Research Article

A putative miR172-targeted CeAPETALA2-like gene is involved in floral patterning regulation of the orchid Cymbidium ensifolium

Published: October 05, 2015
Genet. Mol. Res. 14 (4) : 12049-12061 DOI: 10.4238/2015.October.5.18

Abstract

APETALA2 plays critical roles in establishing meristem and organ identity during plant floral development. In this study, we obtained a CeAP2-like gene by using the mRNA differential display technique to analyze the wild type and a multitepal mutant of the orchid Cymbidium ensifolium. The full-length cDNA encoding the CeAP2-like transcription factor shows significant similarity to the cDNA of AP2 from Erycina pusilla and contains nucleotides complementary to miR172. Using a transient gene expression system of Arabidopsis protoplasts, we found that the accumulation of CeAP2-like protein and transcripts was negatively regulated by miR172, indicating this gene as a putative target of miR172. Northern blotting revealed that CeAP2-like is dominantly expressed in the sepals and petals of the wild-type flower, and shows low expression in the gynostemium. In contrast, the accumulation of CeAP2-like transcripts decreased significantly, especially in the central part of the mutant flower, corresponding to its abnormal petals and the absence of the gynostemium. Furthermore, we found an antagonistic expression pattern between CeAP2-like and AGAMOUS in the wild type, representing A- and C-class genes that specify floral organ fate. However, this antagonistic distribution was modified in the multitepal mutant, and both genes showed lower expression than that in the wild type. This result suggested that the balance between CeAP2-like and AGAMOUS activity was important for the regulation of floral patterning in C. ensifolium. This study represents the first report on a class A gene and its regulatory role for floral development in the orchid C. ensifolium.

APETALA2 plays critical roles in establishing meristem and organ identity during plant floral development. In this study, we obtained a CeAP2-like gene by using the mRNA differential display technique to analyze the wild type and a multitepal mutant of the orchid Cymbidium ensifolium. The full-length cDNA encoding the CeAP2-like transcription factor shows significant similarity to the cDNA of AP2 from Erycina pusilla and contains nucleotides complementary to miR172. Using a transient gene expression system of Arabidopsis protoplasts, we found that the accumulation of CeAP2-like protein and transcripts was negatively regulated by miR172, indicating this gene as a putative target of miR172. Northern blotting revealed that CeAP2-like is dominantly expressed in the sepals and petals of the wild-type flower, and shows low expression in the gynostemium. In contrast, the accumulation of CeAP2-like transcripts decreased significantly, especially in the central part of the mutant flower, corresponding to its abnormal petals and the absence of the gynostemium. Furthermore, we found an antagonistic expression pattern between CeAP2-like and AGAMOUS in the wild type, representing A- and C-class genes that specify floral organ fate. However, this antagonistic distribution was modified in the multitepal mutant, and both genes showed lower expression than that in the wild type. This result suggested that the balance between CeAP2-like and AGAMOUS activity was important for the regulation of floral patterning in C. ensifolium. This study represents the first report on a class A gene and its regulatory role for floral development in the orchid C. ensifolium.