Theses Doctoral

Genetic and Infectious Causes of Microcephaly: NDE1 Mutations Compared to the Zika Virus

Doobin, David J.

Brain development is an exquisitely coordinated process of progenitor cell proliferation followed by the migration of progeny to their final location in the developing brain. There are a myriad of points at which this process can be disturbed, and the examination of these perturbations help us further understand basic science, as well as epidemics sweeping through the world around us. Microcephaly, which is defined as a head circumference greater than 2 standard deviations below the mean, can occur through genetic, infectious, vascular, or metabolic etiologies, and the studies herein examine two forms by which microcephaly occurs. First, we investigate the role of the dynein regulatory protein Nde1 in the development of the neocortex, which is the outer region of the forebrain. NDE1 mutations are associated with severe microcephaly, and we find that unlike most microcephaly genes whose products have one role in the cell cycle, Nde1 is required at three discrete points in neuronal progenitors, termed radial glia progenitors (RGPs). We initially find that Nde1 is required to recruit dynein to the nuclear envelope to allow for interkinetic nuclear migration (INM) during G2. Additionally, Nde1 helps to initiate primary cilia resorption at the G1-to-S transition. Finally, there is a necessity for Nde1 at the G2-to-M transition after the completion of INM and prior to nuclear envelope breakdown. These three distinct roles for Nde1 illustrate the breadth of functions that the protein has during RGP proliferation, and help to explain why patients with NDE1 mutations have such severe microcephaly.
As this work was ongoing there was a global outbreak of a new pathogen that had previously been dormant throughout Africa and Asia, only to emerge at epidemic proportions in the Western Hemisphere. This pathogen, the Zika Virus (ZIKV), is particularly alarming because of its subclinical course in adults but devastating consequences for fetal development, with the hallmark symptom being microcephaly. Using our organotypic brain slice model system, we demonstrate the ability of a variety of ZIKV isolates to infect and replicate in embryonic brain tissue. All ZIKV isolates that infect the organotypic slices lead to increases in apoptosis, though these increases are particularly pronounced in isolates from the Asian/American lineages. Notably, one isolate from a patient in Nigeria (termed 30656) does not replicate in mouse neuronal tissue, but electroporation of the 30656 ZIKV genome allows for a single cycle replication, suggesting that this isolate is unable to enter RGPs. All infectious isolates are pathogenic in early- and mid- gestation embryonic tissue, but only one isolate infects and replicates in late- gestation embryonic tissue. This was the most recently isolated sample tested, and it demonstrates a predilection for neurons, suggesting that ZIKV may be mutating as it spreads. These results provide foundational insight into the pathogenesis of ZIKV- associated microcephaly, and illustrate how studies of genetic forms of microcephaly can enhance and facilitate our understanding of infectious causes of the disease.


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More About This Work

Academic Units
Neurobiology and Behavior
Thesis Advisors
Vallee, Richard
Ph.D., Columbia University
Published Here
January 17, 2017