The retraction of the protoplast during PCD is an active, and interruptible, calcium-flux driven process

Abstract

The protoplast retracts during apoptosis-like programmed cell death (AL-PCD) and, if this retraction is an active component of AL-PCD, it should be used as a defining feature for this type of programmed cell death. We used an array of pharmacological and genetic tools to test if the rates of protoplast retraction in cells undergoing AL-PCD can be modulated. Disturbing calcium flux signalling, ATP synthesis and mitochondrial permeability transition all inhibited protoplast retraction and often also the execution of the death programme. Protoplast retraction can precede loss of plasma membrane integrity and cell death can be interrupted after the protoplast retraction had already occurred. Blocking calcium influx inhibited the protoplast retraction, reduced DNA fragmentation and delayed death induced by AL-PCD associated stresses. At higher levels of stress, where cell death occurs without protoplast retraction, blocking calcium flux had no effect on the death process. The results therefore strongly suggest that retraction of the protoplast is an active biological process dependent on an early Ca-mediated trigger rather than cellular disintegration due to plasma membrane damage. Therefore this morphologically distinct cell type is a quantifiable feature, and consequently, reporter of AL-PCD.

The nitrogen-fixing symbiotic cyanobacterium, Nostoc punctiforme can regulate plant programmed cell death

“…To investigate whether N. punctiforme can regulate PCD, we used a well-established method involving a 10-min, 51°C temperature stress to induce PCD in A. thaliana suspension cell cultures (Kacprzyk et al, 2017) following a 1-hr pre-incubation with N. punctiformeconditioned medium (CM; 20% v/v). This workflow is depicted in Fig.…”

“…5 × 10 4 cells ml -1 ) and left to pre-incubate for 1 hr under standard culturing conditions. Next, cells were either incubated at room temperature (~21°C) or placed in a water bath set to 51°C for 10 min with constant agitation at 120 rpm (Kacprzyk et al, 2017). After returning to standard culturing conditions for 24 hr, the frequency of viable, PCDcommitted and necrotic cells was measured.…”

Determining the effect of calcium on cell death rate and perforation formation during leaf development in the novel model system, the lace plant (Aponogeton madagascariensis)

“…To date, the details of the PCD process and unique signalling cascades have been more thoroughly investigated in animals compared to plants (Dauphinee & Gunawardena, ). Similarities between plant and animal PCD or apoptosis have been found, however, including nuclear condensation, nuclear DNA (nDNA) fragmentation, release of cytochrome c (cyt‐c) and the involvement of caspase‐like proteases (Lord & Gunawardena, ; Kacprzyk et al ., ). Increasing our knowledge of PCD in plants is important as it holds significant relevance for the agricultural and biotechnology industries.…”

“…However, some overlap does exist between these two categories. Based on the current literature, key effectors of PCD include the balance between antioxidants and reactive oxygen species (ROS) (de Pinto et al ., ; Wang et al ., ; Serrano et al ., ; Dauphinee et al ., ), hormones such as ethylene (Trobacher, ; Dauphinee et al ., ), release of factors such as cyt‐c (Balk et al ., ; Yu et al ., ) and influx and efflux of signalling ions like calcium (O'Brien & Ferguson, ; Stael et al ., ; Kacprzyk et al ., ).…”

“…Previous investigations have employed different calcium modulators to elucidate the involvement of Ca 2+ in the regulation of cell functioning (Blevins et al ., ; O'Brien & Ferguson, ; Minibayeva et al ., ; Liu et al ., ; Kacprzyk et al ., ). Exposure to the calcium ionophore A23187 increases oxygen consumption and heat production in wheat ( Triticum aestivum ) roots (Minibayeva et al ., ), reduces nodule number and weight in soybeans ( Glycine max ) (Blevins et al ., ) and stimulates the apoptotic pathway in tobacco ( Nicotiana plumbaginifolia ) (O'Brien & Ferguson, ).…”