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The Ohio State University

1. Groves, Norman R. Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>.

Degree: PhD, Molecular Genetics, 2019, The Ohio State University

Proteins residing at the inner nuclear membrane (INM) in animals and yeast play a variety of roles within the cell, including nuclear organization, nuclear movement and chromatin organization. INM proteins in animals and yeast have been shown to have a variety of topologies and structures. Over 100 nuclear envelope transmembrane proteins (NETs) have been shown to be enriched at nuclear membranes, but have yet to be tested for INM localization. Amongst known INM proteins in animals and yeast, only SUN proteins are encoded for in plant genomes. SUN proteins form the INM component of linker of nucleoskeleton and cytoskeleton (LINC) complexes and are conserved from animals and yeast to plants. Besides SUN proteins, cation channels have been identified that localize to the INM, in the form of <i>Medicago truncatula</i> DMI1 and CNGC15. Given the lack of homologs of mammalian INM proteins in plants, proteins likely exist that are structurally divergent from animal and yeast INM proteins, but functionally serve similar roles within the cell.In both plants and animals, protein trafficking to the INM is poorly understood relative to other protein targeting pathways. One model for protein trafficking to the INM is transport factor-mediated trafficking, in which nuclear localization signals (NLSs) are necessary for localization at the INM. To determine if such a pathway to the INM exists in plants, I fused an NLS to a tail-anchored ER protein, PICL, and determined that a GFP-tagged, NLS-fused PICL is enriched at the NE in <i>Nicotiana benthamiana</i> leaf epidermal cells. Localization at the INM compared to outer nuclear membrane (ONM) was resolved using Airyscan sub-diffraction limited confocal microscopy, and NLS-fused PICL was shown to access the INM. Nine monopartite and bipartite NLSs were tested for their sufficiency to enrich a chimeric membrane protein, Heh2linker-TMD, at the NE. All nine NLSs were sufficient for enrichment of the chimeric membrane at the NE, and a monopartite and bipartite NLS were each shown to be sufficient for localization to the INM. This data is the first evidence that an NLS-mediated pathway to the INM exists in plants. Presence of an NLS and transmembrane domain (TMD) was used as criterium to identify novel INM proteins in plants. The <i>Arabidopsis thaliana</i> proteome was queried for presence of both domains, resulting in a shortlist of 32 putative inner nuclear envelope proteins (PINEs). A secondary search for <i>Arabidopsis</i> homologs of mammalian nuclear envelope transmembrane proteins (NETs) yielded 16 proteins, <i>Arabidopsis</i> SUN1 among them. The results of these queries are presented here.While the INM component of LINC complexes, SUN proteins, are conserved from animals and yeast to plants, the ONM components of plant LINC complexes has diverged in plants. Those proteins, KASH proteins, are defined by a conserved C-terminus consisting of a TMD and NE luminal tail. KASH proteins vary in their cytoplasmic domains, which interact directly or indirectly with the cytoskeleton. While… Advisors/Committee Members: Meier , Iris (Advisor).

Subjects/Keywords: Cellular Biology; Molecular Biology; Plant Biology; Plant Sciences; Protein trafficking, Tail-anchored protein, Nuclear Localization signal, Plant, Inner Nuclear Membrane, Outer Nuclear Membrane, KASH

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APA (6th Edition):

Groves, N. R. (2019). Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>. (Doctoral Dissertation). The Ohio State University. Retrieved from http://rave.ohiolink.edu/etdc/view?acc_num=osu1557145385558223

Chicago Manual of Style (16th Edition):

Groves, Norman R. “Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>.” 2019. Doctoral Dissertation, The Ohio State University. Accessed November 17, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=osu1557145385558223.

MLA Handbook (7th Edition):

Groves, Norman R. “Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>.” 2019. Web. 17 Nov 2019.

Vancouver:

Groves NR. Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>. [Internet] [Doctoral dissertation]. The Ohio State University; 2019. [cited 2019 Nov 17]. Available from: http://rave.ohiolink.edu/etdc/view?acc_num=osu1557145385558223.

Council of Science Editors:

Groves NR. Determining features sufficient for protein trafficking to the plant inner nuclear membrane and identification of putative nuclear envelope-associated proteins in <i>Arabidopsis thaliana</i>. [Doctoral Dissertation]. The Ohio State University; 2019. Available from: http://rave.ohiolink.edu/etdc/view?acc_num=osu1557145385558223

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