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University of North Carolina

1. Tribble, Emily K. On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol.

Degree: Cell Biology and Physiology, 2011, University of North Carolina

Lipid metabolism within the nuclear matrix of mammalian cells is robust. Numerous lipid-signaling pathway components are found within purified nuclear fractions, depleted of nuclear envelope and cellular debris using detergents ("endonuclear" fractions). Interestingly, membrane-stripped nuclei also contain measurable quantities of the substrate lipids, notably phospholipids (PLs), despite the absence of detectable bilayer structures. According to published work, these PLs are i) major constituents of the endonuclear space (10-16% by volume), ii) distinct from cytosolic/cellular lipids in molecular species profiles and relative abundance, and iii) generated or consumed in response to physiological cues. The abundance and diversity of PL molecular species in nuclei suggests endonuclear lipids not only signal within the nuclear space, but also impact nuclear structure and function on the whole. That the elements of functional endonuclear phosphoinositide signaling pathways (enzymes and PLs) are found within nuclei has received much attention, as their cytosolic counterparts regulate numerous cellular events. While new evidence suggests that nuclear-generated phosphoinositides participate as cofactors in essential nuclear processes, there are still unanswered questions regarding the regulation, location, and organization of endonuclear phosphoinositides. Phosphatidylinositol, the PL from which all phosphoinositides are generated, enters the nuclear space via an unknown mechanism. This dissertation originated in testing a candidate importer protein, the phosphatidylinositol transfer protein alpha, in the nuclear supply of Phosphatidylinositol. Such experiments require envelope-stripped nuclei that meet high purity standards. Initial attempts at purifying nuclei via published protocols were unsuccessful, and a new method for purifying nuclei from mouse embryonic fibroblasts for use in this context is described. Nuclei purified according to the new protocol meet an expanded, quantitative quality control suite. These highly purified nuclei contain several orders of magnitude less PL than previously reported in LC/MS/MS mass analyses. In addition, pulse-labeling and comparative dynamic lipidomic studies of mouse embryonic fibroblast nuclei, either wild type or genetically knocked-out for Phosphatidylinositol transfer protein alpha, purified with this new protocol demonstrated that our candidate shuttle protein was not an obligate requirement in the import of Phosphatidylinositol. Overall, this dissertation demonstrates that proper execution of nuclear preparations is an essential component of studies of endonuclear lipid regulation. Advisors/Committee Members: Tribble, Emily K., Bankaitis, Vytas A..

Subjects/Keywords: School of Medicine; Department of Cell Biology and Physiology

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APA (6th Edition):

Tribble, E. K. (2011). On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol. (Thesis). University of North Carolina. Retrieved from https://cdr.lib.unc.edu/record/uuid:689ebf04-478a-42f7-a946-ac62b546ce3e

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

Chicago Manual of Style (16th Edition):

Tribble, Emily K. “On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol.” 2011. Thesis, University of North Carolina. Accessed November 25, 2020. https://cdr.lib.unc.edu/record/uuid:689ebf04-478a-42f7-a946-ac62b546ce3e.

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

MLA Handbook (7th Edition):

Tribble, Emily K. “On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol.” 2011. Web. 25 Nov 2020.

Vancouver:

Tribble EK. On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol. [Internet] [Thesis]. University of North Carolina; 2011. [cited 2020 Nov 25]. Available from: https://cdr.lib.unc.edu/record/uuid:689ebf04-478a-42f7-a946-ac62b546ce3e.

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

Council of Science Editors:

Tribble EK. On the role of the phosphatidylinositol transfer protein alpha in the nuclear import of phosphatidylinositol. [Thesis]. University of North Carolina; 2011. Available from: https://cdr.lib.unc.edu/record/uuid:689ebf04-478a-42f7-a946-ac62b546ce3e

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

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