Cooperative autoinhibition and multi-level activation mechanisms of calcineurin

Abstract

The Ca 2+ /calmodulin-dependent protein phosphatase calcineurin (CN), a heterodimer composed of a catalytic subunit A and an essential regulatory subunit B, plays critical functions in various cellular processes such as cardiac hypertrophy and T cell activation. It is the target of the most widely used immunosuppressants for transplantation, tacrolimus (FK506) and cyclosporin A. However, the structure of a large part of the CNA regulatory region remains to be determined, and there has been considerable debate concerning the regulation of CN activity. Here, we report the crystal structure of full-length CN (β isoform), which revealed a novel autoinhibitory segment (AIS) in addition to the well-known autoinhibitory domain (AID). The AIS nestles in a hydrophobic intersubunit groove, which overlaps the recognition site for substrates and immunosuppressant-immunophilin complexes. Indeed, disruption of this AIS interaction results in partial stimulation of CN activity. More importantly, our biochemical studies demonstrate that calmodulin does not remove AID from the active site, but only regulates the orientation of AID with respect to the catalytic core, causing incomplete activation of CN. Our findings challenge the current model for CN activation, and provide a better understanding of molecular mechanisms of CN activity regulation.

Investigating the human Calcineurin Interaction Network using the πɸLxVP SLiM

“…1B). This is the same pocket occupied by LxVP-containing inhibitors1318. As expected, the NFATc1 residues that interact most extensively with CN are Leu387 NFATc1 and Val389 NFATc1 .…”

Calcineurin promotes APC/C activation at meiotic exit by acting on both XErp1 and Cdc20

“…In the absence of calmodulin, pSer335 was weakly dephosphorylated by CaN confirming previous reports [29] that CaN even in the absence of its co-activator has some basal activity. Studies on mammalian Emi2 have suggested that phosphorylation of Ser385 (Ser335 in XErp1) increases the affinity for B 0 56 roughly 2,6-fold [25].…”

Calcineurin signaling in the heart: The importance of time and place

“…1A). Binding of Ca 2+ to the regulatory sites initiates a series of conformational changes that allow binding of a calmodulin/Ca 2+ complex and a change in the orientation of the AID to expose the active site [13, 14] (Fig. 1B).…”

“…PxIxIT domains vary in their binding affinities, allowing for substrate selection based both on concentration and binding strength. The second docking motif, LxVP, binds to a hydrophobic pocket at the CnA/CnB interface, which is only accessible when calcineurin is active [14, 56]. An alternative model for LxVP binding has been proposed in which the LxVP docking site overlaps with the PxIxIT docking site [57].…”