A single nucleotide substitution in the coding region of Ogura male sterile gene, orf138, determines effectiveness of a fertility restorer gene, Rfo, in radish

<jats:title>Abstract</jats:title><jats:p>Cytoplasmic male sterility (CMS) observed in many plants leads defect in the production of functional pollen, while the expression of CMS is suppressed by a fertility restorer gene in the nuclear genome. Ogura CMS of radish is induced by a mitochondrial <jats:italic>orf138</jats:italic>, and a fertility restorer gene, <jats:italic>Rfo</jats:italic>, encodes a P-type PPR protein, ORF687, acting at the translational level. But, the exact function of ORF687 is still unclear. We found a Japanese variety showing male sterility even in the presence of <jats:italic>Rfo</jats:italic>. We examined the pollen fertility, <jats:italic>Rfo</jats:italic> expression, and <jats:italic>orf138</jats:italic> mRNA in progenies of this variety. The progeny with Type H <jats:italic>orf138</jats:italic> and <jats:italic>Rfo</jats:italic> showed male sterility when their <jats:italic>orf138</jats:italic> mRNA was unprocessed within the coding region. By contrast, all progeny with Type A <jats:italic>orf138</jats:italic> were fertile though <jats:italic>orf138</jats:italic> mRNA remained unprocessed in the coding region, demonstrating that ORF687 functions on Type A but not on Type H. In silico analysis suggested a specific binding site of ORF687 in the coding region, not the 5′ untranslated region estimated previously, of Type A. A single nucleotide substitution in the putative binding site diminishes affinity of ORF687 in Type H and is most likely the cause of the ineffectiveness of ORF687. Furthermore, fertility restoration by RNA processing at a novel site in some progeny plants indicated a new and the third fertility restorer gene, <jats:italic>Rfs</jats:italic>, for <jats:italic>orf138</jats:italic>. This study clarified that direct ORF687 binding to the coding region of <jats:italic>orf138</jats:italic> is essential for fertility restoration by <jats:italic>Rfo</jats:italic>.</jats:p>