M. Sari Gorla

Organisation and structural evolution of the rice glutathione S-transferase gene family

Glutathione S-transferases (GSTs) comprise a large family of key defence enzymes against xenobiotic toxicity. Here we describe the comprehensive characterisation of this important multigene family in the model monocot species rice [Oryza sativa (L.)]. Furthermore, we investigate the molecular evolution of the family based on the analysis of (1) the patterns of within-genome duplication, and (2) the phylogenetic relationships and evolutionary divergence among rice, Arabidopsis, maize and soybean GSTs. By in-silico screening of the EST and genome divisions of the Genbank/EMBL/DDBJ database we have isolated 59 putative genes and two pseudogenes, making this the largest plant GST family characterised to date. Of these, 38 (62%) are represented by genomic and EST sequences and 23 (38%) are known only from their genomic sequences. A preliminary survey of EST collections shows a large degree of variability in gene expression between different tissues and environmental conditions, with a small number of genes (13) accounting for 80% of all ESTs. Rice GSTs are organised in four main phylogenetic classes, with 91% of all rice genes belonging to the two plant-specific classes Tau (40 genes) and Phi (16 genes). Pairwise identity scores range between 17 and 98% for proteins of the same class, and 7 and 21% for interclass comparisons. Rapid evolution by gene duplication is suggested by the discovery of two large clusters of 7 and 23 closely related genes on chromosomes 1 and 10, respectively. A comparison of the complete GST families in two monocot and two dicot species suggests a monophyletic origin for all Theta and Zeta GSTs, and no more than three common ancestors for all Phi and Tau genes.

The extent of gametophytic-sporophytic gene expression in maize

SummaryTo determine the extent of gametophytic gene expression and the type of transcription, haploid or haplo-diploid, of the genes, isozymes were used as genetic markers. Fifteen enzymatic systems, including thirty-four isozymes, were studied. The determination of the type of expression of genes coding for multimeric enzymes was based on the comparison of electrophoretic patterns of pollen and of sporophytic tissues from plants heterozygous for electrophoretic mobility: if gene expression in pollen is of a gametophytic (haploid) origin, pollen, unlike the sporophyte, would reveal only the parental homomultimeric bands. The enzymes analyzed can be grouped in three categories according to type of gene expression: i) enzymes present in both pollen and sporophyte, coded by the same gene with haplo-diploid expression; ii) enzymes controlling analogous functions in pollen and sporophyte, coded by different genes, expressed in only one of the two phases; iii) enzymes present in two or more forms in the sporophyte and only in one form in the gametophyte. The data allow the proportion of haplo-diploid gene expression in the loci examined to be estimated at 0.72; 0.22 and 0.06 being the proportions attributable to the sporophytic and gametophytic domains, respectively.

Male gametophytic selection in maize

SummaryThere is evidence that male gametophyte selection is a widespread phenomenon in higher plants. The pollen tube growth rate is one of the main components of gametophyte selective value; genetic variability for this trait, due to the effect of single genes or to quantitative variation, has been described in maize. However, indication of gametophytic selection has been indirectly obtained; its effect was revealed by the positive relation observed between gametophyte competitive ability and sporophyte metrical traits.This paper considers the results of selection applied to gametophyte populations produced from single plants. The competitive ability of the lines was evaluated in comparison with that of a standard line by means of the pollen mixture technique. Sporophytic traits were measured in the hybrid progeny obtained by crossing selected S3 and S4 families with an unrelated single cross and an inbred line. Gametophyte selection produced inbred lines with high gametophyte competitive ability. In view of the selection procedure adopted, this result was interpreted as an indication of haploid expression of genes involved in the control of pollen tube growth. Moreover, this gametophytic trait was positively correlated with sporophytic traits (seedling weight, kernel weight and root tip growth in vitro), indicating that both groups of characters have a common genetic basis.

Molecular markers (RFLPs and HSPs) for the genetic dissection of thermotolerance in maize.

SummaryCellular membrane stability (CMS) is a physiological index widely used to evaluate thermostability in plants. The genetic basis of the character has been studied following two different approaches: restriction fragment length polymorphism (RFLP) analysis, and the effects of segregating heat shock protein (HSP) loci. RFLP analysis was based on a set of recombinant inbreds derived from the T32 × CM37 F1 hybrid and characterized for about 200 RFLP loci. Heritability of CMS estimated by standard quantitative analysis was 0.73. Regression analysis of CMS on RFLPs detected a minimum number of six quantitative trait loci (QTL) accounting for 53% of the genetic variability. The analysis of the matrices of correlation between RFLP loci, either within or between chromosomes, indicates that no false assignment was produced by this analysis. The effect of HSPs on the variability of the CMS was tested for a low-molecular-weight peptide (HSP-17) showing presence-absence of segregation in the B73 × Pa33 F2 population. Although the genetic variability of the character was very high (h2=0.58) the effect of HSP-17 was not significant, indicating either that the polypeptide is not involved in the determination of the character or that its effect is not statistically detectable.

