Abstract
Zea mays (maize) Opaque-2 (ZmO2) protein is an important bZIP transcription factor that regulates the expression of major storage proteins (22-kD zeins) and other important genes during maize seed development. ZmO2 is subject to functional regulation through protein-protein interactions. To unveil the potential regulatory network associated with ZmO2, a protein-protein interaction study was carried out using the truncated version of ZmO2 (O2-2) as bait in a yeast two-hybrid screen with a maize seed cDNA library. A protein with homology to Taxilin was found to have stable interaction with ZmO2 in yeast and was designated as ZmTaxilin. Sequence analysis indicated that ZmTaxilin has a long coiled-coil domain containing three conserved zipper motifs. Each of the three zipper motifs is individually able to interact with ZmO2 in yeast. A GST pull-down assay demonstrated the interaction between GST-fused ZmTaxilin and ZmO2 extracted from developing maize seeds. Using onion epidermal cells as in vivo assay system, we found that ZmTaxilin could change the sub-cellular distribution of ZmO2. We also demonstrated that this change significantly repressed the transcriptional activity of ZmO2 on the 22-kD zein promoter. Our study suggests that a Taxilin-mediated change in sub-cellular distribution of ZmO2 may have important functional consequences for ZmO2 activity.
Highlights
The Zea mays protein Opaque-2 (ZmO2) is a bZIP transcription factor that is primarily expressed in the sub-aleurone layers of maize endosperm [1,2,3,4]
We have identified a ZmTaxilin protein that interacts with Zea mays (maize) protein Opaque-2 (ZmO2) supported by the following evidence
Full length ZmO2 and ZmTaxilin can interact in yeast cells and in vitro
Summary
The Zea mays (maize) protein Opaque-2 (ZmO2) is a bZIP transcription factor that is primarily expressed in the sub-aleurone layers of maize endosperm [1,2,3,4]. Previous studies revealed that a few proteins such as OHP1 (Opaque-2 heterodimerising protein 1), PBF (prolamin-box binding factor), GCN5 (general control of amino-acid synthesis protein 5) and ADA2 (transcriptional adaptor 2) can interact with ZmO2. OHP1 is a bZIP transcription factor and can bind to the ZmO2 target site as a homodimer or as a heterodimeric complex with ZmO2 [16]. PBF belongs to the Dof class of plant zinc-finger transcription factors and binds to the prolamin-box (P-box), which lies 20 bp upstream of the ZmO2 target site in the 22-kD zein gene promoter [17]. The DNA-binding activity of ZmO2 is regulated diurnally by a phosphorylation/dephosphorylation mechanism; consequentially it is likely that ZmO2 protein interacts with kinase (s) and phosphatase (s) [19,20,21]. To obtain additional information about the ZmO2 interaction network, a yeast two-hybrid screen was performed to identify proteins that interact with ZmO2
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