Abstract
Though cytoplasmic male sterility (CMS) in peppers is associated with the orf507 gene, definitive and direct evidence that it directly causes male sterility is still lacking. In this study, differences in histochemical localization of anther cytochrome c oxidase between the pepper CMS line and maintainer line were observed mainly in the tapetal cells and tapetal membrane. Inducible and specific expression of the orf507 gene in the pepper maintainer line found that transformants were morphologically similar to untransformed and transformed control plants, but had shrunken anthers that showed little dehiscence and fewer pollen grains with lower germination rate and higher naturally damaged rate. These characters were different from those of CMS line which does not produce any pollen grains. Meanwhile a pollination test using transformants as the male parent set few fruit and there were few seeds in the limited number of fruits. At the tetrad stage, ablation of the tapetal cell induced by premature programmed cell death (PCD) occurred in the transformants and the microspores were distorted and degraded at the mononuclear stage. Stable transmission of induced semi-male sterility was confirmed by a test cross. In addition, expression of orf507 in the maintainer lines seemed to inhibit expression of atp6-2 to a certain extent, and lead to the increase of the activity of cytochrome c oxidase and the ATP hydrolysis of the mitochondrial F1Fo-ATP synthase. These results introduce the premature PCD caused by orf507 gene in tapetal cells and semi-male sterility, but not complete male sterility.
Highlights
Cytoplasmic male sterility (CMS) is a maternally inherited trait that prevents a plant from producing functional pollen grains
We found that mitochondrial cytochrome c oxidase and F1Fo-ATPase were dysfunctional in the cytoplasmic male sterility (CMS) line HW203A (Ji et al, 2013), but it remains unclear whether expression of orf507 is responsible for the dysfunction of the mitochondrial enzyme
The results showed that osmium black was localized on the membrane of the anther wall at the MMC stage in the CMS line, while hardly any osmium black particles were found in the microspore mother cell and the tapetum cell (Figure 1)
Summary
Cytoplasmic male sterility (CMS) is a maternally inherited trait that prevents a plant from producing functional pollen grains. The CMS lines, that can-not produce functional pollen grains, are often used as female lines in hybrid seed production, to increase the purity of seeds (Schnable and Wise, 1998). The several bottlenecks faced during the development of CMS lines through orf507 and pepper male sterility conventional breeding require biotechnological intervention. The plant mitochondrial genome cannot be manipulated directly, so investigations of mitochondrial contributions to male sterility must be made by engineering nuclear genes encoding mitochondria-targeted proteins, including the expression of unedited forms of mitochondrial genes in the maintainer line (Hernould and Suharsono, 1993), and the expression of the CMSassociated mitochondrial orf s (Yamamoto et al, 2008). CMS was related with premature degradation of the tapetal cell, a sporogenous tissue that nurtures the pollen mother cells (Hernould et al, 1998). Premature DNA fragmentation and programmed cell death (PCD) were observed at meiosis stage in the anthers of sterile plants (Nizampatnam et al, 2009)
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