Crystallographic Studies of the Cerebral Cavernous Malformations Proteins
- 340 Downloads
Cerebral cavernous malformations (CCM) are dysplasias that primarily occur in the neurovasculature, and are associated with mutations in three genes: KRIT1, CCM2, and PDCD10, the protein products of which are KRIT1 (Krev/Rap1 Interaction Trapped 1; CCM1, cerebral cavernous malformations 1), CCM2 (cerebral cavernous malformations 2; OSM, osmosensing scaffold for MEKK3), and CCM3 (cerebral cavernous malformations 3; PDCD10, programmed cell death 10). Until recently, these proteins were relatively understudied at the molecular level, and only three folded domains were documented. These were a band 4.1, ezrin, radixin, moesin (FERM), and an ankyrin repeat domain (ARD) in KRIT1, and a phosphotyrosine-binding (PTB) domain in CCM2. Over the past 10 years, a crystallographic approach has been used to discover a series of previously unidentified domains within the CCM proteins. These include a non-functional Nudix (or pseudonudix) domain in KRIT1, a harmonin homology domain (HHD) in CCM2, and dimerization and focal adhesion targeting (FAT)-homology domains within CCM3. Many of the roles of these domains have been revealed by structure-guided studies that show the CCM proteins can directly interact with one another to form a signaling scaffold, and that the “CCM complex” functions in signal transduction by interacting with other binding partners, including ICAP1, RAP1, and MEKK3. In this chapter, we describe the crystallization of CCM protein domains alone, and with their interaction partners.
Key wordsCCM proteins X-ray crystallography Protein purification KRIT1 CCM2 CCM3 ICAP1 RAP1 MEKK3
We wish to thank Amy Stiegler Wyler, Byunghak Ha, and David Calderwood for helpful discussions. This work is funded by National Institutes of Health grants R01GM114621 and R01NS085078 to TJB.
- 7.Fisher OS, Liu W, Zhang R, Stiegler AL, Ghedia S, Weber JL, Boggon TJ (2015) Structural basis for the disruption of the cerebral cavernous malformations 2 (CCM2) interaction with Krev interaction trapped 1 (KRIT1) by disease-associated mutations. J Biol Chem 290(5):2842–2853. https://doi.org/10.1074/jbc.M114.616433CrossRefPubMedGoogle Scholar
- 15.Gingras AR, Puzon-McLaughlin W, Ginsberg MH (2013) The structure of the ternary complex of Krev Interaction Trapped 1 (KRIT1) bound to both the Rap1 GTPase and the Heart of Glass (HEG1) cytoplasmic tail. J Biol Chem 288(33):23639–23649. https://doi.org/10.1074/jbc.M113.462911CrossRefPubMedPubMedCentralGoogle Scholar
- 18.Wang X, Ding J, Wang D (2012) Crystallization and preliminary X-ray analysis of the C-terminal domain of CCM2, part of a novel adaptor protein involved in cerebral cavernous malformations. Acta Crystallogr Sect F Struct Biol Cryst Commun 68(Pt 6):683–686. https://doi.org/10.1107/S1744309112016181CrossRefPubMedPubMedCentralGoogle Scholar
- 19.Ding J, Wang X, Li DF, Hu Y, Zhang Y, Wang DC (2010) Crystal structure of human programmed cell death 10 complexed with inositol-(1,3,4,5)-tetrakisphosphate: a novel adaptor protein involved in human cerebral cavernous malformation. Biochem Biophys Res Commun 399(4):587–592CrossRefPubMedGoogle Scholar
- 24.Liquori CL, Berg MJ, Siegel AM, Huang E, Zawistowski JS, Stoffer T, Verlaan D, Balogun F, Hughes L, Leedom TP, Plummer NW, Cannella M, Maglione V, Squitieri F, Johnson EW, Rouleau GA, Ptacek L, Marchuk DA (2003) Mutations in a gene encoding a novel protein containing a phosphotyrosine-binding domain cause type 2 cerebral cavernous malformations. Am J Hum Genet 73(6):1459–1464CrossRefPubMedPubMedCentralGoogle Scholar
- 25.Denier C, Goutagny S, Labauge P, Krivosic V, Arnoult M, Cousin A, Benabid AL, Comoy J, Frerebeau P, Gilbert B, Houtteville JP, Jan M, Lapierre F, Loiseau H, Menei P, Mercier P, Moreau JJ, Nivelon-Chevallier A, Parker F, Redondo AM, Scarabin JM, Tremoulet M, Zerah M, Maciazek J, Tournier-Lasserve E (2004) Mutations within the MGC4607 gene cause cerebral cavernous malformations. Am J Hum Genet 74(2):326–337CrossRefPubMedPubMedCentralGoogle Scholar
- 27.Bergametti F, Denier C, Labauge P, Arnoult M, Boetto S, Clanet M, Coubes P, Echenne B, Ibrahim R, Irthum B, Jacquet G, Lonjon M, Moreau JJ, Neau JP, Parker F, Tremoulet M, Tournier-Lasserve E (2005) Mutations within the programmed cell death 10 gene cause cerebral cavernous malformations. Am J Hum Genet 76(1):42–51CrossRefPubMedGoogle Scholar