Arabidopsis Acyl-CoA-binding protein ACBP2 interacts with an ethylene-responsive element-binding protein, AtEBP, via its ankyrin repeats.
Cytosolic acyl-CoA-binding proteins (ACBP) bind long-chain acyl-CoAs and act as intracellular acyl-CoA transporters and maintain acyl-CoA pools. Arabidopsis thaliana ACBP2 shows conservation at the acyl-CoA-binding domain to cytosolic ACBPs but is distinct by the presence of an N-terminal transmembrane domain and C-terminal ankyrin repeats. The function of the acyl-CoA-binding domain in ACBP2 has been confirmed by site-directed mutagenesis and four conserved residues crucial for palmitoyl-CoA binding have been identified. Results from ACBP2:GFP fusions transiently expressed in onion epidermal cells have demonstrated that the transmembrane domain functions in plasma membrane targeting, suggesting that ACBP2 transfers acyl-CoA esters to this membrane. In this study, we investigated the significance of its ankyrin repeats in mediating protein-protein interactions by yeast two-hybrid analysis and in vitro protein-binding assays; we showed that ACBP2 interacts with the A. thaliana ethylene-responsive element-binding protein AtEBP via its ankyrin repeats. This interaction was lacking in yeast two-hybrid analysis upon removal of the ankyrin repeats. When the subcellular localizations of ACBP2 and AtEBP were further investigated using autofluorescent protein fusions in transient expression by agroinfiltration of tobacco leaves, the DsRed:ACBP2 fusion protein was localized to the plasma membrane while the GFP:AtEBP fusion protein was targeted to the nucleus and plasma membrane. Co-expression of DsRed:ACBP2 and GFP:AtEBP showed a common localization of both proteins at the plasma membrane, suggesting that ACBP2 likely interacts with AtEBP at the plasma membrane.
Li
Hong-Ye
HY
Department of Botany, University of Hong Kong, Pokfulam Road, Hong Kong, China.
Chye
Mee-Len
ML
D000595:Amino Acid Sequence; D017089:Ankyrin Repeat / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014026:Nicotiana / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques
C419263:ACBP2 protein, Arabidopsis; D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins