title: Caffeine-Induced Premature Chromosome Condensation Results in the Apoptosis-Like Programmed Cell Death in Root Meristems of Vicia faba.
abstract: We have demonstrated that the activation of apoptosis-like programmed cell death (AL-PCD) was a secondary result of caffeine (CF) induced premature chromosome condensation (PCC) in hydroxyurea-synchronized Vicia faba root meristem cells. Initiation of the apoptotic-like cell degradation pathway seemed to be the result of DNA damage generated by treatment with hydroxyurea (HU) [double-stranded breaks (DSBs) mostly] and co-treatment with HU/CF [single-stranded breaks (SSBs) mainly]. A single chromosome comet assay was successfully used to study different types of DNA damage (neutral variant-DSBs versus alkaline-DSBs or SSBs). The immunocytochemical detection of H2AXS139Ph and PARP-2 were used as markers for DSBs and SSBs, respectively. Acridine orange and ethidium bromide (AO/EB) were applied for quantitative immunofluorescence measurements of dead, dying and living cells. Apoptotic-type DNA fragmentation and positive TUNEL reaction finally proved that CF triggers AL-PCD in stressed V. faba root meristem cells. In addition, the results obtained under transmission electron microscopy (TEM) further revealed apoptotic-like features at the ultrastructural level of PCC-type cells: (i) extensive vacuolization; (ii) abnormal chromatin condensation, its marginalization and concomitant degradation; (iii) formation of autophagy-like vesicles (iv) protoplast shrinkage (v) fragmentation of cell nuclei and (vi) extensive degeneration of the cells. The results obtained have been discussed with respect to the vacuolar/autolytic type of plant-specific AL-PCD.
authors.affiliation: Department of Cytophysiology, Institute of Experimental Biology, Faculty of Biology and Environmental Protection, University of Łódź, Łódź, Poland.
authors.forename: Dorota
authors.initials: D
authors.lastname: Rybaczek
authors.affiliation: Department of Cytophysiology, Institute of Experimental Biology, Faculty of Biology and Environmental Protection, University of Łódź, Łódź, Poland.
authors.forename: Marcelina Weronika
authors.initials: MW
authors.lastname: Musiałek
authors.affiliation: Department of Molecular Biophysics, Faculty of Biology and Environmental Protection, University of Łódź, Łódź, Poland.
authors.forename: Aneta
authors.initials: A
authors.lastname: Balcerczyk
mesh_terms: D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D032461:Chromosomes, Plant / Q000378:metabolism*; D004249:DNA Damage / Q000187:drug effects; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D018519:Meristem / Q000166:cytology / Q000187:drug effects*; D018517:Plant Roots / Q000166:cytology; D014617:Vacuoles / Q000187:drug effects; D031307:Vicia faba / Q000166:cytology*
chemical_list: D002110:Caffeine; D006918:Hydroxyurea