How does targeted proteolysis regulate ABA signalling?

Through our genetic and transcriptomic analyses we have shown that the N-end rule pathway is key to removing ABA sensitivity for germination, that it does this by interacting with the ABA transduction pathway, and that it influences the transcriptome by relieving the ABA-associated repression of a cohort of genes through removal of an ABA signal transduction-related protein substrate(s). Therefore our identification of the role of this protein degradation pathway offers a timely and unique window of opportunity to understand how targeted proteolysis regulates ABA signalling. The key hypotheses that need addressing are: 1. That germination induced endo-peptidase activity initiates the degradation of a key component(s) of ABA signalling through the N-end rule pathway. 2. That the ABA-related transcription factor ABI3 represents a substrate for the N-end rule pathway. 3. That the endosperm is the major site of action of the N-end rule pathway in the control of germination potential. 4. That the N-end rule pathway influences germination potential through a genetic network of mutually-exclusive repression and activation of key enzymes of endosperm functionality, via the transcription factors ABI3 and EIL4. 5. That the N-end rule pathway may have other functions during plant growth and development associated with removal of ABA sensitivity.

We have identified a key role for the N-end rule pathway of protein degradation in the regulation of ABA signalling in imbibed seeds. The key hypotheses that will be addressed in this grant are: 1. That germination induced endo-peptidase activity initiates the degradation of a key component(s) of ABA signalling through the N-end rule pathway. 2. That the ABA-related transcription factor ABI3 represents a substrate for the N-end rule pathway. 3. That the endosperm is the major site of action of the N-end rule pathway in the control of germination potential. 4. That the N-end rule pathway influences germination potential through a genetic network of mutually-exclusive repression and activation of key enzymes of endosperm functionality, via the transcription factors ABI3 and EIL4. 5. That the N-end rule pathway may have other functions during plant growth, development and environmental interaction associated with removal of ABA sensitivity. Outputs from the work programme: With the proposed work plan we will: - determine the complete pathway of regulation of ABA sensitivity of imbibed seeds. - identify the endo-peptidase activity(s) that initiates the pathway, and investigate potential substrates acted upon by the protease, - determine in which seed compartment the pathway functions, - determine the topology of the genetic network regulating endosperm functionality. - determine whether this pathway is also important for other ABA-related processes during growth, development and environmental response. ABA sensitivity of germination is a important determinant of agronomically important traits including Pre-Harvest Sprouting in wheat, a major determinant of seed quality and therefore food security. Characterisation of this pathway will provide candidate genes for testing in cereals, providing strategies to overcome these negative traits.