Chloroplast nucleoids are highly dynamic in ploidy, number, and structure during angiosperm leaf development.

Stephan Greiner,Tommaso Pellizzer,Irina Malinova,Ralph Bock,Hieronim Golczyk,Thomas Börner,Reinhold G Herrmann

doi:10.1111/tpj.14658

Abstract

Chloroplast nucleoids are large, compact nucleoprotein structures containing multiple copies of the plastid genome. Studies on structural and quantitative changes of plastid DNA (ptDNA) during leaf development are scarce and have produced controversial data. We have systematically investigated nucleoid dynamics and ptDNA quantities in the mesophyll of Arabidopsis, tobacco, sugar beet, and maize from the early post-meristematic stage until necrosis. DNA of individual nucleoids was quantified by DAPI-based supersensitive epifluorescence microscopy. Nucleoids occurred in scattered, stacked, or ring-shaped arrangements and in recurring patterns during leaf development that was remarkably similar between the species studied. Nucleoids per organelle varied from a few in meristematic plastids to >30 in mature chloroplasts (corresponding to about 20-750 nucleoids per cell). Nucleoid ploidies ranged from haploid to >20-fold even within individual organelles, with average values between 2.6-fold and 6.7-fold and little changes during leaf development. DNA quantities per organelle increased gradually from about a dozen plastome copies in tiny plastids of apex cells to 70-130 copies in chloroplasts of about 7μm diameter in mature mesophyll tissue, and from about 80 plastome copies in meristematic cells to 2600-3300 copies in mature diploid mesophyll cells without conspicuous decline during leaf development. Pulsed-field electrophoresis, restriction of high-molecular-weight DNA from chloroplasts and gerontoplasts, and CsCl equilibrium centrifugation of single-stranded and double-stranded ptDNA revealed no noticeable fragmentation of the organelle DNA during leaf development, implying that plastid genomes in mesophyll tissues are remarkably stable until senescence.

Highlights

The plastid genome represents one of three spatially separated cellular subgenomes constituting the genetic system of plants
The multiple copies of the plastid genome are condensed in nucleoids that reside in the stroma and exhibit prokaryotic properties, consistent with the cyanobacterial ancestry of the plastid
Leaflets and leaves from which samples were taken are described in Material and Methods, some examples are photographically documented in Golczyk et al (2014), and for sugar beet, in Rauwolf et al (2010)

Summary

Introduction

The plastid genome (plastome; Renner, 1934) represents one of three spatially separated cellular subgenomes constituting the genetic system of plants. The development of chloroplasts from undifferentiated proplastids present in meristems is accompanied by an increase of plastids in both size and number per cell (cf Butterfass, 1979) This includes a substantial increase in nucleoid number and plastome copies per cell, while nuclear DNA amounts remain constant (e.g., Herrmann and Kowallik, 1970, Selldén and Leech, 1981, Boffey and Leech, 1982, Hashimoto, 1985, Miyamura et al, 1986, Baumgartner et al, 1989, Miyamura et al, 1990, Fujie et al, 1994, Rauwolf et al, 2010, Golczyk et al, 2014, Ma and Li, 2015)

Results

Discussion

Conclusion