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Structural insights into the negative regulation of BRI1 signaling by BRI1-interacting protein BKI1.

ABSTRACT: Brassinosteroids (BRs) are essential steroid hormones that have crucial roles in plant growth and development. BRs are perceived by the cell-surface receptor-like kinase brassinosteroid insensitive 1 (BRI1). In the absence of BRs, the cytosolic kinase domain (KD) of BRI1 is inhibited by its auto-inhibitory carboxyl terminus, as well as by interacting with an inhibitor protein, BRI1 kinase inhibitor 1 (BKI1). How BR binding to the extracellular domain of BRI1 leads to activation of the KD and dissociation of BKI1 into the cytosol remains unclear. Here we report the crystal structure of BRI1 KD in complex with the interacting peptide derived from BKI1. We also provide biochemical evidence that BRI1-associated kinase 1 (BAK1) plays an essential role in initiating BR signaling. Steroid-dependent heterodimerization of BRI1 and BAK1 ectodomains brings their cytoplasmic KDs in the right orientation for competing with BKI1 and transphosphorylation.

SUBMITTER: Wang J

PROVIDER: S-EPMC4220157 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Structural insights into the negative regulation of BRI1 signaling by BRI1-interacting protein BKI1.

Wang Jie J Jiang Jianjun J Wang Jue J Chen Lei L Fan Shi-Long SL Wu Jia-Wei JW Wang Xuelu X Wang Zhi-Xin ZX

Cell research 20141021 11

Brassinosteroids (BRs) are essential steroid hormones that have crucial roles in plant growth and development. BRs are perceived by the cell-surface receptor-like kinase brassinosteroid insensitive 1 (BRI1). In the absence of BRs, the cytosolic kinase domain (KD) of BRI1 is inhibited by its auto-inhibitory carboxyl terminus, as well as by interacting with an inhibitor protein, BRI1 kinase inhibitor 1 (BKI1). How BR binding to the extracellular domain of BRI1 leads to activation of the KD and dis ...[more]

PMID: 25331450

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Project description:Cannabinoid CB1 receptors (CB1Rs) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid receptor-interacting protein 1a (CRIP1a) binds to the CB1R C-terminus and can attenuate constitutive CB1R-mediated inhibition of Ca(2+) channel activity. We now demonstrate cellular colocalization of CRIP1a at neuronal elements in the CNS and show that CRIP1a inhibits both constitutive and agonist-stimulated CB1R-mediated guanine nucleotide-binding regulatory protein (G-protein) activity. Stable overexpression of CRIP1a in human embryonic kidney (HEK)-293 cells stably expressing CB1Rs (CB1-HEK), or in N18TG2 cells endogenously expressing CB1Rs, decreased CB1R-mediated G-protein activation (measured by agonist-stimulated [(35)S]GTP?S (guanylyl-5'-[O-thio]-triphosphate) binding) in both cell lines and attenuated inverse agonism by rimonabant in CB1-HEK cells. Conversely, small-interfering RNA-mediated knockdown of CRIP1a in N18TG2 cells enhanced CB1R-mediated G-protein activation. These effects were not attributable to differences in CB1R expression or endocannabinoid tone because CB1R levels did not differ between cell lines varying in CRIP1a expression, and endocannabinoid levels were undetectable (CB1-HEK) or unchanged (N18TG2) by CRIP1a overexpression. In CB1-HEK cells, 4-hour pretreatment with cannabinoid agonists downregulated CB1Rs and desensitized agonist-stimulated [(35)S]GTP?S binding. CRIP1a overexpression attenuated CB1R downregulation without altering CB1R desensitization. Finally, in cultured autaptic hippocampal neurons, CRIP1a overexpression attenuated both depolarization-induced suppression of excitation and inhibition of excitatory synaptic activity induced by exogenous application of cannabinoid but not by adenosine A1 agonists. These results confirm that CRIP1a inhibits constitutive CB1R activity and demonstrate that CRIP1a can also inhibit agonist-stimulated CB1R signaling and downregulation of CB1Rs. Thus, CRIP1a appears to act as a broad negative regulator of CB1R function.

Dataset Information

Structural insights into the negative regulation of BRI1 signaling by BRI1-interacting protein BKI1.

Publications

Structural insights into the negative regulation of BRI1 signaling by BRI1-interacting protein BKI1.

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