Cytoplasmic Male Sterility and Fertility Restoration

Cytoplasmic Male Sterility and Fertility Restoration

Cytoplasmic male sterility (CMS), a condition under which a plant is unable to produce functional pollen, is widespread among higher plants. CMS systems represent a valuable tool in the production of hybrid seed in self-pollinating crop species, including maize, rice, cotton, and a number of vegetable crops. Hybrids often exhibit heterosis, more commonly known as hybrid vigor, whereby hybrid progeny exhibit superior growth characteristics relative to either of the parental lines. CMS systems can be of considerable value in facilitating efficient hybrid seed production.

There is growing interest in improving hybrid technology both to help supply food for the world’s increasing population and to contribute to land conservation efforts. For example, the use of hybrid rice enabled China to reduce the total amount of land planted to rice from 36.5 Mha in 1975 to 30.5 Mha in 2000 while at the same time increasing total production from 128 to 189 million tons, representing a yield increase of 3.5 to 6.2 tons/ha ( Understanding the molecular basis of CMS, as well as other hybrid production methods involving self-incompatibility and apomixis, is critical for continued improvements in hybrid technology.

CMS is a maternally inherited trait that is often associated with unusual open reading frames (ORFs) found in mitochondrial genomes (Chase and Babay-Laughnan, 2004;Hanson and Bentolila, 2004). In many cases, it has been found that male fertility can be restored by nuclear-encoded fertility restorer (Rf) gene(s). CMS/Rf systems therefore are also of value in the study of interactions between nuclear and mitochondrial genomes. On the one hand, sterility results from mitochondrial genes causing cytoplasmic dysfunction, and on the other, fertility restoration relies on nuclear genes that suppress cytoplasmic dysfunction.

CMS can arise spontaneously in breeding lines, as a result of wide crosses or the interspecific exchange of nuclear and cytoplasmic genomes, or following mutagenesis (Hanson and Bentolila, 2004). For example, CMS-WA (wild abortive) rice was developed inindica rice cultivars from a male-sterile plant found in a natural population of the wild riceOryza rufipogon Griff. CMS-Boro II rice arose from a wide cross based on the cytoplasm of Chinsurah Boro II (O. sativa subsp indica) and the nucleus of Taichung 65 (subsp japonica). The well-known male-sterile Texas cytoplasm in maize arose spontaneously in a breeding line, and CMS-PET1 cytoplasm of sunflower arose from an interspecific cross betweenHelianthus petiolaris and H. annuus.

There are a number of different types of CMS systems with distinct genetic features, both within and among different species, but key features that appear to be shared across different types are (1) CMS is associated with chimeric mitochondrial ORFs, and (2) fertility restoration is often associated with genes encoding pentatricopeptide repeat (PPR) proteins (Chase and Babay-Laughnan, 2004Hanson and Bentolila, 2004). In this issue of The Plant CellWang et al. (pages 676–687) describe details of the molecular basis of CMS and fertility restoration in the CMS-Boro II system in rice, which are likely to have far-reaching implications for CMS systems in general. First, the authors show that the mitochondrialorf79 associated with CMS-Boro II encodes a cytotoxic peptide responsible for CMS, and second, they show that two PPR proteins encoded by the Rf-1 locus in the Boro II system block the production of this cytotoxic peptide by distinct mechanisms (endonucleolytic cleavage and degradation of the dicistronic mRNA).

