Identification of candidates for interacting partners of the tail domain of DcNMCP1, a major component of the Daucus carota nuclear lamina-like structure.
NMCP/CRWN (NUCLEAR MATRIX CONSTITUENT PROTEIN/CROWDED NUCLEI) is a major component of a protein fibrous meshwork (lamina-like structure) on the plant inner nuclear membrane. NMCP/CRWN contributes to regulating nuclear shape and nuclear functions. An NMCP/CRWN protein in Daucus carota (DcNMCP1) is localized to the nuclear periphery in interphase cells, and surrounds chromosomes in cells in metaphase and anaphase. The N-terminal region and the C-terminal region of DcNMCP1 are both necessary for localizing DcNMCP1 to the nuclear periphery. Here candidate interacting partners of the amino acid position 975-1053 of DcNMCP1 (T975-1053), which is present in the C-terminal region and contains a conserved sequence that plays a role in localizing DcNMCP1 to the nuclear periphery, are screened for. Arabidopsis thaliana nuclear proteins were subjected to far-Western blotting with GST-fused T975-1053 as a probe, and signals were detected at the positions corresponding to ∼70, ∼40, and ∼18 kDa. These ∼70, ∼40, and ∼18 kDa nuclear proteins were identified by mass spectrometry, and subjected to a yeast 2-hybrid (Y2H) analysis with T975-1053 as bait. In this analysis, the ∼40 kDa protein ARP7, which is a nuclear actin-related protein possibly involved in regulating chromatin structures, was confirmed to interact with T975-1053. Independently of the far-Western blotting, a Y2H screen was performed using T975-1053 as bait. Targeted Y2H assays confirmed that 3 proteins identified in the screen, MYB3, SINAT1, and BIM1, interact with T975-1053. These proteins might have roles in NMCP/CRWN protein-mediated biologic processes.
a Laboratory of Crop Physiology, Research Faculty of Agriculture , Hokkaido University , Sapporo-shi , Hokkaido , Japan.
Ryota
R
Mochizuki
a Laboratory of Crop Physiology, Research Faculty of Agriculture , Hokkaido University , Sapporo-shi , Hokkaido , Japan.
Daisuke
D
Tsugama
a Laboratory of Crop Physiology, Research Faculty of Agriculture , Hokkaido University , Sapporo-shi , Hokkaido , Japan.
Michihiro
M
Yamazaki
a Laboratory of Crop Physiology, Research Faculty of Agriculture , Hokkaido University , Sapporo-shi , Hokkaido , Japan.
Kaien
K
Fujino
a Laboratory of Crop Physiology, Research Faculty of Agriculture , Hokkaido University , Sapporo-shi , Hokkaido , Japan.
Kiyoshi
K
Masuda
D018552:Daucus carota / Q000166:cytology* / Q000378:metabolism*; D034881:Nuclear Lamina / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D000072417:Protein Domains; D021381:Protein Transport
D010940:Plant Proteins
actin; chromatin; lamina-like structure; nuclear membrane; plants; protein-protein interaction; transcription factor