Novel strategy for yeast construction using δ-integration and cell fusion to efficiently produce ethanol from raw starch

We developed a novel strategy for constructing yeast to improve levels of amylase gene expression and the practical potential of yeast by combining δ-integration and polyploidization through cell fusion. Streptococcus bovis α-amylase and Rhizopus oryzae glucoamylase/α-agglutinin fusion protein genes were integrated into haploid yeast strains. Diploid strains were constructed from these haploid strains by mating, and then a tetraploid strain was constructed by cell fusion. The α-amylase and glucoamylase activities of the tetraploid strain were increased up to 1.5- and tenfold, respectively, compared with the parental strain. The diploid and tetraploid strains proliferated faster, yielded more cells, and fermented glucose more effectively than the haploid strain. Ethanol productivity from raw starch was improved with increased ploidy; the tetraploid strain consumed 150 g/l of raw starch and produced 70 g/l of ethanol after 72 h of fermentation. Our strategy for constructing yeasts resulted in the simultaneous overexpression of genes integrated into the genome and improvements in the practical potential of yeasts.     

Ptiotosynthetic activity and chloroplast membrane polypeptides in euploid cells of Ricinus

The possible effects of altered nuclear complement size on the poiypeptide composition and photochemical activity of chloroplasts in haploid, diploid, and tetraploid cells, of Ricinus communis L. have been evaluated. The electron transport capacity in isolated chloroplasts decreases with the increase in nuclear genome size. Both Photosystem II (DCPIP reduction) and Photosystem 1 oxygen uptake (TMPD to methyl viologen) activities were lower in plastid preparations from tetraploid individuals than in diploid and haploid cell preparations. Photosynthetic O₂-evolution and CO₂-fixation rates in leaf tissue from euploid individuals were also found to decrease with the increase in size of the nuclear genome. Specific activity levels of RuBP-carboxylase were observed to increase with ploidy. Electrophoretic examination of the poiypeptide composition of thylakoid membranes from haploid, diploid, and tetraploid celis revealed no difference in the relative proportions of the constituent polypeptides of these membranes. The regulation of chloroplast development and the basis for altered plastid function in the presence of altered nuclear genome size are discussed.     

Epigenetic inheritance and genome regulation: is DNA methylation linked to ploidy in haplodiploid insects?

Organisms show great variation in ploidy level. For example, chromosome copy number varies among cells, individuals and species. One particularly widespread example of ploidy variation is found in haplodiploid taxa, wherein males are typically haploid and females are typically diploid. Despite the prevalence of haplodiploidy, the regulatory consequences of having separate haploid and diploid genomes are poorly understood. In particular, it remains unknown whether epigenetic mechanisms contribute to regulatory compensation for genome dosage. To gain greater insights into the importance of epigenetic information to ploidy compensation, we examined DNA methylation differences among diploid queen, diploid worker, haploid male and diploid male Solenopsis invicta fire ants. Surprisingly, we found that morphologically dissimilar diploid males, queens and workers were more similar to one another in terms of DNA methylation than were morphologically similar haploid and diploid males. Moreover, methylation level was positively associated with gene expression for genes that were differentially methylated in haploid and diploid castes. These data demonstrate that intragenic DNA methylation levels differ among individuals of distinct ploidy and are positively associated with levels of gene expression. Thus, these results suggest that epigenetic information may be linked to ploidy compensation in haplodiploid insects. Overall, this study suggests that epigenetic mechanisms may be important to maintaining appropriate patterns of gene regulation in biological systems that differ in genome copy number.