Expression of sunflower cytoplasmic male sterility-associated open reading frame, orfH522 induces male sterility in transgenic tobacco plants

Abstract

Sterility in the universally exploited PET1-CMS system of sunflower is associated with the expression of orfH522, a novel mitochondrial gene. Definitive evidence that ORFH522 is directly responsible for male sterility is lacking. To test the hypothesis that ORFH522 is sufficient to induce male sterility, a set of chimeric constructs were developed. The cDNA of orfH522 was cloned in-frame with yeast coxIV pre-sequence, and was expressed under tapetum-specific promoter TA29 (construct designated as TCON). For developing control vectors, orfH522 was cloned without the transit peptide under TA29 promoter (TON) or orfH522 was cloned with or without transit peptide under the constitutive CaMV35S promoter (SCOP and SOP). Among several independent transformants obtained with each of the gene cassettes, one third of the transgenics (6/17) with TCON were completely male sterile while more than 10 independent transformants obtained with each of the control vectors were fertile. The male sterile plants were morphologically similar to fertile plants, but had anthers that remained below the stigmatic surface at anthesis. RT-PCR analysis of the sterile plants confirmed the anther-specific expression of orfH522 and bright-field microscopy demonstrated ablation of the tapetal cell layer. Premature DNA fragmentation and programmed cell death was observed at meiosis stage in the anthers of sterile plants. Stable transmission of induced male sterility trait was confirmed in test cross progeny. This constitutes the first report at demonstrating the induction of male sterility by introducing orfH522 gene that could be useful for genetic engineering of male sterility.

Cost of Having the Largest Mitochondrial Genome: Evolutionary Mechanism of Plant Mitochondrial Genome

“…A transgenic approach sometimes (but not always) works well, wherein plants expressing specific unique ORFs fused with mitochondrial import signal peptides express male sterility or toxin sensitivity [79,[101][102][103][104][105]. Microorganisms such as E coli can also be used as hosts to examine the toxin sensitivities conferred by specific ORFs [36,76].…”

Early flowering and genetic containment studies in transgenic poplar

“…Various sterility gene constructs have been tested with different levels of success in crop plants, e.g., by expression of deleterious genes in flower organs, like barnase (Paddon & Hartley 1986, Mariani et al 1990, García-Sogo et al 2010, orfH522 (Nizampatnam et al 2009), monooxygenase (MNX - Gan et al 2010), stilbene synthase (STS -Fisher et al 1997, Höfig et al 2006, the gene for ribosome inactivating protein (Palmiter et al 1987) and RNA interference (Nawaz-ul-Rehman et al 2007). Specific floral regulatory promoters to direct expression of genes in reproductive structures have been found in crop plants, e.g., TA29 promoter from Nicotiana tabacum (Koltunow et al 1990, Mariani et al 1990) and forest tree species, e.g., PrMA-LE1 from Pinus radiata (Höfig et al 2003) and PTD from poplar (Skinner et al 2003).…”

Characterization of a male sterile mutant from progeny of a transgenic plant containing a leaf senescence-inhibition gene in wheat

“…Male sterility induced by genetic transformation or alien DNA introduction has been reported in many studies (Ji et al 2003;Cheon et al 2004;Yang et al 2004;Wang et al 2005;Höfig et al 2006;Ge et al 2008;Wan et al 2008;Nizampatnam et al 2009). However, most of the male sterile lines were not genetically characterized and it is not known if the male sterility of the lines was stably inherited or if the lines had potential value for crop improvement.…”

“…Constitutive expression of a human gene affected the morphologies of transgenic plants such that vegetative growth of transgenic tobacco was retarded, and male sterility was induced in transgenic tobacco and Arabidopsis (Cheon et al 2004). Tapetum specific expression of mitochondrial gene orfH522 in tobacco made onethird of the transgenics completely male sterile (Nizampatnam et al 2009). Cone-specific expression of a stilbene synthase gene (STS) in anthers of transgenic tobacco plants resulted in complete nuclear male sterility in 70% of transformed plants (Höfig et al 2006).…”