Regulation of signaling directionality revealed by 3D snapshots of a kinase:regulator complex in action
- Institut Pasteur de Montevideo, Uruguay
- Universidad de Buenos Aires, Argentina
Abstract
Two-component systems (TCS) are protein machineries that enable cells to respond to input signals. Histidine kinases (HK) are the sensory component, transferring information toward downstream response regulators (RR). HKs transfer phosphoryl groups to their specific RRs, but also dephosphorylate them, overall ensuring proper signaling. The mechanisms by which HKs discriminate between such disparate directions, are yet unknown. We now disclose crystal structures of the HK:RR complex DesK:DesR from Bacillus subtilis, comprising snapshots of the phosphotransfer and the dephosphorylation reactions. The HK dictates the reactional outcome through conformational rearrangements that include the reactive histidine. The phosphotransfer center is asymmetric, poised for dissociative nucleophilic substitution. The structural bases of HK phosphatase/phosphotransferase control are uncovered, and the unexpected discovery of a dissociative reactional center, sheds light on the evolution of TCS phosphotransfer reversibility. Our findings should be applicable to a broad range of signaling systems and instrumental in synthetic TCS rewiring.
Data availability
-
Crystal structure of phosphorylated DesKCPublicly available at the RCSB Protein Data Bank (accession no: 5IUM).
-
Crystal structure of the DesK-DesR complex in the phosphatase statePublicly available at the RCSB Protein Data Bank (accession no: 5IUN).
-
Crystal structure of the DesK-DesR complex in the phosphotransfer state with low Mg2+ (20 mM)Publicly available at the RCSB Protein Data Bank (accession no: 5IUJ).
-
Crystal structure of the DesK-DesR complex in the phosphotransfer state with high Mg2+ (150 mM)Publicly available at the RCSB Protein Data Bank (accession no: 5IUK).
-
Crystal structure of the DesK-DesR complex in the phosphotransfer state with high Mg2+ (150 mM) and BeF3Publicly available at the RCSB Protein Data Bank (accession no: 5IUL).
Article and author information
Author details
Alejandro Buschiazzo
Funding
Agencia Nacional de Investigación e Innovación (FCE2009_1_2679)
- Felipe Trajtenberg
- Alejandro Buschiazzo
Agence Nationale de la Recherche (PCV06_138918)
- Alejandro Buschiazzo
FOCEM (COF 03/11)
- Alejandro Buschiazzo
Centro de Biologia Estructural del Mercosur
- Alejandro Buschiazzo
Agencia Nacional de Investigación e Innovación (FCE2007_219)
- Felipe Trajtenberg
- Alejandro Buschiazzo
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Michael Laub, Massachusetts Institute of Technology, United States
Version history
- Received: September 11, 2016
- Accepted: December 9, 2016
- Accepted Manuscript published: December 12, 2016 (version 1)
- Version of Record published: January 12, 2017 (version 2)
- Version of Record updated: September 5, 2017 (version 3)
Copyright
© 2016, Trajtenberg et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,068
- views
-
- 490
- downloads
-
- 48
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Regulation of signaling directionality revealed by 3D snapshots of a kinase:regulator complex in action
eLife 5:e21422.
https://doi.org/10.7554/eLife.21422
Further reading
-
- Cell Biology
- Structural Biology and Molecular Biophysics
Acetylation of α-tubulin at the lysine 40 residue (αK40) by αTAT1/MEC-17 acetyltransferase modulates microtubule properties and occurs in most eukaryotic cells. Previous literatures suggest that acetylated microtubules are more stable and damage resistant. αK40 acetylation is the only known microtubule luminal post-translational modification site. The luminal location suggests that the modification tunes the lateral interaction of protofilaments inside the microtubule. In this study, we examined the effect of tubulin acetylation on the doublet microtubule (DMT) in the cilia of Tetrahymena thermophila using a combination of cryo-electron microscopy, molecular dynamics, and mass spectrometry. We found that αK40 acetylation exerts a small-scale effect on the DMT structure and stability by influencing the lateral rotational angle. In addition, comparative mass spectrometry revealed a link between αK40 acetylation and phosphorylation in cilia.
-
- Structural Biology and Molecular Biophysics
The dimeric two-pore OSCA/TMEM63 family has recently been identified as mechanically activated ion channels. Previously, based on the unique features of the structure of OSCA1.2, we postulated the potential involvement of several structural elements in sensing membrane tension (Jojoa-Cruz et al., 2018). Interestingly, while OSCA1, 2, and 3 clades are activated by membrane stretch in cell-attached patches (i.e. they are stretch-activated channels), they differ in their ability to transduce membrane deformation induced by a blunt probe (poking). Here, in an effort to understand the domains contributing to mechanical signal transduction, we used cryo-electron microscopy to solve the structure of Arabidopsis thaliana (At) OSCA3.1, which, unlike AtOSCA1.2, only produced stretch- but not poke-activated currents in our initial characterization (Murthy et al., 2018). Mutagenesis and electrophysiological assessment of conserved and divergent putative mechanosensitive features of OSCA1.2 reveal a selective disruption of the macroscopic currents elicited by poking without considerable effects on stretch-activated currents (SAC). Our results support the involvement of the amphipathic helix and lipid-interacting residues in the membrane fenestration in the response to poking. Our findings position these two structural elements as potential sources of functional diversity within the family.