In the early 1990s, several groups reported that rice CMS-Boro II is associated with an abnormal copy of the mitochondrial gene apt6 (Kadowaki et al., 1990Iwabuchi et al., 1993) that produces aberrant mRNA transcripts containing an additional ORF named orf79(Akagi et al., 1994). Interestingly, in a number of well-characterized systems, CMS is associated with alterations in promoter regions and portions of coding regions of mitochondrial ATP synthase subunit genes, which raises the possibility that impaired ATP synthase activity could be a causal factor in the disrupted pollen development in CMS lines in a number of species (Hanson and Bentolila, 2004). It is more often considered that transcription of unusual or aberrant ORFs is causally related to CMS, and it has been shown that CMS-associated mitochondrial ORFs encode proteins with cytotoxic properties in sunflower (Nakai et al., 1995) and Brassicaceae species (Duroc et al., 2005). However, it was unknown if CMS-Boro II resulted from incorrect translation of atp6 or from translation of downstream sequences including orf79Akagi et al. (1994) found that orf79 encodes a predicted transmembrane protein with a novel C-terminal region and an N terminus showing similarity to the rice mitochondrial cytochrome oxidase subunit I. It was suspected that orf79 causes CMS in Boro II cytoplasm, but definitive proof has been lacking.

Wang et al. examined the role of orf79 in CMS, first by testing for possible cytotoxicity of the ORF79 peptide in Escherichia coli. Expression of the protein was found to be lethal to the host E. coli cells, with cell lysis leading to a rapid decrease in cell density and lethality depending on the presence of a five–amino acid segment of the C-terminal region. The authors next tested whether ORF79 causes male sterility by transforming a normal fertile rice line with orf79 carrying a mitochondrion-transit signal under the control of the cauliflower mosaic virus 35S promoter. The transgenic plants exhibited semi-male-sterility wherein 50% or more of the pollen grains were aborted (see figure). Semi-male-sterility was observed because the transgene is present in only a portion of pollen grains after meiosis. Female fertility was unaffected in the transgenic lines, and the semi-male-sterility phenotype of the T1 progeny cosegregated with the presence of the transgene with a 1:1 segregation ratio, indicating that the orf79 transgene was transmitted normally through the female germline but poorly or not all through pollen. These results show that orf79encodes a cytotoxic peptide that causes CMS in rice.

View larger version:


Pollen Grains of a Normal Fertile Rice Line and orf79-Transgenic Rice Plants.

Top, normal fertile rice line; middle, orf79-transgenic rice plants. Fertile pollen grains are darkly stained, and sterile grains are lightly stained. Bars = 50 μm. Bottom panel shows cosegregation of the orf79transgene and male sterility (s) in transgenic plants. f, male-fertile.

In addition, using immunoblot analysis, Wang et al. show that, despite the constitutive RNA expression of the gene, ORF79 protein accumulates specifically in the microspores of a CMS line but is absent in sporophytic tissues (as well as the microspores of fertility-restored plants). They propose that there may be a posttranslational regulatory mechanism that suppresses the accumulation of the protein, which could explain the genetic feature of gametophytic male sterility and why orf79 does not disrupt the development of sporophytic tissues.

Wang et al. then sought to characterize the nature of the complex Rf1 chromosomal region and clarify the molecular mechanism underlying fertility restoration. The restoring alleleRf-1 is present in some indica rice lines, whereas most lines of the subspecies japonicacarry a nonrestoring rf-1 allele. Previous research had shown that Rf-1 encodes a PPR protein that functions in fertility restoration of CMS-Boro II (Kazama and Toriyama, 2003;Akagi et al., 2004Komori et al., 2004). Akagi et al. (2004) showed that Rf-1 is a complex locus containing multiple copies of genes encoding PPR proteins. One of the genes, Rf-1A, encoded a predicted PPR protein of 791 amino acids that contained a mitochondria-targeting signal and cosegregated with fertility restoration. This was the same gene cloned by Kazama and Toriyama (2003) (called PPR8-1) and Komori et al. (2004) (called PPR791) and thus has been considered to be the single gene responsible for fertility restoration.Akagi et al. (2004) also identified two other genes thought to be nonfunctional in fertility restoration: Rf-1B, which encoded a truncated protein that lacked a mitochondrial-transit signal, and Rf-1C, which encoded a protein of high similarity to Rf-1A but was outside of the crossover point of the experimental recombinants.

