Large scale gene expression profiling of metabolic shift of mammalian cells in culture.
The metabolic state of hybridoma cells in continuous culture varies with the cultivation condition from which the culture is initiated. At a metabolically shifted state, cells have markedly reduced glucose and other nutrient consumption and lactate production as compared to cells in batch culture or in continuous culture without a metabolic shift. Taking a combined genomics and proteomics approach, we investigated the molecular mechanism of metabolic shift. Cells from continuous cultures at two different steady states with a glucose consumption to lactate production molar ratio (DeltaL/DeltaG) of 0.08 and 1.4 were studied. Affymetrix GeneChips as well as cDNA microarrays were employed to identify differentially expressed mRNA transcripts, and the differentially expressed proteins were identified using the 2D gel electrophoresis-mass spectrometry approach. The decrease in glucose metabolism upon metabolic shift is accompanied by a decrease in gene expression of a number of genes involved in its metabolism. However, the number of genes differentially expressed and the extent of differential expression upon metabolic shift are relatively moderate. The change in the expression of metabolic genes at the transcriptional level was confirmed by real time PCR. The results suggest that metabolic shift is a combined effect of both biochemical events at reaction level and gene expression at transcription and translation level. This approach of integrating transcriptional profiling, proteomic techniques and biochemical analysis provides a more global view of the metabolism of mammalian cells in culture.
Korke
Rashmi
R
Department of Chemical Engineering and Materials Science, University of Minnesota, 421 Washington Avenue SE, Minneapolis, MN 55455-0132, USA.
Gatti
Marcela de Leon
Mde L
Lau
Ally Lei Yin
AL
Lim
Justin Wee Eng
JW
Seow
Teck Keong
TK
Chung
Maxey Ching Ming
MC
Hu
Wei-Shou
WS
D000818:Animals; D002460:Cell Line; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000502:physiology*; D005947:Glucose / Q000378:metabolism; D006825:Hybridomas / Q000378:metabolism*; D019344:Lactic Acid / Q000096:biosynthesis; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D020543:Proteome / Q000235:genetics* / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D014158:Transcription, Genetic / Q000502:physiology
D020543:Proteome; D019344:Lactic Acid; D005947:Glucose