Protein kinases are an abundant class of enzymes which play important roles in numerous signal transduction systems. Arabidopsis TOUSLED kinase is a serine/threonine kinase which is essential for cell-cell communication within the shoot meristem. TOUSLED is encoded by a single gene in Arabidopsis. Recessive mutants in this gene show mild vegetative defects and severe floral abnormalities including a random reduction in the number of floral organs produced and defects in the formation of the gynoecium. This thesis describes the cloning and characterization of three maize genes with homology to TOUSLED. These genes are known as the TOUSLED-LIKE KINASE (TLK) genes. Partial genomic and cDNA clones of the maize TLK genes have been sequenced and analyzed. These genes show remarkable similarity to each other and to TOUSLED over the region corresponding to the TOUSLED catalytic domain. The TLK genes fall into two distinct classes on the basis of nucleotide and amino acid sequence. Both classes appear to be expressed throughout the plant. In addition, database searches reveal that TOUSLED-like genes are present in a diverse array of other eukaryotes, indicating that the TLK genes are members of a widespread, evolutionarily conserved class. Two approaches have been taken to find mutants in the TLK genes. This thesis describes the tassel-less1 (tls1) mutant, a possible mutant in one of the TLK genes. The tls1 mutation maps to the same chromosomal location as one of the TLK genes, and may represent a lesion in this TLK gene. Characterization of the t1s1 mutant reveals that disruption of the TLS1 gene results in variable, progressive vegetative defects and severe reduction of the reproductive structures. The tls1 phenotype is consistent with the hypothesis that TLS1 plays a role in regulating meristem activity. In addition, TLK sequences have been used in a reverse genetics screen to isolate families which contain Mutator transposable element insertions into the TLK genes. These families are currently being analyzed for phenotype and allelism to tls1. / Graduation date: 1997

Experiments were performed to assess regulatory factors governing maize embryo maturation and vivipary. Both visual and molecular markers of embryo development were used to examine the roles of the hormones abscisic acid (ABA) and gibberellins (GAs), as well as water stress in governing transit from early embryogeny to maturation-phase development. A differential screen identified cDNAs whose expression is impaired in maize viviparous mutants which fail to undergo maturation and instead precociously germinate. The cDNAs isolated in this screen absolutely required both ABA and the Viviparousl (Vpl) gene product for expression both in vivo and in vitro. Two novel clones were isolated: a maize homologue of the wheat metallothionein gene E[subscript]c and a second clone which may encode a novel seed storage protein of maize. In a separate screen, a maize cDNA encoding a Lea group 3 protein was isolated. Like many maturation-associated genes, maize Lea 3 was shown to ABA-inducible but is also expressed in response to water stress in the absence of ABA or the Vp 1 gene. We examined whether gibberellins might also be a factor modulating precocious germination. Gibberellin inhibitors applied to cultured wildtype embryos suppressed precocious germination and enhanced anthocyanin accumulation in a developmentally specific manner. These behaviors mimicked the effect of ABA and they were reversed by the addition of exogenous GA���. Vivipary in vivo resulting from diminished ABA levels could be suppressed by either chemical or genetic reduction of GA levels in immature kernels and resulted in desiccation-tolerant seed. In contrast, reduction of endogenous gibberellins did not suppress vivipary of the ABA-insensitive mutant vp1. Temporal analysis of gibberellin accumulation in developing kernels revealed the accumulation of two bioactive species (GA��� and GA���) during a developmental window just prior to peak ABA levels. It is suggested that these species stimulate a developmental program leading to vivipary in the absence of sufficient levels of ABA and that reduction of GA levels reestablishes a hormone balance appropriate for suppression of germination and induction of maturation in ABA-deficient kernels. The failure to suppress vivipary via reduction of GA levels in the ABA-insensitive mutant vp1 suggests that the wildtype gene product functions downstream of the sites of GA and ABA action in regulation of maturation versus germination. / Graduation date: 1995

The nucleotide sequence of a cloned RAPD DNA marker (OPI08) linked to a disease resistance gene in L. sativa (lettuce) revealed homology with the conserved domain of the reverse transcriptase of Ty3/gypsy retrotransposons. To further characterize the presence of Ty3/gypsy-like sequences in plants, sets of degenerate primers deduced from archetype retrotransposons were used for PCR amplification of a sequence domain characteristic of the reverse transcriptase and the integrase of Ty3/gypsy retrotransposons. The nucleotide sequence of two cloned DNA fragments of Z. mays (maize) and A. thaliana proved to be homologous with the conserved domains of the reverse transcriptase and the integrase of Ty3/gypsy retrotransposons. Southern blot analysis also demonstrated homology of the Z. mays clone to Lycopersicon (tomato) and L. sativa. This is the first report of Ty3/gypsy-like sequences in A. thaliana, and L. sativa. This research brings to six the number of plant species where this type of element has been reported, in contrast to the large number of plant Ty1/copia transposable elements described. It is not known whether these elements are actively transposing in plant genomes.

No description available.