- Title
- Investigating the role of cell wall invertase in ovule development in Arabidopsis thaliana
- Creator
- Liao, Shengjin
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2019
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- Ovule development is essential for the realization of crop yield as it determines seed number. Cell wall invertase (CWIN)-mediated sucrose metabolism and signalling is central to plant reproductive development including the formation of pollen fertility and seed- and fruit- set as well as their stress responses. It remains elusive, however, whether and how CWIN controls ovule development. Here, we provide a set of molecular evidence that CWIN functions as a positive regulator in ovule initiation and development. First, in situ RNA hybridization experiments revealed that the two reproductive specific genes CWIN2 and CWIN4 were highly expressed at the placenta region where ovule primordia initiated. To experimentally validate the potential role of CWIN in ovule initiation and subsequent development, we generated pSTK::amiRCWIN24 transgenic plants, in which an ovule-specific SEEDSTICK promoter (pSTK) was used to drive the artificial microRNAs (amiRCWIN24) specifically targeting CWIN2 and CWIN4. The pSTK::amiRCWIN24 transgenic plants exhibited a significant decrease in the transcript levels of CWIN2 and 4 and considerable reduction in ovule number and increased ovule abortion, which collectively results in over 50% seed loss as compared to WT. Further analysis indicates that over 65% of the seed loss was due to blockage of ovule initiation; whereas less than 35% of the loss was resulted from ovule abortion in the transgenic plants. The data clearly showed that the blockage of ovule initiation is the primary cause of seed number reduction in the pSTK::amiRCWIN24 transgenic plants. The degree of seed number loss is further aggravated by the ovule abortion for those ovules that did initiate. These findings demonstrated that CWIN plays an important role in determining the formation of ovules at the early reproductive stage. To explore the underlying mechanism of CWIN-mediated ovule initiation, RNA-Seq analysis was conducted to compare the transcript profiles between transgenic and WT plants at the ovule initiation stage by using pooled flower bud samples harvested at floral stage 8-10, which covers a stage prior to (stage 8), during (stage 9) and immediately after (stage 10) ovule initiation. Through this exercise, we identified several potential pathways that may participate in the CWIN-mediated ovule initiation. The data indicate that CWIN may exert its control over ovule initiation through suppressing SPL8- and SPL14- pathway, in which SPL8 and SPL14 may function as negative regulators repressing MADS transcription factor such as STK, AGL66 and AGL104 to block ovule initiation. Alternatively, CWIN-mediated ovule initiation may be achieved through a positive regulation of SAURs, in which SAURs inhibit the expression of D clade subfamily of type 2C protein phosphatases (PP2C.Ds) to promote ovule initiation. Since ovule primordia connect the placenta symplasmically during ovule initiation in Arabidopsis, while CWIN functions in the apoplasm, it is unlikely that the inhibition of ovule formation is casued by the lack of supply of carbon nutrients in the CWIN-suppression transgenic plants. This, together with the emerging evidence that CWIN-mediated sucrose hydrolysis plays a major role in sugar signalling to impact plant development, suggests that CWIN may regulate ovule initiation through sugar signalling. Relevently, our RNA-Seq analysis identified a cohort of putative extracellular receptor-like-kinases (RLKs) and intracellular small GTPases (Rops) that were also differentially expressed in response to CWIN-silencing in the transgenic plants. Together, these findings suggest that CWIN-mediated sugar signalling may be perceived by or transmitted through RLKs and Rop signalling components to regulate ovule formation by modulating the expression of downstream candidate genes such as , STK, AGL66, AGL104 and early auxin response genes, SAURs.
- Subject
- cell wall invertase; ovule initiation; Arabidopsis thaliana; sugar signalling; ovule abortion
- Identifier
- http://hdl.handle.net/1959.13/1397777
- Identifier
- uon:34354
- Rights
- Copyright 2019 Shengjin Liao
- Language
- eng
- Full Text
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