The DELLA protein family and gibberellin signal transduction : : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Biochemistry, Massey University, Palmerston North, New Zealand
Gibberellins (GA) are plant hormones that promote important aspects of growth such as seed
germination, leaf expansion, trichome initiation, transition to flowering, stem elongation,
flower and fruit development. Genetic and molecular data indicate that growth and
development are a default state and that the DELLA proteins are key repressors of GAmediated
growth and development. Mutant analysis indicates that GA does not directly
promote growth; rather it overcomes the repression of the DELLA proteins by causing them to
be degraded. The N-terminal domain of the DELLA proteins is involved in the perception of the
GA signal and the C-terminal domain mediates the repression of GA responses. A GA-bound
receptor recognises the DELLA proteins and interacts with an F-box E3 ligase, the DELLA
proteins are then poly-ubiquitinated and degraded through the ubiquitin-26S proteasome
system. However, the DELLA proteins are also post-translationally modified which affects their
activity. It is believed the DELLAs are modified both with O-linked N-Acetyl glucosamine for
stability and phosphate groups to mark them for F-box recognition. However, the precise
nature, and role of these modifications is yet to be shown. DELLA-repressive action is mediated
by interaction with other proteins and not through direct DNA binding. Few DELLA-interacting
proteins are known.
Apple and Kiwifruit DELLA repressor, GID1 GA receptor and SLY1/ GID2 F-box orthologues were
identified in their respective sequence databases. Relative amount and location of the
orthologous transcript sequences was examined through qPCR and reporter gene experiments.
Apple qPCR experiments indicated relatively high levels of DELLA transcripts in
developmentally arrested tissues. Kiwifruit experiments present a more complicated picture,
with high relative levels of DELLAs in the actively expanding tissues, however, concomitant
with this were high relative levels of the GID1 and GID2 transcripts. Each transcript was found
in every tissue studied and indicated complex developmental transcriptional control.
Both direct and indirect immunoprecipitation experiments utilising a novel tag were
performed in GA-deficient plant backgrounds in order to isolate DELLA proteins and their
interacting proteins likely targeted by DELLA repressive function. Proteins from these
experiments were identified from their peptides in mass spectrometry analysis and database
query. Several transcription factors, kinases, proteins involved in RNA processing and protein
components involved in hormonal signalling pathways other than GA were present.
DELLA repression complex formation was also investigated with two-dimensional
electrophoresis and western blotting, and indicated a dominant repressive complex at
approximately 160 kDa, with additional multiple larger complexes of up to 600 kDa.
Papers removed due to copyright restrictions: Foster, T., Kirk, C. A., Jones, W. T., Allan, A. C., Espley, R., Karunairetnam, S. & Rakonjac, J.
(2007). Characterisation of the DELLA subfamily in Apple (Malus x domestica Borkh.).
Tree Genetics and Genomes, 3: 187-197.;
Jones, W. T., Harvey, D., Kirk, C., Rakonjac, J., Sun, X., Frearson, N. & Al-Samarrai, T. A. (2007).
A Novel Peptide tag for Detection and Purification of Recombinant Expressed Proteins.
Protein Expression and Purification, 53: 404-410.;
Sun, X., Frearson, N., Kirk, C., Jones, W. T., Harvey, D., Rakonjac, J., Foster, T. & Al-Samarrai, T.
(2008). An E. coli Expression System optimized for DELLA Proteins. Protein Expression
and Purification, 58: 168-174.;
Sun, X. L., Jones, W. T., Harvey, D., Edwards, P. J. B., Pascal, S. M., Kirk, C., Considine, T.,
Sheerin, D. J., Rakonjac, J., Oldfield, C. J., Xue, B., Dunker, A. K. & Uversky, V. N. (2010).
N-terminal Domains of DELLA Proteins Are Intrinsically Unstructured in the Absence of
Interaction with GID1/Gibberellic Acid Receptors. Journal of Biological Chemistry, 285:
Sheerin, D. J., Buchanan, J., Kirk, C., Harvey, D., Sun, X., Spagnuolo, J., Li, S., Liu, T., Woods, V.
A., Foster, T., Jones, W. T. & Rakonjac, J. (2011). Inter- and Intra-Molecular Interactions
of Arabidopsis thaliana DELLA protein RGL1. Biochemical Journal, 435: 629-639.