Caspase inhibitors affect the kinetics and dimensions of tracheary elements in xylogenic Zinnia (Zinnia elegans) cell cultures

Abstract

BackgroundThe xylem vascular system is composed of fused dead, hollow cells called tracheary elements (TEs) that originate through trans-differentiation of root and shoot cambium cells. TEs undergo autolysis as they differentiate and mature. The final stage of the formation of TEs in plants is the death of the involved cells, a process showing some similarities to programmed cell death (PCD) in animal systems. Plant proteases with functional similarity to proteases involved in mammalian apoptotic cell death (caspases) are suggested as an integral part of the core mechanism of most PCD responses in plants, but participation of plant caspase-like proteases in TE PCD has not yet been documented.ResultsConfocal microscopic images revealed the consecutive stages of TE formation in Zinnia cells during trans-differentiation. Application of the caspase inhibitors Z-Asp-CH2-DCB, Ac-YVAD-CMK and Ac-DEVD-CHO affected the kinetics of formation and the dimensions of the TEs resulting in a significant delay of TE formation, production of larger TEs and in elimination of the 'two-wave' pattern of TE production. DNA breakdown and appearance of TUNEL-positive nuclei was observed in xylogenic cultures and this was suppressed in the presence of caspase inhibitors.ConclusionsTo the best of our knowledge this is the first report showing that caspase inhibitors can modulate the process of trans-differentiation in Zinnia xylogenic cell cultures. As caspase inhibitors are closely associated with cell death inhibition in a variety of plant systems, this suggests that the altered TE formation results from suppression of PCD. The findings presented here are a first step towards the use of appropriate PCD signalling modulators or related molecular genetic strategies to improve the hydraulic properties of xylem vessels in favour of the quality and shelf life of plants or plant parts.

The Plant Vascular System: Evolution, Development and FunctionsF

“…This result suggested the involvement of ATMC9 in PCD. However, application of caspase inhibitors significantly delays the time of tracheary element formation and inhibits DNA breakdown and appearance of TUNEL‐positive nuclei in Zinnia xylogenic cell culture (Twumasi et al 2010).…”

Xylogenesis in zinnia (Zinnia elegans) cell cultures: unravelling the regulatory steps in a complex developmental programmed cell death event

“…However, in addition to vacuole expansion and collapse and cellular autolysis, typical for vacuolar cell death, also other PCD features have been observed: nucleus condensation, oxidative stress-related processes, laddering type of DNA fragmentation and activation of PCD-associated enzymes such as CLPs (Bonneau et al 2008; Twumasi et al 2010a; Woltering 2010; Han et al 2012; Petzold et al 2012 and references therein). This suggests that zinnia TE differentiation may involve signalling pathways of both vacuolar and necrotic PCD classes.…”

Post mortem function of AtMC9 in xylem vessel elements

“…ZINNIA ENDONUCLEASE1 (ZEN1), an S1-type nuclease, was shown to be required for post mortem DNA degradation in Zinnia elegans TEs (Ito & Fukuda, 2002). Serine and cysteine protease activities have been shown to increase during TE differentiation (Minami & Fukuda, 1995;Ye & Varner, 1996;Beers & Freeman, 1997;Groover & Jones, 1999) and, consequently, pharmacological inhibition of cysteine proteases in xylogenic cell cultures blocked or delayed TE differentiation, but only if applied before formation of secondary wall thickenings (Fukuda, 1996;Woffenden et al, 1998;Twumasi et al, 2010). While this suggests a role for cysteine proteases in early signalling of TE differentiation, it does not exclude roles during autolysis.…”