Microtubules are assembled from ?/? tubulin heterodimers. In mammals, a small multigene family encodes ?- and ?-tubulins, with each gene product (termed an isotype) having a distinct developmentally regulated and tissue-specific pattern of expression. Recently, mutations have been identified in genes (e.g. TUBA1A and TUBB2B) encoding these isotypes that cause certain human neuronal migration disorders, all of which are associated with severe cognitive disabilities. These discoveries reinforce the notion that microtubule based events play a central role in neuronal migration during brain development, and that disruption of critical microtubule processes results in cortical dysgeneses. Analysis of the properties of disease causing mutations in TUBA1A suggests two broad classes of mechanism: 1. Defects in the complex tubulin heterodimer assembly pathway This pathway involves sequential interactions of newly synthesized ?- and ?-tubulin polypeptides with multiple chaperone proteins (including the cytosolic chaperonin CCT and five tubulin specific chaperones, TBCA-TBCE). CCT facilitates productive folding by providing a sequestered environment in which folding can occur in the absence of off-pathway interactions that might otherwise lead to aggregation, while the tubulin specific chaperones function downstream of CCT as an ?/? tubulin heterodimer assembly machine. 2. Defective microtubule dynamics and/or interactions with Microtubule Associated Proteins (MAPs) Disease causing mutations might compromise microtubule dynamics or interfere with interaction(s) between microtubules and MAPs that are critical for directing proper neuronal migration. The experiments proposed here constitute a multifaceted approach towards understanding the mechanism of these diseases. 1) We will exploit the mutation-induced defective interactions between CCT-generated ?-tubulin folding intermediates and TBCB to define this interaction. These experiments will establish the mechanism of action of TBCB as a critical player in proper corticogenesis. 2) We will generate populations of isotypically homogeneous wild type and mutant tubulin heterodimers. These will be used to identify MAPs (such as the microtubule polymerase TOGp) that bind differently to microtubules polymerized from wild type and disease-causing mutant heterodimers. 3) For those disease-causing tubulin mutations that do not interfere with the heterodimer assembly machinery, we will examine their effect on microtubule dynamics in single cell experiments (including cultured neurons) in vivo. 4) The mechanism of disease will be explored via the construction and analysis of transgenic mice in which one copy of wild type tuba1a (the mouse homolog of TUBA1A) is replaced with the same allele harboring a disease-causing mutation. The brains of these mice will be examined in terms of their development and the behavior of their microtubules in vitro and in vivo.

Public Health Relevance

We will establish the molecular basis of naturally occurring neuronal migration diseases caused by mutations in the ?-tubulin gene TUBA1A. We will use structural, biochemical, cell biological and animal model approaches in order to understand the significance of these mutations in terms of the development of the mammalian brain.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM097376-02
Application #
8517751
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Gindhart, Joseph G
Project Start
2012-08-01
Project End
2017-05-31
Budget Start
2013-06-01
Budget End
2015-05-31
Support Year
2
Fiscal Year
2013
Total Cost
$460,586
Indirect Cost
$188,854
Name
New York University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
121911077
City
New York
State
NY
Country
United States
Zip Code
10016
Poirier, Karine; Lebrun, Nicolas; Broix, Loic et al. (2013) Mutations in TUBG1, DYNC1H1, KIF5C and KIF2A cause malformations of cortical development and microcephaly. Nat Genet 45:639-47