). "The linoleic acid derivative DCP-LA selectively activates PKC-epsilon, possibly binding to the phosphatidylserine binding site". J. Lipid Res. 47 (6): 1146–56. doi:. PMID 16520488.
- ^ Li T, Paudel HK (March 2006). "Glycogen synthase kinase 3beta phosphorylates Alzheimer's disease-specific Ser396 of microtubule-associated protein tau by a sequential mechanism". Biochemistry 45 (10): 3125–33. doi:. PMID 16519507.
- ^ Mustafa H, Strasser B, Rauth S, Irving RA, Wark KL (April 2006). "Identification of a functional nuclear export signal in the green fluorescent protein asFP499". Biochem. Biophys. Res. Commun. 342 (4): 1178–82. doi:. PMID 16516151.
- ^ He TC, Zhou S, da Costa LT, Yu J, Kinzler KW, Vogelstein B (March 1998). "A simplified system for generating recombinant adenoviruses". Proc. Natl. Acad. Sci. U.S.A. 95 (5): 2509–14. PMID 9482916.
- ^ Dautzenberg FM, Higelin J, Teichert U (February 2000). "Functional characterization of corticotropin-releasing factor type 1 receptor endogenously expressed in human embryonic kidney 293 cells". Eur. J. Pharmacol. 390 (1-2): 51–9. PMID 10708706.
- ^ Meyer zu Heringdorf D, Lass H, Kuchar I, et al (March 2001). "Stimulation of intracellular sphingosine-1-phosphate production by G-protein-coupled sphingosine-1-phosphate receptors". Eur. J. Pharmacol. 414 (2-3): 145–54. PMID 11239914.
- ^ Luo J, Busillo JM, Benovic JL (April 2008). "M3 Muscarinic Acetylcholine Receptor-Mediated Signaling is Regulated by Distinct Mechanisms". Mol. Pharmacol.. doi:. PMID 18388243.
- ^ Zagranichnaya TK, Wu X, Villereal ML (August 2005). "Endogenous TRPC1, TRPC3, and TRPC7 proteins combine to form native store-operated channels in HEK-293 cells". J. Biol. Chem. 280 (33): 29559–69. doi:. PMID 15972814.
