Gatekeeper-Activation Loop Cross-Talk Determines Distinct Autoactivation States of Symbiosis Receptor Kinase

Bhattacharya, Avisek and Paul, Anindita and Chakrabarti, Dipanjan and DasGupta, Maitrayee (2019) Gatekeeper-Activation Loop Cross-Talk Determines Distinct Autoactivation States of Symbiosis Receptor Kinase. In: BIOCHEMISTRY, 58 (19). pp. 2419-2431.

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Abstract

Plant receptor-like kinases (RLKs) have a Tyr in the ``gatekeeper'' position adjacent to the hinge region. The gatekeeper is phosphorylated in several RLKs, including symbiosis receptor kinase (SYMRK), but the significance of this remains unknown. Gatekeeper substitution did not inactivate Arachis hypogaea SYMRK but affected autophosphorylation at selected sites. Herein, we show that nonphosphorylatable gatekeepers (Y670F and Y670A) restrict SYMRK to be a Ser/Thr kinase with a basal level of phosphorylation (similar to 5 P/polypeptide, termed state I) whereas phosphorylatable gatekeepers (Y670 and Y670T) allowed SYMRK to be dual specific (Ser/Thr/Tyr) with a maximal level of phosphorylation (similar to 10 P/polypeptide, termed state II). State II SYMRKs were phosphorylated on gatekeeper residues, and the phosphocode in their activation segment was distinct from state I. The k(cat)/K-m for substrate phosphorylation was similar to 10-fold higher for state II, though for autophosphorylation, it was comparable with those of state I SYMRKs. To identify other determinants of state I features, we mutagenized all nine sites where phosphorylation was affected by nonphosphorylatable gatekeepers (Y670F and Y670A). Only two such mutants, S754A and S757A, located on the activation loop failed to phosphorylate gatekeeper Tyr and restricted SYMRK in state I. Double mutants like Y670F/S754A retained the features of state I, but Y670F/S757A was significantly inactivated, indicating a nonphosphorylatable gatekeeper can bypass phosphorylation of 5754 but not 5757 in the activation segment. We propose a working model for the hierarchical phosphorylation of SYMRK on gatekeeper and activation segments for its pS757-mediated activation as a Ser/Thr kinase in selfie mode (autophosphorylation) to a pS754/pY670-mediated activation as a Ser/Thr/Tyr kinase that functions in dual mode (both autophosphorylation and substrate phosphorylation).

Item Type:	Journal Article
Publication:	BIOCHEMISTRY
Publisher:	AMER CHEMICAL SOC
Additional Information:	copyright for this article belongs to BIOCHEMISTRY
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	04 Jun 2020 09:51
Last Modified:	04 Jun 2020 09:51
URI:	http://eprints.iisc.ac.in/id/eprint/62873

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