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Originally published In Press as doi:10.1074/jbc.M110226200 on December 21, 2001

J. Biol. Chem., Vol. 277, Issue 9, 7282-7286, March 1, 2002
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Residues 137 and 153 at the N Terminus of the XylS Protein Influence the Effector Profile of This Transcriptional Regulator and the sigma  Factor Used by RNA Polymerase to Stimulate Transcription from Its Cognate Promoter*

Raquel RuizDagger and Juan L. Ramos§

From the Department of Biochemistry and Molecular and Cellular Biology of Plants, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Apartado de Correos 419, E-18008 Granada, Spain

The 321-residue XylS and XylS1 proteins, encoded by the pWW0 and pWW53 plasmids respectively, differ in only 5 residues at positions 4, 53, 90, 137, and 153. As a result, the effector profile of XylS is wider than that of XylS1, and XylS mediates higher levels of transcription from its cognate-regulatable promoter than does XylS1. We generated a series of XylS-pWW0 mutants and found that the single mutants Asp-137 right-arrow Glu and His-153 right-arrow Asn exhibited an activation pattern different from that of the wild-type regulator. In the double-mutant XylSD137E,H153N the effector profile for benzoates was similar to that of XylS1. This suggests that these two residues are crucial for effector recognition and regulator activation to stimulate transcription. XylS-dependent transcription from its cognate promoter is mediated by RNA polymerase with sigma 32 or sigma 38, whereas XylS1 uses RNA polymerase with sigma 32 or sigma 70. We also found that point mutations at positions 137 and 153 of XylS led RNA polymerase to mediate transcription with sigma 70 rather than with sigma 38, as demonstrated by primer extension analysis in a sigma 70-thermosensitive background proficient and deficient in sigma 38. This suggests that a positive transcriptional regulator can choose the RNA polymerase complex that mediates transcription from a given promoter.

* This study was supported by European Commission Grant QLK3-2000-0170 and the Spanish Comisión Interministerial de Ciencia y Tecnología Grant BIO2000-0964.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Dagger Recipient of a fellowship from the Spanish Ministry of Education and Culture.

§ To whom correspondence should be addressed. Tel: 34-958-121011; Fax: 34-958-129600; E-mail: jlramos@eez.csic.es.

Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
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