C The Creamer Lab
Research in the Creamer Lab
Calcineurin
The primary focus of the Creamer laboratory is on the function and regulation of the serine/threonine phosphatase calcineurin (CaN). CaN is ubiquitously expressed and highly conserved in all eukaryotes. As an integral component of several signaling pathways, this enzyme dephosphorylates, and thereby regulates, a number of important proteins such as the NFAT family of transcription activators, the microtubule-associated tau, and nitric oxide synthase. CaN plays central roles in neuronal signaling, cardiac growth and immune system activation. Dysregulation of CaN contributes to a number of disorders including Alzheimer’s disease, Down syndrome, mental retardation, cardiac hypertrophy, and autoimmune diseases.
CaN is a heterodimer with an ~58-66kDa A chain and a 19kDa B chain. There are three isoforms of the A and two of the B chain. These combine to give three variants. The α variant is expressed primarily in neurons, β is ubiquitously expressed and γ is expressed solely in testis. The A chain possesses catalytic, regulatory and autoinhibitory domains. The B chain, which appears to be important for structural reasons, is highly homologous to the calcium binding protein calmodulin. CaN activity is tightly regulated by a number of other proteins. Calmodulin (CaM) binds to and activates CaN. Other proteins, including Rcan1, CHP1 and cabin/cain, are endogenous inhibitors of CaN. Yet others, such as AKAP79, serve to localize CaN to specific regions within a cell. Breakdowns in any of these regulatory systems can lead to pathologies associated with the diseases listed above. For example, Rcan1 is overexpressed as a consequence of the trisomy underlying Down syndrome. This in turn leads to inappropriate inhibition of CaN. Notably, the CaN regulatory mechanisms are poorly understood at the molecular/structural level.Ongoing projects-
1) Upon an increase in cellular calcium levels, CaM binds to and activates CaN. CaM binds to a regulatory domain within CaN which leads to a conformational change that in turn ejects an autoinhibitory domain from CaN’s active site. The regulatory domain of CaN appears to be devoid of structure prior to CaM binding. In other words this domain is intrinsically disordered. Upon CaM binding the CaN regulatory domain gains structure. We are currently studying this disorder-to-order transition and how it leads to CaN activity. 2) Rcan1 (regulator of calcineurin 1) is an endogenous inhibitor of CaN that has been associated with Down syndrome and Alzheimer’s disease. This protein is known to bind to the catalytic domain of CaN, blocking access to the active site. Rcan1 possesses two domains: a well-structured domain of unknown function, and the CaN binding domain which appears to be completely unstructured. We therefore have an enzyme with an intrinsically disordered regulatory domain being inhibited by a protein with an intrinsically disordered inhibitory domain. Rcan1 binding to CaN is a potential drug target in Alzheimer’s and Down syndrome, but little is known regarding the details of the interactions between these proteins. We are studying this system in order to gain an understanding of the structural details of these interactions. 3) CHP1 (calcineurin B homology protein 1) is another endogenous inhibitor of CaN. What is remarkable about this protein is that, like the CaN B chain, it is structurally homologous to CaM. CaN is thus regulated by three proteins with the same structure, but with three different roles. CaM activates, the CaN B chain has a structural role, and CHP1 inhibits. Other than the fact that it inhibits CaN, little is known about how CHP1 functions. Our initial data suggests that it might bind to the same region of the regulatory domain as CaM, but does not induce a disorder-to-order transition, thereby leaving the autoinhibitory domain bound in CaN’s active site.Calmodulin
Calmodulin binds to and regulates a great many proteins including calcineurin. It has been suggested that calmodulin binds to disordered regioins within these proteins. Consequently we are interested in other calmodulin targets.