Transcriptional profiling of batch and fed-batch protein-free 293-HEK cultures using DNA microarray

Abstract

HEK (Human Embryonic Kidney) cells were observed to reach 4 times the cell density in low-glutamine protein-free fed-batch cultures when compared to batch cultures. This was accompanied with a reduction in lactate and ammonia production in fed-batch cultures. Studies with virus infection of both batch and fed-batch cultures have also shown an improvement in virus production of about 10,000-fold in fed- batch. An endeavor was undertaken to decipher the cellular transcriptional regulation underlying the fed-batch process using microarray. This study presents results focusing on the genes related to amino acid metabolism, tRNA processing and energy metabolism. These processes are intimately related to cell growth and protein production, two issues of importance to bioprocessing. Our results showed that amino acid metabolism enzymes (eg. ASNS, GLUD1, GOT1) and a number of tRNA synthetases (eg. EPRS, YARS, WARS, GARS) were found to have consistent differences in expression patterns between batch and fed-batch, possibly due to differences in nutrient environment of the two cultures. The expression patterns of 3 energy metabolism-related genes, SLC25A5, COX6B and SUCLG2, were also found to be dissimilar in batch compared to fed- batch, indicating disparity in energy efficiency of the cells. Our results suggest that the microarray platform can effectively be utilized as a tool to monitor transcriptional events of cells in culture. These observations, together with other insights gleaned from further analysis of the data, might be valuable in a rational approach to engineering of robust cell-lines with improved cellular metabolism.