Proteomic analysis of highly purified prolamellar bodies reveals their significance in chloroplast development

Abstract

The prolamellar body (PLB) proteome of dark-grown wheat leaves was characterized. PLBs are formed not only in etioplasts but also in chloroplasts in young developing leaves during the night, yet their function is not fully understood. Highly purified PLBs were prepared from 7-day-old dark-grown leaves and identified by their spectral properties as revealed by low-temperature fluorescence spectroscopy. The PLB preparation had no contamination of extra-plastidal proteins, and only two envelope proteins were found. The PLB proteome was analysed by a combination of 1-D SDS-PAGE and nano-LC FTICR MS. The identification of chlorophyll synthase in the PLB fraction is the first time this enzyme protein was found in extracts of dark-grown plants. This finding is in agreement with its previous localization to PLBs using activity studies. NADPH:protochlorophyllide oxidoreductase A (PORA), which catalyses the reduction of protochlorophyllide to chlorophyllide, dominates the proteome of PLBs. Besides the identification of the PORA protein, the PORB protein was identified for the first time in dark-grown wheat. Altogether 64 unique proteins, representing pigment biosynthesis, photosynthetic light reaction, Calvin cycle proteins, chaperones and protein synthesis, were identified. The in number of proteins' largest group was the one involved in photosynthetic light reactions. This fact strengthens the assumption that the PLB membranes are precursors to the thylakoids and used for the formation of the photosynthetic membranes during greening. The present work is important to enhance our understanding of the significance of PLBs in chloroplast development.

Three-Dimensional Visualization of the Tubular-Lamellar Transformation of the Internal Plastid Membrane Network during Runner Bean Chloroplast Biogenesis

“…Although our study deals with the chloroplasts of an angiosperm, which differ from the chloroplasts of algae, it can be speculated that, during the thylakoid biogenesis of the angiosperm chloroplast, PLBs with connected PTs and the pyrenoid of the algae chloroplast can play a similar structural role (Rast et al, 2015). Regularity of PLBs in the bean plastids (Figure 1) was observed simultaneously with the band at 653 nm corresponding to the Pchlide-LPOR-NADPH complex ( Figure 9A) (Abdelkader et al, 2007;Blomqvist et al, 2008;Schoefs and Franck, 2008;Adam et al, 2011, and references therein). The transformation of PLBs observed after 1 h of illumination was seen as a loss of regular connections through interconnecting tubules (Figure 2).…”

“…We have demonstrated that the paracrystalline structure of the PLB transforms directly to lamellar structures that are porous from the beginning. This direct transformation can explain the presence of so many photosynthetic proteins in the PLB in the darkness, which results in photomorphogenic growth upon illumination (Blomqvist et al, 2008). Already at the beginning of the untangling of the paracrystalline network, tubules were transformed into porous flat slats even in the midst of the degrading PLB (Figure 2).…”

“…The longer wavelength forms are bound to PLBs, while this shorter wavelength form, unbound to LPOR, is mainly found in PTs. In addition to LPOR, PLBs contain enzymes of the chlorophyll and carotenoid biosynthesis pathways, enzymes of the Calvin cycle, and proteins involved in photosynthetic light reactions (subunits of ATP synthase, the oxygen-evolving complex, cytochrome b 6 f, plastocyanin, and ferrodoxin-NADPH oxidoreductase) and other proteins necessary during photomorphogenesis with the exception of the core subunits and the antenna complexes of the two photosystems (von Zychlinski et al, 2005;Kleffmann et al, 2007;Blomqvist et al, 2008;Adam et al, 2011).…”

Gain and Loss of Photosynthetic Membranes during Plastid Differentiation in the Shoot Apex of Arabidopsis

“…During this process, etioplasts, which contain prolamellar bodies (PLBs) and perforated lamellae called prothylakoids, are converted into functional chloroplasts that possess fully mature thylakoid networks. PLBs contain many of the building blocks of the thylakoid membrane, including MGDG, the chlorophyll precursor protochlorophyllide, enzymes involved in chlorophyll and carotenoid biosynthesis, and proteins involved in photosynthetic light reactions (Lonosky et al, 2004;Kleffmann et al, 2007;Blomqvist et al, 2008;Kanervo et al, 2008). This transition is therefore quite different from the one occurring at the shoot apex (from the L2 to the leaf primordia), both in its starting point (PLBs and prothylakoidcontaining etioplasts versus thylakoid-less proplastids) and its end point (fully mature versus partially mature networks).…”

Margareta Ryberg (1946–2012): a personal tribute

“…Sixty-four unique proteins were identified in prolamellar bodies, catalyzing pigment synthesis and various photosynthetic reactions. One POR protein, POR A, was found to dominate the proteome of prolamellar bodies, and POR B was found for the first time in dark-grown wheat (Blomqvist et al 2008). Margareta has over 60 publications on this topic in the Web of Science.…”