Large scale gene expression profiling of metabolic shift of mammalian cells in culture

Rashmi Korke, Marcela De Leon Gatti, Ally Lei Yin Lau, Justin Wee Eng Lim, Teck Keong Seow, Maxey Ching Ming Chung, Wei Shou Hu

Research output: Contribution to journalArticlepeer-review

73 Scopus citations

Abstract

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 (ΔL/ΔG) 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.

Original languageEnglish (US)
Pages (from-to)1-17
Number of pages17
JournalJournal of Biotechnology
Volume107
Issue number1
DOIs
StatePublished - Jan 8 2004

Bibliographical note

Funding Information:
This work was supported in part by a grant to WSH from the National Science Foundation, USA (BES-97272) and by the Academic Health Center at the University of Minnesota. cDNA libraries and clones derived from brain regions of adult mouse (strain C57BL6) were obtained from Research Genetics, Inc. These clones and libraries were constructed by M. Bento Soares, Ph.D. at the University of Iowa under contract NO1 MH80014, which was awarded by the National Institute of Mental Health (NIMH) and the National Institute of Neurological Disorders and Stroke (NINDS) as part of the Brain Molecular Anatomy Project (BMAP). BMAP is an NIH initiative to support molecular carography of the mammalian nervous system through the localization of genes and the analysis of their expression patterns. We thank Pam Skinner, the Biomedical Genomics Center (BMGC) and the UMN mouse group for microarray preparation. The generous gifts from Merck Research Laboratories and Sankyo Co. are also gratefully acknowledged.

Keywords

  • Affymetrix GeneChips
  • Cell culture
  • DNA microarrays
  • Genomics
  • Mass spectrometry
  • Metabolic shift
  • Metabolism
  • Proteomics
  • Two-dimensional gel electrophoresis

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