Wang et al. used map-based cloning to sequence a 37-kb region surrounding Rf-1. They identified two genes that encode PPR proteins, called Rf1a and Rf1b, which were found by complementation testing to function in fertility restoration in this system. Rf1acorresponds to Rf1A/PPR8-1/PPR791 identified in previous studies, whereas Rf1b is another gene at this locus that has not been described previously. Wang et al. show that the RF1A and RF1B proteins both function to restore male fertility by blocking ORF79 production via somewhat different mechanisms. RF1A is shown to mediate endonucleolytic cleavage of the dicistronic atp6/orf79 mRNA at three major regions, each with multiple cleaving sites, whereas RF1B mediates degradation of atp6/orf79 mRNA with no detectable intermediates. RF1A function was found to be epistatic over RF1B, such that when both were present, atp6/orf79 mRNA was preferentially cleaved by RF1A, and the cleavage products were not susceptible to further degradation mediated by RF1B. The authors suggest that different fertility restorer lines may carry either or both functional copies ofRf1a and Rf1b, and the restorer lines in previous studies presumably did not carry a functional Rf1b allele.

PPR proteins constitute a large family, with >400 members in Arabidopsis and rice that are thought to be RNA binding proteins involved in posttranscriptional processes (RNA processing and translation) in mitochondria and chloroplasts, but little data exist on the functions of individual proteins in this family (Lurin et al., 2004). Lurin et al. (2004)hypothesized that they function as sequence-specific adaptors for a variety of other RNA-associated proteins. This idea was supported by Schmitz-Linneweber et al. (2005), who showed that the maize PPR protein CRP1 influences expression of chloroplast genes through association with specific mRNAs, and Kotera et al. (2005), who showed that PPR proteins are involved in mRNA editing in chloroplasts. Bentolila et al. (2002) suggested an mRNA processing function for Rf PPR function after identifying the first Rf gene in petunia, which was found to encode the PPR protein Rf-PPR592. They observed that the presence of the restorer Rf-PPR592 led to a decrease in the gene product of the aberrant mitochondrial ORF pcf and concluded that Rf-PPR592 is likely involved in mediating a reduction in mRNA accumulation. Wang et al. provide definitive support for this hypothesis, showing that RfPPR proteins participate both in endonucleolytic cleavage (RF1A) and degradation (RF1B) of specific mRNAs. They further show that RF1A functions to promote the editing of normalatp6 mRNA independently of its cleavage of dicistronic atp6/orf79 mRNA and role in fertility restoration and suggest that this may be its primary function.

The results of Wang et al. have important implications for other CMS systems. For example, the A3 CMS system in sorghum has similar features with the CMS-Boro II system in rice: the mitochondrial CMS gene of sorghum, orf107, is similar to rice orf79 (Tang et al., 1996), and fertility restoration in sorghum is associated with the editing status of mitochondrial atp6 (Pring et al., 1999). In addition, previous work has suggested that Rfloci associated with fertility restoration in the genetically distinct rice CMS systems CMS-WA (Zhang et al., 2002) and CMS-HL (Liu et al., 2004) map to the same PPR gene cluster as the CMS-Boro II Rf-1 locus. Wang et al. hypothesize that a number of different genes within this PPR cluster have been recruited as fertility restorers with divergent molecular functions. Analyses of these clusters in other CMS systems are needed for a complete understanding of the evolution and molecular basis of CMS.

  1. Nancy A. Eckardt, News and Reviews Editor
  3. doi: http:/​/​dx.​doi.​org/​10.​1105/​tpc.​106.​041830The Plant Cell March 2006 vol. 18 no. 3 515-517
  1. PPT Slides of All Figures


Akagi, H., Nakamura, A., Yokozeki-Misono, Y., Inagaki, A., Takahashi, H., Mori, K., and Fujimura, T. (2004). Positional cloning of the rice Rf-1 gene, a restorer of BT-type cytoplasmic male sterility that encodes a mitochondria-targeting PPR protein. Theor. Appl. Genet. 108, 1449–1457.


