Doctor of Philosophy
Agriculture and Agri-Food Canada
The nuclear pore complex (NPC) is the gateway between the nucleus and cytoplasm, which provides the passage for transport of RNA, protein, and other molecules into and out of the nucleus. NPC is conserved across all eukaryotes and plays a vital role in various cellular processes. However, compared to other organisms, the study of NPC in plants is limited. Although more than 30 different types of nucleoporin proteins in the model plant Arabidopsis thaliana have been identified, none of those proteins has been studied in detail. In this thesis, I focused on one such protein named NUCLEOPORIN1 (NUP1) and investigated its role in seed germination and pollen development.
Since the expression of NUP1 is highest in seeds, I proposed and tested the hypothesis of NUP1 participation in seed germination especially under various abiotic stresses. Genome-wide transcriptomic analysis revealed alteration in abscisic acid (ABA) related genes in nup1. I also found that NUP1 interacts and regulates ABA INSENSITIVE 5 (ABI5), a major negative regulator of germination. Another important function of NUP1 is its role in fertility. NUP1 also plays a major role in pollen development. I compared the development of pollen in the Col-0 to that of nup1 mutant plants. Pollen study indicates the possible role of NUP1 in pollen nuclei migration during pollen tube growth, which directly affects the double fertilization in Arabidopsis.
The results from my thesis contribute to the understanding of the roles of NUP1 during seed germination and pollen development, which are both very important in agriculture. The knowledge generated from this work will help to develop better crops in the future.
Summary for Lay Audience
Plant cells contain a nucleus where most of the DNA is stored. The nucleus is surrounded by a nuclear membrane, which has nuclear pore complexes (NPCs). NPCs act as the gates between the nucleus and cytoplasm. Proteins and RNAs are transported through these pores, which is vital for cellular activity. For my study, I chose one protein NUCLEOPORIN1 (NUP1) from the NPC. To study the function of this protein, I obtained a nup1 Arabidopsis mutant line and characterized it as I can see how it affects the plants if this protein is non-functional. I focused on two functions of NUP1 in this model plant.
First, I investigated the role of NUP1 in seed germination, as its expression is highest in seeds. The mutant seed was able to germinate, similar to the Col-0 (control). However, when a plant hormone ABA was added, the germination of the mutant seed was delayed compared to Col-0. The addition of the ABA hormone mimics abiotic stresses to the plants in the field conditions. This suggests that NUP1 is required for the germination of seeds under abiotic stress conditions. I did a comprehensive analysis of gene expression changes during this germination process. Based on this, I found there are three genes related to germination namely ABA INSENSITIVE 3 (ABI3), ABA INSENSITIVE 4 (ABI4), and ABA INSENSITIVE 5 (ABI5), which could be affected by a mutation in NUP1. Out of these three genes, only a mutation in ABI5 can rescue the sensitive phenotype of nup1 seeds. NUP1 physically interacts with ABI5 and moves from the nucleus to cytoplasm for degradation. ABI5 is predominantly expressed in the nucleus in Col-0 plants, while in mutant plants, it cannot go out to the cytoplasm for degradation. Therefore, NUP1 likely plays a role in exporting of ABI5 for degradation.
Second, I investigated the role of NUP1 in pollen development as NUP1 expression is also high in pollen. The pollen morphology between Col-0 and mutant plants was highly similar with no significant difference in pollen germination. However, the fertility of the mutant plants was reduced compared to Col-0. Therefore, I studied different stages of pollen development to identify when the potential defect. I found that during pollen tube growth the position of nuclei was different in mutants from that in Col-0 plants. This difference in nuclear position may contribute to reduced fertility.
In summary, I studied two functions of the NUP1 protein in seed germination and pollen development. NUP1 is required for seed germination during stress conditions and nuclear positioning in pollen development. This knowledge will be useful to create better crops in the future.
Thapa, Raj K., "Multiple roles of NUP1 in Arabidopsis growth and development" (2020). Electronic Thesis and Dissertation Repository. 7428.
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Available for download on Thursday, December 01, 2022