title: Identification of microRNAs specific for high producer CHO cell lines using steady-state cultivation.
abstract: MicroRNAs are short non-coding RNAs that play an important role in the regulation of gene expression. Hence, microRNAs are considered as potential targets for engineering of Chinese hamster ovary (CHO) cells to improve recombinant protein production. Here, we analyzed and compared the microRNA expression patterns of high, low, and non-producing recombinant CHO cell lines expressing two structurally different model proteins in order to identify microRNAs that are involved in heterologous protein synthesis and secretion and thus might be promising targets for cell engineering to increase productivity. To generate reproducible and comparable data, the cells were cultivated in a bioreactor under steady-state conditions. Global microRNA expression analysis showed that mature microRNAs were predominantly upregulated in the producing cell lines compared to the non-producer. Several microRNAs were significantly differentially expressed between high and low producers, but none of them commonly for both model proteins. The identification of target messenger RNAs (mRNAs) is essential to understand the biological function of microRNAs. Therefore, we negatively correlated microRNA and global mRNA expression data and combined them with computationally predicted and experimentally validated targets. However, statistical analysis of the identified microRNA-mRNA interactions indicated a considerable false positive rate. Our results and the comparison to published data suggest that the reaction of CHO cells to the heterologous protein expression is strongly product- and/or clone-specific. In addition, this study highlights the urgent need for reliable CHO-specific microRNA target prediction tools and experimentally validated target databases in order to facilitate functional analysis of high-throughput microRNA expression data in CHO cells.
authors: Austrian Centre of Industrial Biotechnology (ACIB GmbH), Muthgasse 11, 1190, Vienna, Austria, andreas.maccani@acib.at.
authors: Andreas
authors: A
authors: Maccani
authors: Matthias
authors: M
authors: Hackl
authors: Christian
authors: C
authors: Leitner
authors: Willibald
authors: W
authors: Steinfellner
authors: Alexandra B
authors: AB
authors: Graf
authors: Nadine E
authors: NE
authors: Tatto
authors: Michael
authors: M
authors: Karbiener
authors: Marcel
authors: M
authors: Scheideler
authors: Johannes
authors: J
authors: Grillari
authors: Diethard
authors: D
authors: Mattanovich
authors: Renate
authors: R
authors: Kunert
authors: Nicole
authors: N
authors: Borth
authors: Reingard
authors: R
authors: Grabherr
authors: Wolfgang
authors: W
authors: Ernst
mesh_terms: D000818:Animals; D019149:Bioreactors; D016466:CHO Cells / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods; D003412:Cricetulus; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D035683:MicroRNAs / Q000378:metabolism*
chemical_list: D035683:MicroRNAs