Microcephaly (abnormally small brain) is a neurodevelopmental disorder that causes neurological symptoms, such as intellectual disability, language delay, and epilepsy. A number of causative genes have been reported, the majority of which encode centrosomal proteins. Exactly how mutations in centrosomal proteins cause microcephaly is not well understood. Previous studies using fly and mouse models suggest that mutations in centrosomal proteins may disrupt proliferation of neural progenitor cells (NPCs) or induce premature differentiation into neurons at the expense of NPCs. However, currently available animal models of microcephaly have pretty mild phenotypes, making it hard to address which molecular and cellular mechanisms are critical to severe microcephaly in humans. This project seeks to develop and establish new animal models for microcephaly with robust phenotypes. We have two hypotheses: (1) because many microcephaly gene products colocalize in the centrosome, some of them may interact biochemically and genetically. For example, ASPM (abnormal spindle-like, microcephaly-associated) and WDR62 (WD repeat domain 62), the two most common causes for human microcephaly when mutated, interact with each other. Thus, heterozygous deletion of one microcephaly gene, which has no phenotype at all by itself, may enhance the mild phenotype in homozygous knockout mice of another microcephaly gene; (2) Unlike mice, ferrets have an enlarged brain, which contains outer radial glial cells, a type of NPCs that is highly abundant in the human cortex. Thus, knockout ferrets of a microcephaly gene may show robust phenotypes compared to knockout mice of the same gene.
In Aim 1, we will examine Aspm-/-; Wdr62+/- mice that have a significantly smaller brain than any control mice (Aspm+/+; Wdr62+/- and Aspm-/-; Wdr62+/+ mice), which have negligible phenotypes. We will characterize interaction between the two proteins as well as asymmetric inheritance of mother versus daughter centrosomes in the developing cortex of Aspm-/-; Wdr62+/- mice.
In Aim 2, we will establish and characterize Aspm-knockout ferrets that we have recently generated using TALEN, a new genome-editing tool. Preliminary data show that they have severe microcephaly. Using immunohistochemistry and adenoviral green fluorescent protein infection followed by time-lapse imaging, we will examine abundance and behaviors of diverse NPCs in Aspm-knockout and wile-type ferrets. The proposed work will provide exciting new animal models for microcephaly and cortical malformation in general, creating an innovative experimental system in the field of cerebral cortical development and evolution. With ever-increasing list of genes from human genetic studies, our approach will demonstrate how to study functional meanings of a gene when knockout mice of the gene do not show robust phenotypes. In addition, it should resolve the current debate over roles of outer radial glial cell during cerebral cortical development, and has the potential to identify novel mechanisms of normal cortical development.

Public Health Relevance

Microcephaly ('abnormally small brain') is a neurodevelopmental disorder causing multiple symptoms afflicting human mental health, such as epilepsy and intellectual disability. Although causative genes have been identified, molecular and cellular mechanisms of microcephaly are not well understood because currently available animal models do not show robust microcephaly phenotypes. This project will develop an enhanced mouse model and a ferret model of microcephaly using a new genome-editing tool, thereby providing new ways to investigate molecular control of brain size.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Exploratory/Developmental Grants (R21)
Project #
7R21NS091865-02
Application #
9170447
Study Section
Developmental Brain Disorders Study Section (DBD)
Program Officer
Riddle, Robert D
Project Start
2015-10-22
Project End
2017-02-28
Budget Start
2015-10-22
Budget End
2016-02-29
Support Year
2
Fiscal Year
2015
Total Cost
Indirect Cost
Name
Yale University
Department
Neurosurgery
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
Johnson, Matthew B; Sun, Xingshen; Kodani, Andrew et al. (2018) Aspm knockout ferret reveals an evolutionary mechanism governing cerebral cortical size. Nature 556:370-375
Jayaraman, Divya; Kodani, Andrew; Gonzalez, Dilenny M et al. (2016) Microcephaly Proteins Wdr62 and Aspm Define a Mother Centriole Complex Regulating Centriole Biogenesis, Apical Complex, and Cell Fate. Neuron 92:813-828
Doan, Ryan N; Bae, Byoung-Il; Cubelos, Beatriz et al. (2016) Mutations in Human Accelerated Regions Disrupt Cognition and Social Behavior. Cell 167:341-354.e12