Delivery of Prolamins to the Protein Storage Vacuole in Maize Aleurone Cells

Abstract

Zeins, the prolamin storage proteins found in maize (Zea mays), accumulate in accretions called protein bodies inside the endoplasmic reticulum (ER) of starchy endosperm cells. We found that genes encoding zeins, a-globulin, and legumin-1 are transcribed not only in the starchy endosperm but also in aleurone cells. Unlike the starchy endosperm, aleurone cells accumulate these storage proteins inside protein storage vacuoles (PSVs) instead of the ER. Aleurone PSVs contain zeinrich protein inclusions, a matrix, and a large system of intravacuolar membranes. After being assembled in the ER, zeins are delivered to the aleurone PSVs in atypical prevacuolar compartments that seem to arise at least partially by autophagy and consist of multilayered membranes and engulfed cytoplasmic material. The zein-containing prevacuolar compartments are neither surrounded by a double membrane nor decorated by AUTOPHAGY RELATED8 protein, suggesting that they are not typical autophagosomes. The PSV matrix contains glycoproteins that are trafficked through a Golgi-multivesicular body (MVB) pathway. MVBs likely fuse with the multilayered, autophagic compartments before merging with the PSV. The presence of similar PSVs also containing prolamins and large systems of intravacuolar membranes in wheat (Triticum aestivum) and barley (Hordeum vulgare) starchy endosperm suggests that this trafficking mechanism may be common among cereals.The cereal endosperm consists of three main cell types: an inner mass of starchy endosperm cells, one to three layers of epidermal aleurone cells, and the transfer cells that contact the maternal vascular tissue (Olsen, 2004). The starchy endosperm accounts for 80 to 90% of the grain weight and contains large amounts of storage proteins and starch. These cells undergo programmed cell death during maturation. Aleurone cells are rich in protein storage vacuoles (PSVs), minerals, and lipid bodies and remain alive during seed development. It is assumed that the breakdown of proteins localized to PSVs in aleurone cells provides an essential source of the amino acids necessary for the synthesis of hydrolytic enzymes required for mobilizing food stored in the starchy endosperm (Filner and Varner, 1967;Jacobsen et al., 1988;Bethke et al., 1998).Besides its biological relevance as a model to study plant development, cell differentiation, and programmed cell death, the cereal endosperm is very important in terms of its nutritional value. Cereal grains contain less protein than do legume seeds, but because cereals are produced and consumed in much larger quantities, they are the main source of protein for the nutrition of humans and livestock (Shewry and Halford, 2002). The major storage proteins in maize (Zea mays) kernels are the alcoholsoluble prolamins, a type of storage proteins present only in grasses. Maize prolamins, referred to as zeins, are divided into different types: a-, b-, g-, and d-zeins (Coleman and Larkins, 1999) that differ in amino acid composition and structural properties (Shewry and Halford, 2002)...

The ATG1/ATG13 Protein Kinase Complex Is Both a Regulator and a Target of Autophagic Recycling in Arabidopsis

“…In plants, autophagy is upregulated under nutrient-limiting conditions to help replenish internal supplies of fixed nitrogen (N) and carbon (C) to support continued biosynthesis and energy production and during developmentally programmed cell death and senescence to encourage nutrient remobilization (Doelling et al, 2002;Bassham, 2009;Reyes et al, 2011). It also promotes survival during pathogen invasion by helping orchestrate the hypersensitive response, whereby host plants undergo localized cell death to discourage pathogen spread Hofius et al, 2009;Yoshimoto et al, 2009;Lenz et al, 2011).…”

A BAR-Domain Protein SH3P2, Which Binds to Phosphatidylinositol 3-Phosphate and ATG8, Regulates Autophagosome Formation in Arabidopsis

“…However, structures that are necessary for autophagosome formation in plants have remained elusive. Double-membrane structures, morphologically recognized as autophagosomes, had been reported in plants but had not been validated (Rose et al, 2006;Katsiarimpa et al, 2011;Reyes et al, 2011;Minibayeva et al, 2012). In our investigation, SH3P2-positive structures in Arabidopsis share several features with autophagosomes in mammalian cells: (1) They arise from the isolation membrane as well as from omegasome-like structures and subsequently wrap together into bi/multilayer-membrane autophagosome structures (Figures 4 and 5A; see Supplemental Figure 4 online); (2) they can expand and mature by fusing with endosomes or by engulfing cytoplasmic cargos ( Figures 5B and 5C); and (3) they reacted positively with the autophagosome marker ATG8 (Figures 2, 6A, and 6C).…”

“…In plants, however, most observations of autophagosome-related structures have been based on the classical features of autophagosomes: their isolated double-membrane appearance, enclosed autophagic cargos, and labeling with an autophagosome marker (Rose et al, 2006;Toyooka et al, 2006;Katsiarimpa et al, 2011;Takatsuka et al, 2011;Hanamata et al, 2012). So far, only one electron microscopy (EM) study has shown direct labeling of a double-membrane autophagosome structure in Arabidopsis thaliana with antibodies against the autophagosomal marker ATG8 (Reyes et al, 2011). Despite this single study, investigations on autophagosome biogenesis in plants have yet to reveal the detailed steps involved in this process and well defined intermediate structures.…”

Traffic of Human α-Mannosidase in Plant Cells Suggests the Presence of a New Endoplasmic Reticulum-to-Vacuole Pathway without Involving the Golgi Complex

“…Moreover, the direct transport of proteins from the ER to the vacuole can be adopted by cells in particular physiological situations such as seed germination (Chrispeels and Herman, 2000) or apoptosis (Hayashi et al, 2001). A recent study has provided evidence for another route in the aleurone layer of maize (Zea mays) seeds that transports zeins, a-globulin, and legumin-1 directly from the ER to the PSV without involving the Golgi apparatus (Reyes et al, 2011). The authors suggested the existence in cereals of an atypical autophagic process that delivers ER proteins directly to PSVs, sequestering them into complex prevacuolar compartments.…”

“…In addition, the route that bypasses the Golgi system seems to be linked to the specific transport of proteins that form large aggregates (Herman and Schmidt, 2004;Herman, 2008). Cereal prolamins, when aggregated in the ER in large polymers, can also be transported directly from the ER to PSVs, apparently by autophagy (Levanony et al, 1992;Reyes et al, 2011). Moreover, many vacuolar enzymes are stored in ER-derived vesicles, which, under certain circumstances such as programmed cell death or seed germination, are directly fused with the vacuolar compartment (Hayashi et al, 2001;Rojo et al, 2003).…”