Synthesis and Deposition of Zein in Protein Bodies of Maize Endosperm

Abstract

The origin of protein bodies in maize (Zen mays L.) endosperm was investigated to determine whether they are formed as highly differentiated organefes or as protein deposits within the rough endoplasmic reticulum. Electron microscopy of developing maize endosperm cells showed that membranes surrounding protein bodies were continuous with rough endo-plasmic reticulum membranes. Membranes of protein bodies and rough endoplasmic reticulum both contained cytochrome c reductase activity indicating a similarity between these membranes. Furthermore, the proportion of alcohol-soluble protein synthesized by polyribosomes isolated from protein body or rough endoplasmic reticulum membranes was similar, and the alcohol-soluble or-insoluble proteins showed identical 114Clleucine labeling. These results demonstrated that protein bodies form simply as deposits within the rough eadoplasmic reticulum. Messenger RNA that directed synthesis of only the smaller molecular weight zein subunit was separated from mRNA that synthesized both subunits by sucrose gradient centrfuation. This result demonstrated that separate but similar sized mRNAs synthesize the major zein components. In vitro translation products of purified mRNAs or polyribosones were approximately 2,000 daltons larger than native zein proteins, suggesting that the proteins are synthesized as zein precursors. When intact rough endoplasmic reticulum was placed in the in vitro protein synthesis system, proteins corresponding in molecular weight to the native zein proteins were obtained. During development of cereal seeds, storage proteins are synthesized in the endosperm and deposited in protein bodies (5, 9, 19). Based on electron micrographs of developing wheat endo-sperm, Morton and co-workers (20) concluded that these protein bodies originated in specialized protein-forming organelles (plas-tids) which contained polyribosomes distinct from those of the general ER. An ultrastructural study of developing maize seeds led Khoo and Wolf (11) to conclude that the endosperm protein bodies originated in vesicles produced by RER or formed at the enlarged ends ofRER cisterna. Recent experiments have demonstrated that protein bodies in maize (3, 13) and barley endosperm (2, 6) contain the major prolamine proteins and that these proteins are synthesized by membrane-bound polyribosomes. In some of these studies membrane bound polyribosomes were isolated from the particulate fraction that pelleted between 500 and 37,000g2 (2, 6, 13). In another study polyribosomes were isolated from a protein body fraction obtained by continuous sucrose gradient centrifugation (3). Because of the attachment of ribosomes to the membrane surrounding protein bodies, several investigators concluded that 'Journal Paper No. 7052 of the Purdue University Experiment Station. 2 Abbreviation: g gravitational force determined at Ray. protein bodies are highly differentiated sites of storage protein synthesis (3, 20, 27), and furthermore that mRNAs isolated from "protein body" polyribosomes represent a homogene...

In vitro synthesis of barley storage proteins

“…Some mRNAs coding for some 'B' type polypeptides appear to b~ present. Similar results and conclusions have been presented for maize storage protein (zein) synthesis (Burr et aI, 1978;Larkins and Hurkman, 1978), and evidence presented that the zein mRNA product can be correctly proce3sed by a heterologous system i.e. Xenopus oocytes .…”

“…These results suggest that the proteins are synthesised as precursors. Larkins and Hurkman (1978) renorted that the products of polysome translation were larger than zein and co-migrated with products of mRNA translation. This observation is contrary to that of Very little work has so far been done on the in vitrosynthesis of wheat storage proteins but a situation similar to maize seems to exist for the hordeins of barley.…”

“…Ribosomes have been shown to be on the outside of the protein body membrane of maize (Burr and Burr, 1976;Larkins and Hurkman, 1978;Viotti at aI, 1978 Miflin et ~ (1980a) and Larkins and Hurkman (1978) suggests that cereal protein bodies are formed by the aggre8ation of storage proteins within the lumen of the endoplasmic reticulum. Ir.…”

Synthesis of Oat Globulin Precursors

“…Preliminary experiments on the immunoprecipitation of in vitro translation products with antibodies raised against avenin have shown the presence of polypeptides migrating near the authentic avenin, some with slightly higher mol wt (not shown), again suggesting synthesis as a precursor containing a signal sequence which is common to other cereal prolamins (5,6,8,12,16 In legumes, a precursor to llS legumin near 60,000 D is synthesized which is later cleaved between disulfide-linked acidic and basic polypeptides (7,22,24). In this respect, the oat globulin is similar.…”

In vivo and in vitro synthesis of CM-proteins (A-hordeins) from barley (Hordeum vulgare L.)

“…The study of protein synthesis and processing in the developing cereal endosperm is of interest both because of the central role played by this tissue in the nutrition of mankind and because of its valué as a plant model system (see Brandt and Ingversen 1978;Burr et al 1978;Larkins and Hurkman 1978;Mathews and Miflin 1980;Viotti et al 1979;Wienand and Feix 1978). Barley endosperm is particularly suitable for such a purpose because it can be easily obtained by extrusión from whole ears (O'Dell and Thompson 1982), and important advances have been made in the study of the genetic control of its main proteins (Dolí and Brown 1979;Shewry et al 1980a;Jensen et al 1980;Munck et al 1970;Dolí et al 1974).…”