title: Novel CCM1, CCM2, and CCM3 mutations in patients with cerebral cavernous malformations: in-frame deletion in CCM2 prevents formation of a CCM1/CCM2/CCM3 protein complex.
abstract: Cerebral cavernous malformations (CCM) are prevalent cerebrovascular lesions predisposing to chronic headaches, epilepsy, and hemorrhagic stroke. Using a combination of direct sequencing and MLPA analyses, we identified 15 novel and eight previously published CCM1 (KRIT1), CCM2, and CCM3 (PDCD10) mutations. The mutation detection rate was >90% for familial cases and >60% for isolated cases with multiple malformations. Splice site mutations constituted almost 20% of all CCM mutations identified. One of these proved to be a de novo mutation of the most 3' acceptor splice site of the CCM1 gene resulting in retention of intron 19. A further mutation affected the 3' splice site of CCM2 intron 2 leading to cryptic splice site utilization in both CCM2 and its transcript variant lacking exon 2. With the exception of one in-frame deletion of CCM2 exon 2, which corresponds to the naturally occurring splice variant of CCM2 on the RNA level and is predicted to result in the omission of 58 amino acids (CCM2:p.P11_K68del), all mutations lead to the introduction of premature stop codons. To gain insight into the likely mechanisms underlying the only known CCM2 in-frame deletion, we analyzed the functional consequences of loss of CCM2 exon 2. The CCM2:p.P11_K68del protein could be expressed in cell culture and complexed with CCM3. However, its ability to interact with CCM1 and to form a CCM1/CCM2/CCM3 complex was lost. These data are in agreement with a loss-of-function mechanism for CCM mutations, uncover an N-terminal CCM2 domain required for CCM1 binding, and demonstrate full-length CCM2 as the essential core protein in the CCM1/CCM2/CCM3 complex.
authors.affiliation: Department of Human Genetics, University of Würzburg, Würzburg, Germany.
authors.forename: Sonja
authors.initials: S
authors.lastname: Stahl
authors.forename: Sabine
authors.initials: S
authors.lastname: Gaetzner
authors.forename: Katrin
authors.initials: K
authors.lastname: Voss
authors.forename: Bettina
authors.initials: B
authors.lastname: Brackertz
authors.forename: Elisa
authors.initials: E
authors.lastname: Schleider
authors.forename: Oguzkan
authors.initials: O
authors.lastname: Sürücü
authors.forename: Ekkehard
authors.initials: E
authors.lastname: Kunze
authors.forename: Christian
authors.initials: C
authors.lastname: Netzer
authors.forename: Christoph
authors.initials: C
authors.lastname: Korenke
authors.forename: Ulrich
authors.initials: U
authors.lastname: Finckh
authors.forename: Mario
authors.initials: M
authors.lastname: Habek
authors.forename: Zdravka
authors.initials: Z
authors.lastname: Poljakovic
authors.forename: Miriam
authors.initials: M
authors.lastname: Elbracht
authors.forename: Sabine
authors.initials: S
authors.lastname: Rudnik-Schöneborn
authors.forename: Helmut
authors.initials: H
authors.lastname: Bertalanffy
authors.forename: Ulrich
authors.initials: U
authors.lastname: Sure
authors.forename: Ute
authors.initials: U
authors.lastname: Felbor
mesh_terms: D051017:Apoptosis Regulatory Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D017931:DNA Primers; D005260:Female; D020786:Hemangioma, Cavernous, Central Nervous System / Q000235:genetics*; D006801:Humans; D000075927:KRIT1 Protein; D008297:Male; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D010375:Pedigree; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion
chemical_list: D051017:Apoptosis Regulatory Proteins; C502544:CCM2 protein, human; D002352:Carrier Proteins; D017931:DNA Primers; D000075927:KRIT1 Protein; C107928:KRIT1 protein, human; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; C495390:PDCD10 protein, human; D011518:Proto-Oncogene Proteins