|dc.description.abstract||The effect of mechanical stimulation in plants has been studied in depth for more
than a century. This type of stress has been shown to trigger alterations in
development such as stunting, thickened stems and differential cell wall deposition.
These responses are very likely to be initiated at a subcellular level, but the
molecular mechanisms transducing mechanical signals into intracellular responses
still remain unknown in plants. In this thesis I test the hypothesis that the
membrane anchored protein Defective Kernel 1 (DEK1) could act as a plant-specific
mechanosensor in plants.
Constitutive overexpression of the cytoplasmic CALPAIN domain DEK1 causes a
phenotype in Arabidopsis, that that resembles that of mechanically stressed plants.
The CALPAIN domain of DEK1 shows a very high homology with animal calpains;
a class of calcium-dependent Cysteine proteases which undergo a calciumstimulated
CALPAIN domain-releasing autolytic cleavage event during activation.
A similar autolytic cleavage event has been observed in DEK1 which, together with
the fact that the CALPAIN domain alone can rescue the embryo-lethality associated
with loss of DEK1 function, has led to the suggestion that this domain represents an
activated form of the protein. I show that like mechanically stressed plants,
CALPAIN overexpressing plants show a modified call wall composition. Consistent
with this, transcriptional analysis of these plants shows a deregulation of genes
encoding cell wall modifying enzymes, amongst others. Other characteristics of
mechanically stimulated plants which I have characterized in CALPAIN
overexpressing lines include late flowering and thickened stems. Therefore, I
proposed a model in which the CALPAIN domain of DEK1 acts as an effector which
is normally activated by mechanical stimulation. In this model, the transmembrane
domains of DEK1 would regulate activation (cleavage) of the CALPAIN domain,
potentially in response to mechanical stress.
In order to test this model further, CALPAIN overexpressing lines were generated
in a dek1 mutant background. If the model is correct, these plants should not only
behave as if responding constitutively to mechanical stimulation, but should also
lack appropriate responses to applied mechanical stimuli due to lack of the
mechanosensory integral membrane domain of DEK1. My results confirm that the
absence of the transmembrane domains of DEK1 is indeed translated into a lack of
some, but not all responses to mechanical stimulation compared to wild-type plants.
Furthermore, the lack of the transmembrane domains of DEK1 correlates with the
absence of a mechanically-triggered calcium flux in the plant. Thus my work
suggests that the transmembrane domains of DEK1 are involved in sensing
mechanical stimulation, via the regulation activity of a mechano-sensitive calcium
flux at the plasma membrane.
In summary, my proposal is that Defective Kernel 1 (DEK1) acts both as a key
mechanosensory cellular component, and as the first effector of the signalling
cascade in response to mechanical stimulation, via an autolytic activation in
response to mechanical stress.||en