The Rice Pentatricopeptide Repeat Protein RF5 Restores Fertility in Hong-Lian Cytoplasmic Male-Sterile Lines via a Complex with the Glycine-Rich Protein GRP162  

Abstract

The cytoplasmic male sterility (CMS) phenotype in plants can be reversed by the action of nuclear-encoded fertility restorer (Rf) genes. The molecular mechanism involved in Rf gene-mediated processing of CMS-associated transcripts is unclear, as are the identities of other proteins that may be involved in the CMS-Rf interaction. In this study, we cloned the restorer gene Rf5 for Hong-Lian CMS in rice and studied its fertility restoration mechanism with respect to the processing of the CMS-associated transcript atp6-orfH79. RF5, a pentatricopeptide repeat (PPR) protein, was unable to bind to this CMSassociated transcript; however, a partner protein of RF5 (GRP162, a Gly-rich protein encoding 162 amino acids) was identified to bind to atp6-orfH79. GRP162 was found to physically interact with RF5 and to bind to atp6-orfH79 via an RNA recognition motif. Furthermore, we found that RF5 and GRP162 are both components of a restoration of fertility complex (RFC) that is 400 to 500 kD in size and can cleave CMS-associated transcripts in vitro. Evidence that a PPR protein interacts directly with a Gly-rich protein to form a subunit of the RFC provides a new perspective on the molecular mechanisms underlying fertility restoration.

An RNA recognition motif-containing protein is required for plastid RNA editing in Arabidopsis and maize

“…GRP162 interacts in vivo with RF5, a PPR protein encoded by the fertility restorer gene Rf5 to the Hong-Lian cytoplasmic male sterility. GRP162 was also shown to bind in vitro and in vivo to atp6-orfH79, the cytoplasmic male sterility-associated transcript (47), which is cleaved in the fertility-restored line. Like GRP162, ORRM1 interacts with a PPR protein and can bind to a transcript targeted for editing.…”

“…However, little is known about the molecular function of either GR-RBPs or mRBPs. Recently a rice GR-RBP protein named GRP162, which is likely orthologous to either Arabidopsis GR-RBP7 or GR-RBP8, was shown to be part of a restoration of fertility complex (47). GRP162 interacts in vivo with RF5, a PPR protein encoded by the fertility restorer gene Rf5 to the Hong-Lian cytoplasmic male sterility.…”

The Rf and Rf-like PPR in higher plants, a fast-evolving subclass of PPR genes

“…Although the HL-CMS is caused by a mitochondrial gene that is almost identical to orf79 45 and that fertility restoration in HL-CMS also involves an endoribonucleolytic cleavage upstream of orf79H, RF5 (RF1A) does not have the ability to bind to the atp6-orfH79 transcript intergenic region in vitro. 17 It has been proposed that nucleotide variations between atp6-orf79 and atp6-orfH79 transcripts may be responsible for the inability of RF5 to bind upstream of orfH79. The search for RF5 protein partners identified the glycine rich 162 protein (GRP162) and further analysis indicated that this protein plays essential role in the processing of the atp6-orfH79 bi-cistronic transcript.…”

“…17,44 The identified gene encodes a PPR protein identical to RF1A, but surprisingly the mechanism associated with HL CMS restoration differs from the BT-type CMS. Although the HL-CMS is caused by a mitochondrial gene that is almost identical to orf79 45 and that fertility restoration in HL-CMS also involves an endoribonucleolytic cleavage upstream of orf79H, RF5 (RF1A) does not have the ability to bind to the atp6-orfH79 transcript intergenic region in vitro.…”

“…In the last years, several Rf genes were cloned from various crop species and the majority of them encode proteins belonging to the pentatricopeptide repeat family (PPR) (summarized in Table 1). 6,[12][13][14][15][16][17] All these restoring PPR proteins (hereafter called Rf-PPR) were shown to target the mitochondrion and act by specifically reducing the accumulation of their cognate CMS-associated mitochondrial RNAs and/or proteins. The prevalence of PPR proteins in plastid and mitochondrial RNA processing is now largely documented 18 and the fact that many Rf genes were found to encode PPR proteins is coherent CMS/Rf systems to serve as precious models to study nucleomitochondrial genetic interactions in plants.…”