Messenger RNA Translation State: The Second Dimension of High-Throughput Expression Screening

Technological advances over the past 10 years have generated powerful tools for parallel analysis of complex biological problems. Among these new technologies, DNA arrays have provided an important experimental approach for identifying changes in the levels of individual mRNA molecules during import...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1999-09, Vol.96 (19), p.10632-10636
Main Authors: Zong, Qin, Schummer, Michel, Hood, Leroy, Morris, David R.
Format: Article
Language:English
Subjects:
3T3 Cells
Animals
Biochemistry
Biological Sciences
Complementary DNA
Deoxyribonucleic acid
DNA
DNA, Complementary - metabolism
Fibroblasts
Fibroblasts - metabolism
Gene Expression Regulation
Genetics
Humans
Messenger RNA
Mice
Mitogens - metabolism
Molecular biology
Nucleic Acid Hybridization - methods
Oligonucleotide Array Sequence Analysis
Polyribosomes
Polyribosomes - metabolism
Protein Biosynthesis
Ribonucleic acid
Ribosomes
RNA
RNA, Messenger - genetics
RNA, Messenger - metabolism
Swiss 3T3 cells
Vimentin
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Summary:Technological advances over the past 10 years have generated powerful tools for parallel analysis of complex biological problems. Among these new technologies, DNA arrays have provided an important experimental approach for identifying changes in the levels of individual mRNA molecules during important cellular transitions. However, cellular behavior is dictated not by mRNA levels, but by the proteins translated from the individual mRNA species. We report a high-throughput method for simultaneously monitoring the translation state and level of individual mRNA species. Messenger RNAs from resting and mitogenically activated fibroblasts were separated, according to degree of ribosome loading, into well-translated and under-translated pools. cDNA probes generated from these fractions were used to interrogate cDNA arrays. Among approximately 1,200 genes analyzed, less than 1% were found to be translationally regulated in response to mitogenic activation, demonstrating the strong selectivity of this regulatory mechanism. This high-throughput approach is shown to be an effective tool for superimposing translation profile on mRNA level for large numbers of genes, as well as for identifying translationally regulated genes for further study.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.96.19.10632