Excision of Ds produces waxy proteins with a range of enzymatic activities.

Abstract

The waxy (wx) locus of maize encodes an enzyme responsible for the synthesis of amylose in endosperm tissue. The phenotype of the Dissociation (Ds) insertion mutant wx‐m1 is characterized by endosperm sectors that contain different levels of amylose. We have cloned the Wx gene from this allele and from two germinal derivatives, S5 and S9, that produce intermediate levels of amylose. The Ds insertion in wx‐m1 is in exon sequences, is 409 bp in length and represents an example of a class of Ds elements that are not deletion derivatives of the Activator (Ac) controlling element. The two germinal derivatives, S5 and S9, lack the Ds element but contain an additional 9 and 6 bp, respectively, at the site of Ds insertion. The level of Wx mRNA and Wx protein in S5 and S9 is essentially the same as in normal endosperm tissue but Wx enzymatic activity is reduced. Thus, the lesions in S5 and S9 lead to the addition of amino acids in the Wx protein, resulting in Wx enzymes with altered specific activities. This work supports the notion that the maize transposable elements may serve a function in natural populations to generate genetic diversity, in this case, proteins with new enzymatic properties.

The maize transposable Ds1 element is alternatively spliced from exon sequences.

“…Molecular cloning and characterization of the wx-ml mutation were reported previously (33). Relevant to this study is the fact that the allele contains a 409-bp Ds element in exon 9 of the Wx gene (Fig.…”

“…Comparison of these sequences with the wx-ml sequence (33) permitted the precise determination of 5' and 3' splice sites. Southern blot analysis of PCR products was as described previously (33).…”

“…Like wx-B4, this allele is null in the absence of Ac (14). The Ds insertion in wx-ml is in exon 9, is 409 bp in length, and is a Ds]-type element (33). Unlike the 4.3-kb Ds of wx-m9 and the 1.4-kb Ds of wx-B4, which are deletion derivatives of the Ac element (6,29), DsJ elements share only about 40 bp of homology with Ac sequences (22,27).…”

Interaction with amylopectin influences the ability of granule-bound starch synthase I to elongate malto-oligosaccharides

“…However, there is some evidence to suggest that GBSSI does interact with the amylopectin component of the starch granule. Studies of amylose-free mutants of several species which have inactive forms of GBSSI [22,23] have shown that GBSSI is still predominantly granule-bound, even though the granule lacks amylose. This suggests that the binding of wild-type GBSSI to starch granules may be largely or entirely due to an interaction with the amylopectin component rather than the amylose component of the granule.…”

“…The soluble extract was diluted 1 : 40 in gel sample buffer, heated to 90 mC for 2 min and loaded onto 6 % SDS/polyacrylamide gels. The gels were blotted on to nitrocellulose and the blots developed with antiserum to (A) pea GBSSI [22] (dilution : 1/2500) or (B) a truncated form of pea SSII [10] (dilution : 1/10 000). Lanes contained : 1, proteins from potato starch ; 2, E. coli cells transformed with pMUT6 and expressing mature potato GBSSI ; 3, E. coli cells transformed with pMUT13 and expressing mature pea SSII ; 4, E. coli cells transformed with the vector pET-3 alone ; and 5, proteins from pea starch.…”