CrossRefMedlineWeb of Science

Akagi, H., Sakamoto, M., Shinjyo, C., Shimada, H., and Fujimura, T. (1994). A unique sequence located downstream from the rice mitochondrial apt6 may cause male sterility. Curr. Genet. 25, 52–58.


CrossRefMedlineWeb of Science

Bentolila, S., Alfonso, A.A., and Hanson, M.R. (2002). A pentatricopeptide repeat-containing gene restores fertility to cytoplasmic male-sterile plants. Proc. Natl. Acad. Sci. USA 99, 10887–10892.


Abstract/FREE Full Text

Chase, C., and Babay-Laughnan, S. (2004). Cytoplasmic male sterility and fertility restoration by nuclear genes. In Molecular Biology and Biotechnology of Plant Organelles, H. Daniell and C. Chase, eds (Dordrecht, The Netherlands: Kluwer Academic Publishers), pp. 593–622.

Duroc, Y., Gaillard, C., Hiard, S., Defrance, M.-C., Pelletier, G., and Budar, F.(2005). Biochemical and functional characterization of ORF138, a mitochondrial protein responsible for Ogura cytoplasmic male sterility in Brassiceae. Biochimie87, 1089–1100.



Hanson, M.R., and Bentolila, S. (2004). Interactions of mitochondrial and nuclear genes that affect male gametophyte development. Plant Cell 16, S154–S169.


FREE Full Text

Iwabuchi, M., Kyozuka, J., and Shimamoto, K. (1993). Processing followed by complete editing of an altered mitochondrial apt6 RNA restores fertility of cytoplasmic male sterile rice. EMBO J. 12, 1437–1446.


MedlineWeb of Science

Kadowaki, K., Suzaki, T., and Kazama, S. (1990). A chimeric gene containing the 5′ portion of atp6 is associated with cytoplasmic male-sterility of rice. Mol. Gen. Genet. 224, 10–16.


MedlineWeb of Science

Kazama, T., and Toriyama, K. (2003). A pentatricopeptide repeat-containing gene that promotes the processing of aberrant apt6 RNA of cytoplasmic male-sterile rice. FEBS Lett. 544, 99–102.


CrossRefMedlineWeb of Science

Komori, T., Ohta, S., Murai, N., Takakura, Y., Kuraya, Y., Suzuki, S., Hiei, Y., Imaseki, H., and Nitta, N. (2004). Map-based cloning of a fertility restorer gene,Rf-1, in rice (Oryza sativa L.). Plant J. 37, 315–325.


CrossRefMedlineWeb of Science

Kotera, E., Tasaka, M., and Shikanai, T. (2005). A pentatricopeptide repeat protein is essential for RNA editing in chloroplasts. Nature 433, 326–330.

CrossRefMedlineWeb of Science

Liu, X.Q., Xu, X., Tan, Y.P., Li, S.Q., Hu, J., Huang, J.Y., Yang, D.C., Li, Y.S., and Zhu, Y.G. (2004). Inheritance and molecular mapping of two fertility-restoring loci for Honglian gametophytic cytoplasmic male sterility in rice (Oryza sativa L.). Mol. Genet. Genomics 271, 586–594.



Lurin, C., et al. (2004). Genome-wide analysis of Arabidopsis pentatricopeptide repeat proteins reveals their essential role in organelle biogenesis. Plant Cell 16, 2089–2103.


Abstract/FREE Full Text

Nakai, S., Noda, D., Kondo, M., and Terachi, T. (1995). High-level expression of a mitochondrial orf-522 gene from the male-sterile sunflower is lethal toEscherichia coli. Breed. Sci. 45, 233–236.


Web of Science

Pring, D.R., Tang, H.V., Howad, W., and Kempken, F. (1999). A unique two-gene gametophytic male sterility system in Sorghum involving a possible role of RNA editing in fertility restoration. J. Hered. 90, 386–393.


