References

6TargetsofSI 85
many of the genes encoding components of the apoptotic machinery, including
caspases, have either not yet been identified in plants or are simply
not present (reviewed by Woltering et al. 2002). Nevertheless, there is no
doubt that PCD occurs in plants and much of the evidence for it has come
from biochemical studies (reviewed by van Doorn and Woltering 2005).
We have used markers of PCD to investigate whether an incompatible SI
response triggers PCD in Papaver pollen.
6.3.2
Programmed Cell Death is Triggered During
the Papaver Self-Incompatibility Response
Cytochrome c release is an early event of SI-induced PCD. Since leakage of
cytochrome c into the cytosol is a key early marker for PCD, we investigated
whether this occurred in pollen tubes undergoing incompatible SI. We
showed that large increases in cytosolic cytochrome c were detected in
incompatible pollen (Thomas and Franklin-Tong 2004). The increase in
cytochrome c release was rapid, beginning at 10 min and increasing up
to 2 h, after SI induction. In controls, no release of cytochrome c was
observed. The first detection of cytochrome c in the cytosol at around
10 min corresponds to the point at which peak phosphorylation and MAP
kinase activity of p56 are detected (Rudd et al. 2003). Our data may therefore
indicate crosstalk between different signalling cascades and may represent
the point at which the pollen tube becomes committed to death.
DNA fragmentation is stimulated by SI. Since DNA fragmentation is a classic
marker for PCD, we investigated whether DNA fragmentation occurred
in incompatible Papaver pollen undergoing SI. DNA fragmentation was
detected in incompatible pollen and was S-specific, as it was not observed
in either untreated pollen tubes or compatible pollen tubes (Jordan et
al. 2000). DNA fragmentation is first detected at 4 h and continues to increase
for at least 16 h after SI induction. Pre-treatment of pollen tubes
with DEVD prior to the induction of SI caused a significant reduction in
the amount of DNA fragmentation, reducing it from levels of 71.6% in
SI to 18.97% in pollen tubes with SI-induced in the presence of DEVD
(Fig. 6.3a) (Thomas and Franklin-Tong 2004). YVAD (used as a negative
control) displayed only a small decrease in DNA fragmentation. These data
implicate the involvement of a caspase-3 like activity in SI-mediated DNA
fragmentation. Furthermore, DEVD pre-treatment of pollen tubes allowed
growth to resume following SI induction, whilst YVAD did not alleviate SIinduced
growth inhibition (Thomas and Franklin-Tong 2004). These data
demonstratethatacaspase-3likeactivityisalsoinvolvedinmechanisms
mediating SI-induced pollen tube inhibition.