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J. Biol. Chem., Vol. 277, Issue 8, 6005-6011, February 22, 2002

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The Cold Box Stem-loop Proximal to the 5'-End of the Escherichia coli cspA Gene Stabilizes Its mRNA at Low Temperature^*

Bing Xia, Haiping Ke, Wei Jiang, and Masayori Inouye^§

From the Department of Biochemistry, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854

The 5'-end region of cspA mRNA contains a Cold Box sequence conserved among several cold-shock mRNAs. This region forms a stable stem-loop structure followed by an AU-rich sequence. Here we show that the Cold Box region is essential for the normal scale of cspA mRNA induction after cold shock because a deletion of the stem-loop significantly destabilizes the mRNA and reduces the cold shock-induced cspA mRNA amount by ~50%. The AU-rich track, however, slightly destabilizes the mRNA. The integrity of the stem is essential for the stabilizing function, whereas that of the loop sequence is less important. Overexpression of a mutant cspA mRNA devoid of both the AUG initiation codon and the coding sequence results in a severe growth inhibition at low temperature along with a derepression of the chromosomal cspA expression. Furthermore, the overexpressed RNA is stably associated with the 30 S and 70 S ribosomes. Our results demonstrate that the AUG initiation codon and the coding region containing the downstream box are not required for cspA mRNA to bind ribosomes and that the 5'-untranslated region by itself has a remarkable affinity to ribosomes at low temperature.

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