Abstract/FREE Full Text

Schmitz-Linneweber, C., Williams-Carrier, R., and Barkan, A. (2005). RNA immunoprecipitation and microarray analysis show a chloroplast pentatricopeptide repeat protein to be associated with the 5′ region of mRNAs whose translation it activates. Plant Cell 17, 2791–2804.


Abstract/FREE Full Text

Tang, H.V., Pring, D.R., Shaw, L.C., Salazar, R.A., Muza, F.R., Yan, B., and Schertz, K.F. (1996). Transcript processing internal to a mitochondrial open reading frame is correlated with fertility restoration in male-sterile sorghum.Plant J. 10, 123–133.


CrossRefMedlineWeb of Science

  1. Wang, Z., et al.(2006). Cytoplasmic male sterility of rice with Boro II cytoplasm is caused by a cytotoxic peptide and is restored by two related PPR motif genes via distinct modes of mRNA silencing. Plant Cell 18, 676–687.

Abstract/FREE Full Text

Zhang, Q., Liu, Y.G., Zhang, G., and Mei, M. (2002). Molecular mapping of the fertility restorer gene Rf4 for WA cytoplasmic male sterility in rice. Yi Chuan Xue Bao 29, 1001–1004.



We recommend

  1. RNA PROCESSING FACTOR3 is crucial for the accumulation of mature ccmC transcripts in mitochondria of Arabidopsis accession Columbia.

Christian Jonietz et al., Plant Physiol, 2011

  1. Cytoplasmic male sterility of rice with boro II cytoplasm is caused by a cytotoxic peptide and is restored by two related PPR motif genes via distinct modes of mRNA silencing.

Zhonghua Wang et al., Plant Cell, 2006

  1. Mitochondrial mRNA polymorphisms in different Arabidopsis accessions.

Joachim Forner et al., Plant Physiol, 2008

  1. The rice pentatricopeptide repeat protein RF5 restores fertility in Hong-Lian cytoplasmic male-sterile lines via a complex with the glycine-rich protein GRP162.

Jun Hu et al., Plant Cell, 2012

  1. A reevaluation of rice mitochondrial evolution based on the complete sequence of male-fertile and male-sterile mitochondrial genomes.

Stéphane Bentolila et al., Plant Physiol, 2011

  1. Disruption of a rice pentatricopeptide repeat protein causes a seedling-specific albino phenotype and its utilization to enhance seed purity in hybrid rice production.

Ning Su et al., Plant Physiol, 2012

  1. Characterization of Raphanus sativus pentatricopeptide repeat proteins encoded by the fertility restorer locus for Ogura cytoplasmic male sterility.

M Uyttewaal et al., Plant Cell, 2008

  1. Two DEAD-box proteins may be part of RNA-dependent high-molecular-mass protein complexes in Arabidopsis mitochondria.

Annemarie Matthes et al., Plant Physiol, 2007

  1. Plastid Genotyping Reveals the Uniformity of Cytoplasmic Male Sterile-T Maize Cytoplasms.

Massimo Bosacchi et al., Plant Physiol, 2015

  1. Suppression of cytoplasmic male sterility by nuclear genes alters expression of a novel mitochondrial gene region.

M Singh et al., Plant Cell, 1991

Powered by TrendMD

Articles citing this article

  • Comparative studies of mitochondrial proteomics reveal an intimate protein network of male sterility in wheat (Triticum aestivum L.)J Exp Bot 2015 66: 6191-6203.
  • Experimental Analysis of the Rice Mitochondrial Proteome, Its Biogenesis, and HeterogeneityPlant Physiol. 2009 149: 719-734.

View this article with LENS

Navigate This Article

  1. Top
  2. References

Cited By

View citing article information

  1. More articles like this
  2. More citing articles

Article Usage Stats




Leave a reply

Mulena Naka Group: Mynaka | Nyuwe | Shop4me | Mulena Infotech 
©2017  - Powered by: Mulena Infotech - All Rights  Reserved

Log in with your credentials


Forgot your details?

Create Account