Abstract
Human enteric adenoviruses (HAdVs; serotypes 40 and 41) are important waterborne and food-borne pathogens. However, HAdVs are fastidious, are difficult to cultivate, and do not produce a clear cytopathic effect during cell culture within a reasonable time. Thus, we examined whether the viral transactivator proteins cytomegalovirus (CMV) IE1 and hepatitis B virus (HBV) X promoted the multiplication of HAdVs. Additionally, we constructed a new 293 cell line expressing CMV IE1 protein for cultivation assays. We analyzed the nucleic acid sequences of the promoter regions of both E1A and hexon genes, which are considered to be the most important regions for HAdV replication. Expression of either HBV X or CMV IE1 protein significantly increased the promoter activities of E1A and hexon genes of HAdVs by as much as 14-fold during cell cultivation. The promotion of HAdV expression was confirmed by increased levels of both adenoviral DNA and mRNA expression. Finally, the newly developed 293 cell line expressing CMV IE1 protein showed an increase in viral DNA ranging from 574% to 619% compared with the conventional 293 cell line. These results suggest that the newly constructed cell line could be useful for efficient cultivation and research of fastidious HAdVs.
Highlights
Human enteric adenoviruses (HAdVs; serotypes 40 and 41) are among the most common etiological agents of gastroenteritis, among children [1, 33]
We examined whether the viral transactivator proteins cytomegalovirus (CMV) IE1 and hepatitis B virus (HBV) X promoted the multiplication of HAdVs
E1A promoter showed 3-fold greater activation than the hexon promoter by the two transactivator proteins, with average values of 1,121% and 1,421%, respectively (P ϭ 0.174 and P ϭ 0.003, respectively). These results suggest that viral transactivators, including HBV X and CMV IE1 proteins, can increase the transcription of essential genes of HAdVs
Summary
Human enteric adenoviruses (HAdVs; serotypes 40 and 41) are among the most common etiological agents of gastroenteritis, among children [1, 33]. HAdVs are cultivable in several cell lines, including 293, A549, PLC/PRF5, and Caco-2 cells, they are fastidious and do not produce a clear and consistent cytopathic effect (CPE) within a reasonable time [6, 17,18,19, 20, 31] They are sensitive to type I interferon (IFN), and the HAdV E1A gene is deficient in its ability to transactivate its own genes [4, 23, 36, 39]. Cellular transcription factors such as AP1 or NF-B can be introduced into cells and can markedly increase gene transcription [35] These biological characteristics can be applied both to increase the levels of target mRNA and to promote the multiplication of fastidious HAdVs in cell culture. The objectives of the present study were to determine whether viral transactivation proteins, including HBV X and CMV IE1, can activate the transcription of essential genes of HAdVs and subsequently promote the replication of HAdVs and to construct a new cell line that promotes the replication of fastidious HAdVs
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