The crucial elements of the ‘last step’ of programmed cell death induced by kinetin in root cortex of V. faba ssp. minor seedlings

Abstract

Kinetin-induced programmed cell death, manifested by condensation, degradation and methylation of DNA and fluctuation of kinase activities and ATP levels, is an autolytic and root cortex cell-specific process. The last step of programmed cell death (PCD) induced by kinetin in the root cortex of V. faba ssp. minor seedlings was explained using morphologic (nuclear chromatin/aggregation) and metabolic (DNA degradation, DNA methylation and kinases activity) analyses. This step involves: (1) decrease in nuclear DNA content, (2) increase in the number of 4',6-diamidino-2-phenylindole (DAPI)-stained chromocenters, and decrease in chromomycin A3 (CMA3)-stained chromocenters, (3) increase in fluorescence intensity of CMA3-stained chromocenters, (4) condensation of DAPI-stained and loosening of CMA3-stained chromatin, (5) fluctuation of the level of DNA methylation, (6) fluctuation of activities of exo-/endonucleolytic Zn(2+) and Ca(2+)/Mg(2+)-dependent nucleases, (7) changes in H1 and core histone kinase activities and (8) decrease in cellular ATP amount. These results confirmed that kinetin-induced PCD was a specific process. Additionally, based on data presented in this paper (DNA condensation and ATP depletion) and previous studies [increase in vacuole, increase in amount of cytosolic calcium ions, ROS production and cytosol acidification "in Byczkowska et al. (Protoplasma 250:121-128, 2013)"], we propose that the process resembles autolytic type of cell death, the most common type of death during development of plants. Lastly, the observations also suggested that regulation of these processes might be under control of epigenetic (methylation/phosphorylation) mechanisms.

Caffeine-Induced Premature Chromosome Condensation Results in the Apoptosis-Like Programmed Cell Death in Root Meristems of Vicia faba

“…faba root meristem cells, DNA degradation was not observed as a typical internucleosomal process, but rather showed as 'a smear' indicating an exonucleolytic type of DNA degradation [ 53 ], despite the internucleosomal DNA degradation observed during PCD in other plants [ 41 , 54 – 55 ]. Following this, more detailed research by Kunikowska et al [ 53 ] revealed digestion by acidic and basic nucleases of faba bean DNA in kinetin-treated (72 h) seedlings [ 56 ].…”

“…: (iii) micro-autophagy; (iv) macro-autophagy, and (v) mega-autophagy [ 59 ]. The type of cell death, and even the nature of the accompanying nuclear DNA fragmentation, appear to depend not only on the cell type (and species) but even on the experimental system used [ 8 , 30 , 40 , 53 , 56 , 60 ]. For this reason in this work (as in the vast majority of other studies) the DNA fragmentation during PCD evaluated by DNA electrophoresis needed to be confirmed by TUNEL reaction.…”

“…On the other hand, transition of the condensed chromatin to the nucleus periphery and the breakup of a nucleus into small fragments as well as its degradation, seem to be characteristic of final cell degradation (i.e. 'last step' [ 41 – 42 , 56 ]). The phenomenon of cell death is accompanied by proteolysis, often mediated by cysteine proteinases–caspases [ 65 ], while a particular role in the synthesis and accumulation of these proteins is ascribed to mitochondria [ 66 ].…”

“…This does not mean of course that only reversible transformations occur in a cell. PCD, in both animal and plant cells, is an example of a cellular process that looks irreversible; especially if the changes in the cell nuclei during PCD reach 'the point beyond which the cell is irreversibly committed to die' [ 42 , 56 ]. The present study provides the first evidence that 'point of no return' can be the consequence of caffeine-induced genotoxic stress in HU-synchronized V .…”

Mechanism of kinetin-induced death of Vicia faba ssp. minor cortex cells

“…Hallmarks of Kin-ECD were extensively 3,4,5,7,8,9 and thoroughly 10 scrutinised. One of the effects of Kin-ECD is degradation of about 50% of the cortex cells in faba bean seedling roots making cell-free spaces known as aerenchyma 3,7,15/8 .…”