The Arabidopsis NF-YA3 and NF-YA8 Genes Are Functionally Redundant and Are Required in Early Embryogenesis

Monica Fornari,Valentina Calvenzani,Katia Petroni,Chiara Tonelli,Simona Masiero,Markus Grebe

doi:10.1371/journal.pone.0082043

Abstract

Nuclear factor Y (NF-Y) is a trimeric transcription factor composed of three distinct subunits called NF-YA, NF-YB and NF-YC. In Arabidopsis thaliana, NF-Y subunits are known to play roles in many processes, such as gametogenesis, embryogenesis, seed development, drought resistance, ABA signaling, flowering time, primary root elongation, Endoplasmic Reticulum (ER) stress response and blue light responses. Here, we report that the closely related NF-YA3 and NF-YA8 genes control early embryogenesis. Detailed GUS and in situ analyses showed that NF-YA3 and NF-YA8 are expressed in vegetative and reproductive tissues with the highest expression being during embryo development from the globular to the torpedo embryo stage. Plants from the nf-ya3 and nf-ya8 single mutants do not display any obvious phenotypic alteration, whereas nf-ya3 nf-ya8 double mutants are embryo lethal. Morphological analyses showed that the nf-ya3 nf-ya8 embryos fail to undergo to the heart stage and develop into abnormal globular embryos with both proembryo and suspensor characterized by a disordered cell cluster with an irregular shape, suggesting defects in embryo development. The suppression of both NF-YA3 and NF-YA8 gene expression by RNAi experiments resulted in defective embryos that phenocopied the nf-ya3 nf-ya8 double mutants, whereas complementation experiments partially rescued the abnormal globular nf-ya3 nf-ya8 embryos, confirming that NF-YA3 and NF-YA8 are required in early embryogenesis. Finally, the lack of GFP expression of the auxin responsive DR5rev::GFP marker line in double mutant embryos suggested that mutations in both NF-YA3 and NF-YA8 affect auxin response in early developing embryos. Our findings indicate that NF-YA3 and NF-YA8 are functionally redundant genes required in early embryogenesis of Arabidopsis thaliana.

Highlights

In plants and animals, developmental processes are controlled by complex networks of transcription factors, often arranged in multiprotein DNA-binding complexes, whose gene regulatory activity derives from intrinsic properties and the properties of their trans-acting partners [1,2,3,4].The NUCLEAR FACTOR Y (NF-Y), called the CCAAT binding factor (CBF) and the heme-activated protein in yeast (HAP), is a heterotrimeric transcription factor that binds with high affinity and sequence specificity the highly conserved core sequence CCAAT
We obtained transgenic plants carrying a 1995 bp genomic region upstream of NF-YA3 start codon fused to the GUS reporter gene and plants carrying a 1000 bp genomic region upstream of NF-YA8 start codon fused to the GUS reporter gene
Detailed analyses of GUS expression in reproductive organs revealed similar expression of NF-YA3 and NF-YA8 in anthers and in pollen grains at floral stage 8 according to Bowman, 1994 (Fig. 1 G,g), whereas in subsequent stages the signal was in filaments of anthers, but not in mature pollen grains

Summary

Introduction

Developmental processes are controlled by complex networks of transcription factors, often arranged in multiprotein DNA-binding complexes, whose gene regulatory activity derives from intrinsic properties and the properties of their trans-acting partners [1,2,3,4].The NUCLEAR FACTOR Y (NF-Y), called the CCAAT binding factor (CBF) and the heme-activated protein in yeast (HAP), is a heterotrimeric transcription factor that binds with high affinity and sequence specificity the highly conserved core sequence CCAAT. NF-YB and NF-YC subunits form a tight dimer through protein structures similar to the Histon Fold Motif (HFM), a conserved protein-protein and DNA-binding interaction module [7,8]. This dimer translocates to the nucleus, where it offers a complex surface for the association of the NF-YA subunit [9,10,11,12]. A reclassification of the plant NF-Y genes, based on Arabidopsis NF-Y nomenclature, aimed to avoid confusion with acronyms and related, but functionally distinct, histone fold domain (HFD) protein was proposed [16]. The conserved heterodimerization capacity of AtNF-Y histone-like subunits and the different affinities of At-NFYAs for the CAAT sequence was determined [17]

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