Bauer, Asli Petra and Leikam, Doris and Krinner, Simone and Notka, Frank and Ludwig, Christine and Laengst, Gernot and Wagner, Ralf (2010) The impact of intragenic CpG content on gene expression. NUCLEIC ACIDS RESEARCH, 38 (12). pp. 3891-3908. ISSN 0305-1048, 1362-4962
Full text not available from this repository. (Request a copy)Abstract
The development of vaccine components or recombinant therapeutics critically depends on sustained expression of the corresponding transgene. This study aimed to determine the contribution of intragenic CpG content to expression efficiency in transiently and stably transfected mammalian cells. Based upon a humanized version of green fluorescent protein (GFP) containing 60 CpGs within its coding sequence, a CpG-depleted variant of the GFP reporter was established by carefully modulating the codon usage. Interestingly, GFP reporter activity and detectable protein amounts in stably transfected CHO and 293 cells were significantly decreased upon CpG depletion and independent from promoter usage (CMV, EF1 alpha). The reduction in protein expression associated with CpG depletion was likewise observed for other unrelated reporter genes and was clearly reflected by a decline in mRNA copy numbers rather than translational efficiency. Moreover, decreased mRNA levels were neither due to nuclear export restrictions nor alternative splicing or mRNA instability. Rather, the intragenic CpG content influenced de novo transcriptional activity thus implying a common transcription-based mechanism of gene regulation via CpGs. Increased high CpG transcription correlated with changed nucleosomal positions in vitro albeit histone density at the two genes did not change in vivo as monitored by ChIP.
Item Type: | Article |
---|---|
Uncontrolled Keywords: | IMMUNODEFICIENCY-VIRUS TYPE-1; TERM TRANSGENE EXPRESSION; MAMMALIAN-CELLS; DNA METHYLATION; BINDING-PROTEIN; CHROMATIN-STRUCTURE; CODON USAGE; HETEROLOGOUS EXPRESSION; TRANSIENT EXPRESSION; IMMUNE-RESPONSES; |
Subjects: | 500 Science > 570 Life sciences 600 Technology > 610 Medical sciences Medicine |
Divisions: | Medicine > Lehrstuhl für Medizinische Mikrobiologie und Hygiene Biology, Preclinical Medicine > Institut für Biochemie, Genetik und Mikrobiologie > Lehrstuhl für Biochemie III > Prof. Dr. Gernot Längst |
Depositing User: | Dr. Gernot Deinzer |
Date Deposited: | 24 Aug 2020 06:38 |
Last Modified: | 24 Aug 2020 06:38 |
URI: | https://pred.uni-regensburg.de/id/eprint/25569 |
Actions (login required)
View Item |