Diverse Roles of Phosphatidylinositol Transfer Protein PITPβ in Eukaryotic Cells Public Deposited

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  • March 22, 2019
Creator
  • Ile, Kristina E.
    • Affiliation: School of Medicine, Department of Cell Biology and Physiology
Abstract
  • Phosphatidylinositol transfer proteins (PITPs) provide a dynamic interface between membrane dynamics and lipid signaling. PITPα and PITPβ encode for two soluble class I, metazoan specific PITPs. However, despite the fact that PITPα and PITPβ share 77 and 95% primary sequence identity and similarity (respectively), PITPα and PITPβ are functionally distinct and play nonredundant functions in cells. Whereas PITPα knockout mice have been created, and these mice display neuronal, intestinal, and liver pathologies, PITPβ functionality remains largely unknown. In this work, I addressed the distinguishing properties of PITPβ and its role in mammalian and zebrafish cells. We identified regions of PITPβ that contributed to its unique properties: Golgi localization and sphingomyelin (SM) binding. Two single point mutations are sufficient to ablate the SM binding properties of PITPβ, and the Golgi localization was attributed to the combination of two regions of the protein. Next, siRNA and morpholino analyses, in mammalian cells and zebrafish respectively, were developed to understand the physiological role of PITPβ in cells. In mammalian cells, PITPβ was essential for cell survival. Particularly, PITPβ-depleted cells displayed disorganized Golgi networks and anterograde trafficking defects. Surprisingly, PITPβ was not required for cell survival or early development in zebrafish. Instead, PITPβ maintains the outer segment of the double cone cells of the eye - a cell type required for detection of red and green light. The lack of a housekeeping role for PITPβ led us to examine the roles of other PITPs in zebrafish. A novel third soluble PITP in zebrafish, PITPγ, was identified, though its function is not yet determined because we have no means of detecting the protein levels to ascertain knockdown. PITPα, however, had an unexpected role in early development: reduction of PITPα led to an arrest at an early stage of development (about 12 hpf), ultimately leading to embryonic lethality. Together, this dissertation presents novel roles for class I PITPs in zebrafish, and represents the first functional characterization of PITPβ.
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  • In Copyright
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  • "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the School of Medicine (Cell and Developmental Biology)."
Advisor
  • Bankaitis, Vytas A.
Degree
  • Doctor of Philosophy
Graduation year
  • 2009
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