ReviewHEK293 in cell biology and cancer research: phenotype, karyotype, tumorigenicity, and stress-induced genome-phenotype evolution
Introduction
293 cell line (original name designates the number of a successful transformation experiment; more widely known as the Human Embryonic Kidney 293 cells (HEK293)) was derived in 1973 by exposing the human primary embryonic kidney cell culture of an aborted embryo to the mechanically sheared DNA of adenovirus type 5 (AD5) (Graham et al., 1977). A recent analysis showed that 293 cell line and its derivatives (e.g., 293T or 293S) harbor 5–6 copies of AD5 DNA fragments encoding early gene 1 (E1) (Lin et al., 2014), and this copy number did not change from that originally reported (4–5 fragments) (Graham et al., 1977). An AD5 DNA integration site is located in pregnancy-specific beta-1-glycoprotein 4 (PSG4) gene, which was mapped to the chromosome region 19q13.2 (Lin et al., 2014, Louis et al., 1997). 293 cell line and its derivatives have been the most frequently used cells after HeLa in cell biology studies and after CHO in biotechnology (Lin et al., 2014) as a vehicle for the production of adenoviral vaccines (Kovesdi and Hedley, 2010) and recombinant proteins (Geisse and Fux, 2009) as well as for analysis of the neuronal synapse formation in mixed-culture assays (Biederer and Scheiffele, 2007), in electrophysiology and neuropharmacology (Thomas and Smart, 2005, Varghese et al., 2006). Also, 293 cells were often used as a model for studying the transforming/oncogenic properties of cancer-associated genes (Table 1) but sometimes incorrectly supposed as non-tumorigenic or even “normal” human cell line (discussed in Kavsan et al., 2011, Stepanenko and Kavsan, 2012). Despite the widespread and historically long-term productive exploitation in cell biology, biotechnology, and cancer research, an origin, phenotype, karyotype instability, and tumorigenicity of 293 cell line are still disputed and debated.
Section snippets
293 cell line: of kidney or adrenal gland origin and epithelial or neuronal phenotype?
293 cells have been considered the kidney embryonic epithelial cells (e.g., Ashokkumar et al., 2006, Cusick et al., 2010) or even fibroblasts (e.g., Cooper et al., 1998, Yung et al., 2012) and shown to express markers of a kidney developmental stage between the condensed mesenchyme and epithelial S-shaped body but not markers of the differentiated tubular segments, proximal tubules or collecting ducts (Torban and Goodyer, 1998). Moreover, an ectopic expression of transcription factor paired box
The complex karyotype of 293 cells and genome instability-driving mechanisms
293 cells are considered the female origin referring to the presence of several X chromosomes and lack of Y chromosome when conventional cytogenetics is employed (Gilbert et al., 2000, Machida et al., 2009, Meir et al., 2011). Actually, the high-coverage genome sequencing did not reveal a trace of Y-chromosome-derived sequence (Lin et al., 2014). Also, the short tandem repeat analysis with amelogenin marker showed female genotype of 293 cells (American Type Culture Collection, ATCC).
Karyotype
Both stable plasmid DNA transfection and overexpression of cancer-associated genes promote genome-phenotype evolution of tumorigenic 293 cell line
To study the dynamics of the 293 genome under the procedures commonly used in laboratory investigations and in biotechnological engineering, Lin et al. analyzed 293 cell line (from ATCC) and its derivatives using the multiplex fluorescence in situ hybridization, high-coverage genome sequencing, SNP array, and exon array: 1) 293T cell line, which expresses a temperature-sensitive allele of the SV40 T antigen; 2) 293S cell line, which was adapted to growth in suspension; 3) 293SG cell line, which
Concluding remarks
We raise a concern about the use of 293 cell line as a model in studying renal functions or as “normal” human cells in transformation transgenic studies and point out to a significant karyotype diversification of 293 cells and their laboratory/biotechnological derivatives. The aberrant karyotype, genomic heterogeneity, and constitutive expression of adenoviral E1 gene make transcriptome of 293 cells severely deregulated, bringing to naught affords to elucidate an authentic cell type of origin
Conflict of interest
No potential conflicts of interest were disclosed.
Acknowledgments
This work was supported in frames of the programs “Fundamental grounds of molecular and cell biotechnologies” №38/14 and “Nanotechnologies and nanomaterials for 2010–2014 years” №5.16.3.14 by the National Academy of Sciences of Ukraine (NASU).
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2023, Advanced Drug Delivery ReviewsCitation Excerpt :Interestingly, expression of recombinant cell-surface mucins can substantially mitigate HEK293-F clumping in suspension[121]. Like CHO cells and other immortalized cell lines, HEK293 cells exhibit genomic instability characterized by chromosomal translocations, copy number alterations, and other events, raising concerns of karyotypic and phenotypic drift during prolonged cultivation[122–124]. The potential for drift in O-glycosylation patterns during long-term cultivation and subcloning in human cell lines must still be addressed.