Genetic Variability of Gametophyte Growth Rate in Maize

SummaryIn order to measure differences in the pollen growth rate of numerous lines of maize and to investigate the main features of their genetic control, gametophyte growth was studied in vitro and in vivo. In vitro pollen tube growth of twenty inbred lines and seven hybrids was measured; a remarkable variability was observed in the growth rate of the inbred lines examined: most lines were distinct, showing different levels of growth.Analysis of frequency distribution of pollen tube lengths for pairs of inbred lines and their F1′ s revealed greater variance among lengths of F1 pollen tubes, presumably indicating the segregation of genetic factors expressed in the gametophyte.Similar frequency distributions of tube lengths in pollen produced by two pairs of reciprocal hybrids virtually excluded the presence of a cytoplasmic component. In vivo competitive ability of pollen tubes was measured as the increase in relative fertilization frequency from apex to base of the ear. Mixtures were made using two types of genetically distinguishable pollen, and were applied to a female common parent. Nine pairs of inbred lines furnished the pollen for the mixtures. In all cases where the B14 line was involved, this pollen type fertilized nearly all the ovules, perhaps indicating the presence of a gametophytic factor. When other lines were compared, the ears contained mixtures of the two possible seed types, the relative proportions of which indicated the differential competitive abilities of the two pollen tube types.A comparison between in vitro and in vivo behavior was made for some genotypes. In vivo results generally agreed with in vitro results. The degree of the differences between lines however was changed, presumably because pollen-style or pollen-pollen interactions are absent in vitro.Differing growth patterns between lines were also revealed in vivo by direct observation of fluorescent pollen tubes within the silks, a finding which may be useful in further studies.

Herbicide-tolerant corn by pollen selection

SummaryMaize pollen was exposed to the herbicide Chlorsulfuron (CS), and segregation for tolerance was observed. The resulting plant generation exhibited significantly greater tolerance to CS than other (control) progeny. Since the increase in tolerance occurred after only one generation of pollen exposure, this result demonstrates that pollen selection can be used to develop herbicide-resistant crop species, even when the species are not amenable to cell culture. It also suggests a possible mechanism for the rapid evolution of herbicide tolerance in weeds.

Male Gametophytic Selection in Higher Plants

Gametophytic selection refers to the differential gene transmission during the haploid phase in higher plants, which begins with the meiosis product and ends with fertilization. The special features of the male gametophyte (large population size, independence of the maternal plant, direct exposure to environmental stresses, competition within the same style) suggest that selection in this phase may be expected to act with greater intensity then in the female gametophyte.

Haplodiploid gene expression in maize and its detection

A method for the demonstration of the gametophytic origin of genetic variability in maize is described. For genes coding dimeric or multimeric enzymes, haploid expression can be demonstrated by means of translocations between A and B chromosomes (TB-A), which make it possible to obtain hyperploid pollen grains, partially diploid and heterozygous for electrophoretic mobility. Comparison of the electrophoretic pattern of this pollen type (three bands) and that of normal grains produced by a heterozygous F/S plant (two bands only) reveals haploid transcription of the monomeric forms. The procedure was tested on ADH-1 and used to demonstrate haploid expression for GOT-1. The data obtained suggest, moreover, that the reduction in male gamete transmission of duplications may be due to differences in pollen competitive ability rather than to processes affecting microspore maturation.

Chromosomal and extrachromosomal transmission of cellular melanization in the Freckled phenotype in Drosophila melanogaster.

Freckled ( Frd ) is a dominant mutant of D. melanogaster , lethal in double dose, which manifests itself by melanization in the adult stage, while nothing can be noticed in the larva and/or in the pupa. Its localization has been determined at nearly 102 of the second chromosome. Among the offspring from backcrosses involving Frd , some individuals that do not carry the Frd gene exhibit a weak Frd manifestation in the adult stage, or a stronger one, mostly lethal, in the larval or pupal stage. Weak Frd , indicated as [ Frd ], is transmitted indefinitely without any recognizable linkage with the chromosomes, although its frequency is influenced by the 1st and, probably, by the 3rd chromosome. No matroclinous transmission has been found. It is concluded that [ Frd ] is probably controlled by an extrachromosomal entity, not located in the cytoplasm. The possible relationships between Frd and [ Frd ] are discussed.