Defective endosperm* (De*)-B30 is a dominant maize (Zea mays) mutation that depresses zein synthesis in the developing endosperm. The mutant kernels have an opaque, starchy phenotype, malformed zein protein bodies, and highly increased levels of binding protein and other chaperone proteins in the endosperm. Immunoblotting revealed a novel ␣-zein protein in De*-B30 that migrates between the 22-and 19-kD ␣-zein bands. Because the De*-B30 mutation maps in a cluster of 19-kD ␣-zein genes, we characterized cDNA clones encoding these proteins from a developing endosperm library. This led to the identification of a 19-kD ␣-zein cDNA in which proline replaces serine at the 15th position of the signal peptide. Although the corresponding gene does not appear to be highly expressed in De*-B30, it was found to be tightly linked with the mutant phenotype in a segregating F 2 population. Furthermore, when the protein was synthesized in yeast cells, the signal peptide appeared to be less efficiently processed than when serine replaced proline. To test whether this gene is responsible for the De*-B30 mutation, transgenic maize plants expressing this sequence were created. T 1 seeds originating from the transformants manifested an opaque kernel phenotype with enhanced levels of binding protein in the endosperm, similar to De*-B30. These results are consistent with the hypothesis that the De*-B30 mutation causes a defective signal peptide in a 19-kD ␣-zein protein.A number of mutations have been identified that affect storage protein synthesis in maize (Zea mays) endosperm (for review, see Coleman and Larkins, 1998). The mutant kernels typically have a starchy endosperm texture and low density, both of which are thought to be associated with the reduced synthesis of the prolamin storage proteins, or zeins. When placed on a light box, endosperm of these mutants does not transmit light, i.e. it is opaque, whereas that of normal, wild-type kernels is vitreous and translucent. Recessive mutations that produce such an opaque kernel phenotype are traditionally classified as "opaque" (e.g. opaque1-15), whereas semidominant opaque mutations are called "floury" (e.g. floury1-3). There are, in addition, several dominant opaque mutations, defective endosperm* (De*)-B30 and Mucuronate (Mc;Soave et al., 1979;Soave and Salamini, 1984). The basis of the relationship between an opaque kernel phenotype and zein synthesis is not well understood. Although several of these mutants have a reduced zein content and show altered expression of specific types of zein genes, the degree to which zein synthesis is reduced among them is variable (Hunter et al., 2002). For example, in o1, zein synthesis is barely affected. Generally, these mutations result in increased expression of genes associated with physiological stress and the unfolded protein response (UPR; Kaufman, 1999) during endosperm development.Because of their influence on zein synthesis and the nature of their pleiotropic effects on other genes expressed in the endosperm, the genetic defects re...