AGO10 and Floral Stem Cell Termination Figure 1. hen6 (ago10-12) and hen7 (ag-10) mutations enhance hua1 hua2. (A) A hua1 hua2 flower with a gynoecium that was slightly enlarged at the top. (B) A hua1 hua2 hen6 (or hua1 hua2 ago10-12) flower with petaloid stamens and a bulged gynoecium containing ectopic floral organs inside. (C) A hua1 hua2 hen7 (or hua1 hua2 ag-10) flower that resembled that of an ag mutant in that the third whorl stamens were transformed into petals and that the sepal-petal-petal pattern was reiterated. (D) Siliques of hua1 hua2. (E) Siliques of hua1 hua2 ago10-12. (F) A representative silique of hua1 ag-10 (left), hua1 ag-10 plants carrying the pAG::AG-GFP transgene (middle), and Ler (right). (G, H) Longitudinal sections of hua1 hua2 and hua1 hua2 ago10-12 stages 10-11 flowers. An ectopic floral meristem inside the fourth whorl carpels was present in hua1 hua2 ago10-12 (H) but not hua1 hua2 (G). ca, carpel; st, stamen. (I) A hua1 hua2 ago10-12 flower carrying AGO10 genomic DNA, which rescued the floral determinacy and homeotic defects of hua1 hua2 ago10-12. Some outer whorl organs were removed to expose the gynoecia in (A), (B), and (I). Scale bars, 50 mm in (G), (H) and 1 mm in all other panels. doi:10.1371/journal.pgen.1001358.g001 [31]. Accordingly, a genetic screen in the hua1 hua2 background effectively identified genes that promote reproductive organ identity specification or floral determinacy [28,32–35]. Among the new mutations we isolated in this screen were hua enhancer6 (hen6) and hua enhancer7 (hen7), recessive alleles in two genes. hen6 enhanced the floral phenotypes of hua1 hua2 in terms of both reproductive organ identities and floral determinacy. While hua1 hua2 flowers have stamens in the third whorl, petaloid stemens were observed in hua1 hua2 hen6 flowers (Figure 1A,1B; Figure S3H), indicating a defect in stamen identity specification. In addition, hua1 hua2 hen6 gynoecia were much shorter than those of hua1 hua2 and appeared bulged or heart-shaped (Figure 1B,1D,1E). Extra floral organs internal to the fourth whorl carpels were present in hua1 hua2 hen6 but not in hua1 hua2 gynoecia; these extra organs were generated from an indeterminate floral meristem as observed in longitudinal sections of flowers (Figure 1G,1H; Table S1). hen6 was mapped to the BAC MQD19 on chromosome 5, from which AGO10 was selected as a candidate gene for sequencing. A C-to-T transition leading to the substitution of leucine (L) 674, which is highly conserved among all Arabidopsis AGO proteins, by phenylalanine (F) was found in AGO10 (Figure S1A,S1B). The homeotic and floral determinacy defects of hua1 PLoS Genetics | www.plosgenetics.org 3 March 2011 | Volume 7 | Issue 3 | e1001358