Allosteric Activation of Bacterial Response Regulators: the Role of the Cognate Histidine Kinase Beyond Phosphorylation

Abstract

Response regulators are proteins that undergo transient phosphorylation, connecting specific signals to adaptive responses. Remarkably, the molecular mechanism of response regulator activation remains elusive, largely because of the scarcity of structural data on multidomain response regulators and histidine kinase/response regulator complexes. We now address this question by using a combination of crystallographic data and functional analyses in vitro and in vivo, studying DesR and its cognate sensor kinase DesK, a two-component system that controls membrane fluidity in Bacillus subtilis. We establish that phosphorylation of the receiver domain of DesR is allosterically coupled to two distinct exposed surfaces of the protein, controlling noncanonical dimerization/tetramerization, cooperative activation, and DesK binding. One of these surfaces is critical for both homodimerization- and kinase-triggered allosteric activations. Moreover, DesK induces a phosphorylation-independent activation of DesR in vivo, uncovering a novel and stringent level of specificity among kinases and regulators. Our results support a model that helps to explain how response regulators restrict phosphorylation by small-molecule phosphoryl donors, as well as cross talk with noncognate sensors.Fil: Trajtenberg, Felipe. Instituto Pasteur de Montevideo; UruguayFil: Albanesi, Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; Argentina. Universidad Nacional de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Ruetalo, Natalia. Instituto Pasteur de Montevideo; UruguayFil: Botti, Horacio. Instituto Pasteur de Montevideo; UruguayFil: Mechaly, Ariel. Instituto Pasteur de Montevideo; Uruguay. Institut Pasteur, Unité de Microbiologie Structurale, Paris; FranciaFil: Nieves, Marcos. Instituto Pasteur de Montevideo, Laboratorio de Biología Celular de Membranas, Montevideo; UruguayFil: Aguilar, Pablo Sebastián. Instituto Pasteur de Montevideo, Laboratorio de Biología Celular de Membranas, Montevideo; UruguayFil: Cybulski, Larisa Estefania. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; Argentina. Universidad Nacional de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Larrieux, Nicole. Instituto Pasteur de Montevideo; UruguayFil: de Mendoza, Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; Argentina. Universidad Nacional de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Buschiazzo, Alejandro. Instituto Pasteur de Montevideo; Uruguay. Institut Pasteur, Département de Biologie Structurale et Chimie, Paris; Franci