A maize ribosome-inactivating protein is controlled by the transcriptional activator Opaque-2.

Abstract

Although synthesis of the cytosolic maize albumin b-32 had been shown to be controlled by the Opaque-2 regulatory locus, its function was unknown. We show here that b-32 is a member of the large and widely distributed class of toxic plant proteins with ribosome-inactivating activity. These ribosome-inactivating proteins (RIPs) are RNA N-glycosidases that remove a single base from a conserved 28s rRNA loop required for elongation factor l u binding. Cell-free in vitro translation extracts were used to show that both maize and wheat ribosomes were resistant to molar excesses of b-32 but not to the dicotyledonous RIP gelonin. We extracted RIP activity from kernels during seed maturation and germination. The amount of RIP activity increased during germination, although the amount of b-32 protein remained fairly constant. Expression of a maize RIP gene under the control of an endosperm-specific transcriptional regulator may be an important clue prompting investigation of the biological basis for RIP expression in seeds of other plants. INTRODUCTIONRibosome-inactivating proteins (RIPs) are a widely distributed group of toxic plant proteins that catalytically inactivate eukaryotic ribosomes (for review, see Stirpe and Barbieri, 1986). RlPs function as N-glycosidases to remove a specific adenine in a conserved loop of the large rRNA . This irreversible modification renders the ribosome unable to bind elongation factor l a , thereby blocking translation. Because this translational inhibitory activity is toxic, RlPs have been tested extensively for use as immunotoxins and antiviral agents and more recently for their effects on protozoa, insects, and fungi (Barbieri and Stirpe, 1982;Cenini et al., 1988;Gatehouse et al., 1990;Leah et al., 1991).RIP activities have been found in the seed, root, leaf, or sap of more than 50 different plant species (Gasperi-Campani et al., 1985). Two forms of RlPs have been described (Stirpe and Barbieri, 1986). Type 1 RlPs such as pokeweed antiviral protein, trichosanthin, the barley translation inhibitor, and gelonin are monomeric enzymes, each with an approximate M, of 30,000 (Irvin, 1975;Stirpe et al., 1980;Asano et al., 1984;Maraganore et al., 1987;Yeung et al., 1988). Type 2 RlPs such as ricin, abrin, and modeccin are highly toxic heterodimeric proteins, each with an approximate M, of 60,000 in which one polypeptide with RIP activity (A-chain) is linked by a disulfide bridge to a galactose-binding lectin (B-chain; Pihl, 1973, 1982;Stirpe et al., 1978).Type 1 RlPs and the A-chain of type 2 RlPs have basic isoelectric points, and many have signal peptides that are not To whom correspondence should be addressed. present in the mature protein (Stirpe and Barbieri, 1986). Although RlPs share biological activity, they typically exhibit similarities of <50%, and antibodies raised against RlPs seldom cross-react with RlPs from distantly related species (Ready et al., 1988). The maize b-32 protein has homology with severa1 previously characterized RIPs, yet it is a singlechain acidic protein tha...

The barley 60 kDa jasmonate‐induced protein (JIP60) is a novel ribosome‐inactivating protein

“…In common with all characterized plant RIP the toxicity of this JIP60 derivative is associated with depurination of reticuIocyte 28S rRNA (Bass et al, 1992;May et al, 1989). Like the Type I endosperm RIP, JIP60 and derivatives do not significantly affect translation by wheat germ extract, suggesting that JIP60 is not toxic to plant cells (Stirpe and Hughes, 1989).…”

“…First, JIP60 and maize RIP differ from the Type I and II RIP in containing extra, internal peptides of 20-25 amino acids. Removal of this linker peptide is required for activation of one of the maize RIPs (Bass et al, 1992;Walsh eta/., 1991). Second, JIP60 and ricin differ from the maize and Type I RIP in containing large C-terminal regions.…”

“…Recently, a third RIP type has been characterized from maize. It is apparently not secreted but is synthesized as a 34 kDa precursor which requires removal of an internal peptide for activation (Bass et aL, 1992;Walsh etaL, 1991).…”

γ-Zeins are essential for endosperm modification in quality protein maize

“…It has been shown that O2 regulates more genes than just the 22-kDa α-and β-zeins (39,40). Furthermore, the QPM phenotype comprises several QTLs, which make quantitative contributions (24).…”

JIPs and RIPs: the regulation of plant gene expression by jasmonates in response to environmental cues and pathogens.

“…The N-terminal half of JIP60 is related to both type I and type II RIPs, such as the Mirabilisjalapa antiviral protein (Habuka et al, 1989), saporin-6 from Saponaria officinalis (Benatti et al, 1989), and ricin from Ricinuscommunis (Lambet al, 1985) (see Reinbothe et al, 1994, for comparison). Whereas type I RlPs are basic proteins that consist of a single polypeptide chain of 25 to 32 kD (for references, see Bass et al, 1992), type II RlPs contain, in addition to the N-terminal RIP domain, a second domain that confers lectinlike properties to these proteins Pihl, 1973, 1982). 60th types of RIP are exceptionally potent inhibitors of eukaryotic protein synthesis (Coleman and Roberts, 1982;Stirpe et al, 1988).…”