TY - JOUR
T1 - Salvador has an extended SARAH domain that mediates binding to Hippo kinase
AU - Cairns, Leah
AU - Tran, Thao
AU - Fowl, Brendan H.
AU - Patterson, Angela
AU - Kim, Yoo Jin
AU - Bothner, Brian
AU - Kavran, Jennifer M.
N1 - Funding Information:
This work was supported by National Institutes of Health Grants T32GM007445 (to L. C.) and T32CA009110 (to T. T.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Funding Information:
This work was supported by National Institutes of Health Grants T32GM007445 (to L. C.) and T32CA009110 (to T. T.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The Montana State University Proteomics, Metabo-lomics, and Mass Spectrometry Facility is supported in part by funding from the Murdock Charitable Trust and National Institutes of Health Grant P20GM103474 of the IDEA program. Plasmids encoding Renilla and firefly luciferase and Gal4DBD-Yki used in the reporter assay were provided by K. Irvine.
Funding Information:
Acknowledgments—The Montana State University Proteomics, Metabo-lomics, and Mass Spectrometry Facility is supported in part by funding from the Murdock Charitable Trust and National Institutes of Health Grant P20GM103474 of the IDEA program. Plasmids encoding Renilla and firefly luciferase and Gal4DBD-Yki used in the reporter assay were provided by K. Irvine.
Publisher Copyright:
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.
PY - 2018/4/13
Y1 - 2018/4/13
N2 - The Hippo pathway controls cell proliferation and differentiation through the precisely tuned activity of a core kinase cassette. The activity of Hippo kinase is modulated by interactions between its C-terminal coiled-coil, termed the SARAH domain, and the SARAH domains of either dRassF or Salvador. Here, we wanted to understand the molecular basis of SARAH domain–mediated interactions and their influence on Hippo kinase activity. We focused on Salvador, a positive effector of Hippo activity and the least well-characterized SARAH domain–containing protein. We determined the crystal structure of a complex between Salvador and Hippo SARAH domains from Drosophila. This structure provided insight into the organization of the Salvador SARAH domain including a folded N-terminal extension that expands the binding interface with Hippo SARAH domain. We also found that this extension improves the solubility of the Salvador SARAH domain, enhances binding to Hippo, and is unique to Salvador. We therefore suggest expanding the definition of the Salvador SARAH domain to include this extended region. The heterodimeric assembly observed in the crystal was confirmed by cross-linked MS and provided a structural basis for the mutually exclusive interactions of Hippo with either dRassF or Salvador. Of note, Salvador influenced the kinase activity of Mst2, the mammalian Hippo homolog. In co-transfected HEK293T cells, human Salvador increased the levels of Mst2 autophosphorylation and Mst2-mediated phosphorylation of select substrates, whereas Salvador SARAH domain inhibited Mst2 autophosphorylation in vitro. These results suggest Salvador enhances the effects of Hippo kinase activity at multiple points in the Hippo pathway.
AB - The Hippo pathway controls cell proliferation and differentiation through the precisely tuned activity of a core kinase cassette. The activity of Hippo kinase is modulated by interactions between its C-terminal coiled-coil, termed the SARAH domain, and the SARAH domains of either dRassF or Salvador. Here, we wanted to understand the molecular basis of SARAH domain–mediated interactions and their influence on Hippo kinase activity. We focused on Salvador, a positive effector of Hippo activity and the least well-characterized SARAH domain–containing protein. We determined the crystal structure of a complex between Salvador and Hippo SARAH domains from Drosophila. This structure provided insight into the organization of the Salvador SARAH domain including a folded N-terminal extension that expands the binding interface with Hippo SARAH domain. We also found that this extension improves the solubility of the Salvador SARAH domain, enhances binding to Hippo, and is unique to Salvador. We therefore suggest expanding the definition of the Salvador SARAH domain to include this extended region. The heterodimeric assembly observed in the crystal was confirmed by cross-linked MS and provided a structural basis for the mutually exclusive interactions of Hippo with either dRassF or Salvador. Of note, Salvador influenced the kinase activity of Mst2, the mammalian Hippo homolog. In co-transfected HEK293T cells, human Salvador increased the levels of Mst2 autophosphorylation and Mst2-mediated phosphorylation of select substrates, whereas Salvador SARAH domain inhibited Mst2 autophosphorylation in vitro. These results suggest Salvador enhances the effects of Hippo kinase activity at multiple points in the Hippo pathway.
UR - http://www.scopus.com/inward/record.url?scp=85045379960&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85045379960&partnerID=8YFLogxK
U2 - 10.1074/jbc.RA117.000923
DO - 10.1074/jbc.RA117.000923
M3 - Article
C2 - 29519817
AN - SCOPUS:85045379960
VL - 293
SP - 5532
EP - 5543
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 15
ER -