Clinical and molecular characterization of rare malformations of cortical development - CRADD mutations as a cause of a novel recessive form of lissencephaly

• Applicant: Professorin Dr. Nataliya Di Donato
• Subject Area: Human Genetics
• Term: from 2014 to 2016
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 263956286

Malformations of cortical development (MCD) are a common cause of intellectual disability and epilepsy. In our initial project we focused on identifying new causative genes for classic disorders of neuronal migration such as lissencephaly (LIS).As part of this work, we recognized a novel anterior predominant form of undulating or thin LIS with reduced number of gyri and shallow sulci (cortex 5-7 mm; normal 3-4 mm, classic LIS 12-20 mm). Unlike most other types of LIS, which are caused by X-linked or de novo heterozygous mutations, this group of LIS appears to have exclusively autosomal recessive inheritance. Using whole exome sequencing we have identified two different homozygous missense mutations in the CRADD gene in four patients from two unrelated families. Intriguingly, the encoded protein, designated RAIDD, does not belong to the microtubule-associated or cytoskeletal protein families frequently associated with LIS and other neuronal migration disorders. Rather, the only known function of CRADD is to induce apoptosis by recruiting caspase-2. The presence of biallelic mutations in CRADD should decrease caspase-2-mediated apoptosis. The consequences of reduced apoptosis in brain have been demonstrated in animal models such as loss of caspase-9 (with large brain size), but never in humans. Therefore, elucidation of the functional consequences of mutations in CRADD may not only provide a cause for a novel form of LIS, but give new insight into regulation of apoptosis during embryonic cortical development. Because both CRADD mutations are homozygous, we propose that the mutations lead to loss of CRADD function. We plan to model in vitro loss of function by viral shRNA-mediated knockdown of Cradd in mouse neural progenitor cell lines. Use of a Sox2Cre transgenic line crossed with a Cre reporter strain (Ai14) will allow easy visualization, selection and tracking of neural progenitor cells. We will estimate the apoptosis rate and also evaluate migration in the Cradd knockdown cell culture, as the association with LIS suggests an additional defect in neuronal migration. A collaborator will simultaneously perform similar experiments in patient-derived fibroblasts. Moreover, we will perform a volumetric MRI analysis and examine brain histology of the Cradd-/- mouse (Cradd tm1Mak). We believe that demonstration of the consequences of decreased apoptosis during neuronal development in humans represents a novel mechanism for human developmental disorders. The host institution - the Dobyns lab in Seattle Childrens Research Institute - has all necessary resources to complete the experiments within the requested 4 months. Some will be performed with assistance from the adjacent lab of Prof. Kathleen Millen. This combination of resources is not readily available elsewhere.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Professor Dr. William Dobyns