Interallelic complementation leading to restoration of the normal phenotype in hetero-allelic combinations of some of the EMS induced sh mutants was reported previously (Chourey, 1971, Genetics 68:435). These studies were, however, restricted to the phenotype and the protein level as the enzymatic nature of the gene product was not known. Recent observations (Chourey and Nelson, 1976, Biochem. Genet. 14:1041) showing the product of the Sh locus (Sh protein) to be the enzyme sucrose synthetase has enabled us to examine some of the EMS induced sh mutants and their hybrids at the enzyme level (Table 1). The following observations are noteworthy:
The mutants sh-F and sh-C are similar to the reference allele of sh mutant (W22 background) as regards amounts of sucrose synthetase activity. The same is true of sh-S if its sucrose synthetase activity is assayed in the direction of sucrose degradation (in the presence of UDP). The genetic basis of this residual enzyme activity in these three mutants is not known at this time, as each of sh-F, sh-C and sh-S is known to code the sh locus specific protein (Chourey and Schwartz, 1971, Mutation Res. 12:151). The sh-S mutant, however, is genetically as well as biochemically unique as compared to all the sh mutants that we have studied so far. The enzyme specified by this mutant showed as much as 30% (as compared to the normal endosperm) activity when assayed in the direction of sucrose synthesis. The protein coded by sh-S locus is probably responsible for most of this enzyme activity. The sucrose synthesis and sucrose cleavage properties of sucrose synthetase are believed to be due to the same protein molecule in various plants including maize. Thus, it seems, there is a differential alteration in the rates of sucrose synthesis and sucrose cleavage in the enzyme molecule coded by the sh-S mutant. The occurrence of such a qualitative change in the enzyme in the sh mutant, which was initially selected on the basis of its shrunken phenotype, further substantiates that the Sh locus is the structural gene for sucrose synthetase.
The phenotype of the F1 hybrid kernel produced by crossing sh-S with either sh-C or sh-F is indistinguishable from the wild type (Sh) kernel. No other hetero-allelic combination of sh mutants complements to produce the wild type phenotype. The endosperm extracts of the complementing hybrids (sh-S x sh-F or sh-S x sh-C) when assayed for sucrose synthesis showed activities that were lower than the sh-S but were higher than the sh-F or sh-C homozygotes. Thus the complementing heterozygotes, which have a wild type phenotype, have a lower enzyme activity than one of the parental types which is mutant in phenotype. However, when the same hybrids were assayed for sucrose cleavage the results were different. The complementing heterozygotes show a distinct elevation in the enzyme activity as compared to the parental types. A slight elevation is also observed for the noncomplementing heterozygote, viz. sh-F x sh-C. It is possible that the critical limit of the enzyme activity to restore the wild type phenotype lies somewhere between the values 7% and 10%, as compared to the wild type.
The physiological mode of sucrose synthetase reaction, i.e., whether sucrose cleavage or sucrose synthesis is the in vivo reaction in a developing kernel, is controversial. The following observations of this investigation are interpreted to suggest that sucrose cleavage is the critical in vivo reaction to mobilize photosynthesized sucrose for starch biosynthesis. It is possible that the sh-S mutant is shrunken in phenotype because of a significant reduction in the catalysis of the sucrose cleavage. Note that the rate of sucrose synthesis is not limiting as it is even higher in this mutant than in the complementing hybrids which have a wild type phenotype. In other organisms, interallelic complementation leading to wild type phenotype has been associated with an elevation in the enzyme activity as compared to the parental types. It is observed that such an elevation takes place in the hybrids only when the enzyme activity was assayed for sucrose cleavage.
Prem S. Chourey* and Oliver E. Nelson
*Department of Agronomy, University of Illinois, Urbana, IL 61801
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