Complementation analysis of the cold-sensitive phenotype of the Escherichia coli csdA deletion strain

Department of Biochemistry, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, New Jersey 08854, USA

	Abstract

The cold-shock response of Escherichia coli is elicited by downshift of temperature from 37°C to 15°C and is characterized by induction of several cold-shock proteins including CsdA during acclimation phase. CsdA, a DEAD-box protein has been proposed to participate in a variety of processes, such as ribosome biogenesis, mRNA decay, translation initiation, and gene regulation. It is not clear which or all of the functions of CsdA play a role in its essential cold-shock function and so far no protein was shown to complement its function in vivo. Our screening of an E. coli genomic library for in vivo counterpart of CsdA that can compensate for its absence at low temperature revealed only one protein, another DEAD-box RNA helicase, RhlE. We also observed that although not detected in our genetic screen, two cold-shock-inducible proteins namely, an RNA chaperone, CspA and an exonuclease, RNase R can also complement cold-shock function of CsdA. Interestingly, absence of CsdA and RNase R leads to increased sensitivity of the cells to even moderate temperature downshifts. The correlation between the helicase activity of CsdA and stability of mRNAs of cold-inducible genes was shown using cspA mRNA, which was significantly stabilized in the csdA cells; an effect counteracted by overexpression of wild-type CsdA or RNase R, but not by the helicase-deficient mutant of CsdA. These results suggest that the primary role of CsdA in cold acclimation of cells is in mRNA decay and its helicase activity is pivotal for promoting degradation of mRNAs stabilized at low temperature.