Prem S. Chourey

Callus formation from protoplasts of a maize cell culture.

SummaryA finely dispersed cell suspension culture from the friable callus of the ‘Black Mexican Sweet’ line of maize was obtained. Protoplasts from this cell culture, when grown in a simplified medium described here, showed sustained cell divisions and gave rise to callus.

Genetic evidence that invertase-mediated release of hexoses is critical for appropriate carbon partitioning and normal seed development in maize

Abstractu2002Cell wall-bound invertase (CWI) is spatially and temporally the first enzyme which metabolizes the incoming sucrose in developing seed of maize (Zea mays). Our previous studies have shown that the cell wall-bound invertase-2 (INCW2) isozyme encoded by the wild-type gene of the Miniature1 (Mn1) seed locus plays a critical role in seed development. Null mutations of the gene, such as the mn1 seed mutant which lacks invertase activity, are associated with a loss of ∼70–80% of the normal seed weight. We show here that under in vitro kernel culture conditions the hexose-based medium was similar to the sucrose-based medium in promoting the normal development of kernels of the Mn1, but not of the mutant mn1, genotype. Anatomical, biochemical, and immunohistological data showed that the mn1 kernels retain their mutant phenotype regardless of the presence of sucrose or hexoses in the culture media. The most drastic changes in the mn1 seed mutant were associated with a significant reduction in the size of the endosperm, but not in the pattern or the level of starch localization. Because Mn1 expression was temporally coincident with the endosperm cell divisions, INCW2 must play a critical role in providing hexose sugars for mitotic division, and only a minor role in generating carbon skeletal substrates for starch biosynthesis in the early stages of endosperm development. Furthermore, a lack of the wild-type seed phenotype of the mn1 mutant in hexose media suggests that a metabolic release of hexoses catalyzed by INCW2, rather than an exogenous source, is critical for both generating appropriate sugar-sensing signals for gene expression and for normal endosperm development.

Anaerobic treatment alters the cell specific expression of Adh-1, Sh, and Sus genes in roots of maize seedlings

SummaryThe expression pattern of Sh and Sus, which encode sucrose synthase isozymes SS1 and SS2, respectively, and Adh-1, which encodes alcohol dehydrogenase 1 (ADH1), was examined in situ in the lower region of the primary root of maize seedlings in response to anaerobiosis. Anaerobic induction of ADH1 RNA was localized to the epidermis and cortex. Induction of Sh was marked by highly elevated SS1 RNA levels in the vascular elements, pith, and epidermis. A significant but less drastic increase in SS protein was found in these same tissues as well as the root cap; the increased level of immunosignal was restricted to cells within approximately 1 cm of the root apex. Sus responded to anaerobic stress with a slight reduction in SS2 RNA and protein levels except in the root cap where SS2 protein, but not SS2 RNA, was induced. Thus, in situ hybridization and/or immunolocalization experiments revealed a unique spatial pattern of expression for each of the three genes in response to anaerobiosis. Furthermore, we conclude that multiple regulatory controls including cell specific post-transcriptional mechanisms modulate SS levels in anaerobically stressed seedlings.

A comparative study on the role of cytokinins in caryopsis development in the maize miniature1 seed mutant and its wild type.

We report here on a comparative developmental profile of plant hormone cytokinins in relation to cell size, cell number and endoreduplication in developing maize caryopsis of a cell wall invertase-deficient miniature1 (mn1) seed mutant and its wild type, Mn1, genotype. Both genotypes showed extremely high levels of total cytokinins during the very early stages of development, followed by a marked and genotype specific reduction. While the decrease of cytokinins in Mn1 was associated with their deactivation by 9-glucosylation, the absolute and the relative part of active cytokinin forms was higher in the mutant. During the exponential growth phase of endosperm between 6 d after pollination and 9 d after pollination, the mean cell doubling time, the absolute growth rate and the level of endoreduplication were similar in the two genotypes. However, the entire duration of growth was longer in Mn1 compared with mn1, resulting in a significantly higher cell number in the Mn1 endosperm. These data correlate with the previously reported peak levels of the Mn1-encoded cell wall invertase-2 (INCW2) at 12 d after pollination in the Mn1 endosperm. A model showing possible crosstalk among cytokinins, cell cycle and cell wall invertase as causal to increased cell number and sink strength of the Mn1 developing endosperm is discussed.

Electrophoretic analysis of sucrose synthetase proteins in the complementing heterozygotes at the Shrunken locus in maize.

SummaryElectrophoretic comparisons of sucrose synthetase (SS) proteins in complementing heterozygotes and the corresponding in vitro mixtures of extracts from the homozygotes are described. The latter revealed two protein bands in the expected fashion. The SS protein pattern in the hybrid was different from that of the mixtures. The possibility of heteromeric SS molecules, formed by random polymerization of subunits of the tetramer coded by each allele in the heterozygote, was considered. Such an interaction was expected to form a multiple of five SS proteins that could be visualized after gel electrophoresis. However, only two SS bands were seen in the hybrids. The basis of this marked deviation remains to be explained.

Cytokinins and Their Possible Role in Seed Size and Seed Mass Determination in Maize

One of the richest sources of CKs in various plants species are developing seeds. In fact, zeatin, a major natural CK, was first discovered in developing seeds of maize (Letham, 1963). Not surprisingly, developing seeds of maize, rice and Lupinus albus have played a major role in CK research (reviewed in Emery & Atkins, 2005). A major focus of these studies has been on the possible role of CKs in sink strength of developing seeds. The role of CKs in controlling sink strength is inferred largely from (a) the well known role of CK in stimulating cell division that may lead to increased organ size of the sink tissue, and (b) the coincidence of CK accumulation with seed cell division profiles in several plant species (Dietrich et al., 1995; Arnau et al., 1999; Yang et al., 2002).