DNA maintenance in plastids and mitochondria of plants

Abstract

The DNA molecules in plastids and mitochondria of plants have been studied for over 40 years. Here, we review the data on the circular or linear form, replication, repair, and persistence of the organellar DNA (orgDNA) in plants. The bacterial origin of orgDNA appears to have profoundly influenced ideas about the properties of chromosomal DNA molecules in these organelles to the point of dismissing data inconsistent with ideas from the 1970s. When found at all, circular genome-sized molecules comprise a few percent of orgDNA. In cells active in orgDNA replication, most orgDNA is found as linear and branched-linear forms larger than the size of the genome, likely a consequence of a virus-like DNA replication mechanism. In contrast to the stable chromosomal DNA molecules in bacteria and the plant nucleus, the molecular integrity of orgDNA declines during leaf development at a rate that varies among plant species. This decline is attributed to degradation of damaged-but-not-repaired molecules, with a proposed repair cost-saving benefit most evident in grasses. All orgDNA maintenance activities are proposed to occur on the nucleoid tethered to organellar membranes by developmentally-regulated proteins.

Keywords: DNA recombination; DNA repair; DNA replication; chloroplast DNA; organellar DNA.

FIGURE 1

Fluorescence microscopic images of ethidium-stained mtDNA and ptDNA molecules. (A) and (B) Images of DNA-protein structure from osmotically lysed tobacco BY-2 mitochondria. Complex branching DNA-protein structure with three bright nodes, long immobile fiber, and several fibers that extend leftward toward the anode (examples: 1 and 2) and rightward when the polarity of the electric field was reversed. (Adapted from Oldenburg and Bendich, 1998b). (C) Maize ptDNA molecules from the well-bound fraction following PFGE. Examples: (1) multigenomic complex structure with a Y-branch and (2) a genome-sized circular molecule. Approximately 84% of the DNA mass was in the large complex form, 11% in small branched molecules, and 4% in circular molecules. The in-gel ptDNA was prepared from 14-day maize seedlings. (Adapted from Oldenburg and Bendich, 2004a) (D) and (E) Images of liverwort mtDNA molecules from the well-bound fraction following PFGE. One large complex structure with two bright nodes of fluorescence that are connected by a bright fiber and several fibers extend from each node toward the anode (1). Two smaller “comet” structures with several “tail” fibers extending from the bright ”head” (2, 3). A few small molecules were moving toward the anode (examples: 4, 5). (Adapted from Oldenburg and Bendich, 1998a) The molecules in (B) and (E) were recorded using an epifluorescence microscope equipped with a CCD camera, video monitor, and recorder. Photographs were then taken of ethidium-stained DNA on the monitor and the respective drawings, (A) and (D), were made by tracing the DNA on the monitor. The molecules in (C) were recorded using an epifluorescence microscope equipped with a digital camera and computer. Broad arrows point toward the anode in (A), (B), (D), and (E).

FIGURE 2

Changes in orgDNA during maize development. Recombination-dependent replication of orgDNA in the basal meristem produces branched, multigenomic chromosomes in proplastids and mitochondria (not depicted). DNA-damaging oxidative stress is minimized, requiring little repair, by maintaining hypoxia, antioxidants, and no ROS-generating photosynthesis or respiration. Early in leaf development, orgDNA damage occurs due to ROS generated in photosynthesis, respiration, and oxidation of pigments and lipids. Later, when the damage level exceeds the repair capacity, orgDNA is fragmented and no longer functions in coding or heredity, mitochondria switch from respiration to photorespiration, and DNA copy number declines faster for mitochondria than for chloroplasts. (Reprinted from Kumar et al., 2014).

FIGURE 3

Single-strand annealing mechanism for plastid DNA replication. This single-strand annealing (SSA), recombination-dependent replication model for ptDNA is based on a replication mechanism for herpes virus DNA (Weller and Sawitzke, 2014). (1) A 3′-overhang is generated by 5′-to-3′ exonuclease digestion at the end of a unit-genome-sized monomer. A single-strand annealing protein (SSAP) binds to a 3′-overhang. (2) Annealing of the 3′-overhang of Molecule 1 to a homologous single-strand gap in another ptDNA molecule (Molecule 2). (3) Replication is initiated by priming at the 3′-end, assembly of the replisome, and formation of a replication fork, leading to a branched-linear structure. A similar model with the same or analogous proteins applies to the replication of mtDNA in plants. We propose that replication occurs only with ptDNA attached to the nucleoid-on-membrane using single-strand end-binding proteins. Although we propose that Whirly proteins serve attachment and SSAP functions, other single-strand-binding proteins, such as the OSB and RecA families, may also participate in ptDNA replication. Other replication and recombination mechanisms have been described (Cox, 2007; Marechal and Brisson, 2010; Weller and Sawitzke, 2014; Morrical, 2015).

FIGURE 4

Schematic representation of changes in the amount of ptDNA per plastid during development in three plant species. Increase in ptDNA amount due to ptDNA replication occurs very early in development in maize (red line), followed by a rapid decline. For Arabidopsis (blue line), the increase in ptDNA occurs slightly later and the decline in ptDNA amount is much later. For tobacco (gray line), ptDNA increases more gradually and the decline is less severe. The Roman numerals indicate stages of leaf development. I–III represent expanding leaves, and IV and V represent expanded leaves. (Reprinted from Rowan and Bendich, 2009).