The binding of lamin B receptor to chromatin is regulated by phosphorylation in the RS region.
Binding of lamin B receptor (LBR) to chromatin was studied by means of an in vitro assay system involving recombinant fragments of human LBR and Xenopus sperm chromatin. Glutathione-S-transferase (GST)-fused proteins including LBR fragments containing the N-terminal region (residues 1-53) and arginine-serine repeat-containing region (residues 54-89) bound to chromatin. The binding of GST-fusion proteins incorporating the N-terminal and arginine-serine repeat-containing regions to chromatin was suppressed by mild trypsinization of the chromatin and by pretreatment with a DNA solution. A new cell-free system for analyzing the cell cycle-dependent binding of a protein to chromatin was developed from recombinant proteins, a Xenopus egg cytosol fraction and sperm chromatin. The system was applied to analyse the binding of LBR to chromatin. It was shown that the binding of LBR fragments to chromatin was stimulated by phosphorylation in the arginine-serine repeat-containing region by a protein kinase(s) in a synthetic phase egg cytosol. However, the binding of LBR fragments was suppressed by phosphorylation at different residues in the same region by a kinase(s) in a mitotic phase cytosol. These results suggested that the cell cycle-dependent binding of LBR to chromatin is regulated by phosphorylation in the arginine-serine repeat-containing region by multiple kinases.
Takano
Makoto
M
Course of Biosphere Science, Graduate School of Science and Technology, Niigata University, Japan.
Takeuchi
Masaki
M
Ito
Hiromi
H
Furukawa
Kazuhiro
K
Sugimoto
Kenji
K
Omata
Saburo
S
Horigome
Tsuneyoshi
T
D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D002843:Chromatin / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D005260:Female; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D008938:Mitosis; D010063:Ovum / Q000737:chemistry / Q000502:physiology; D010449:Peptide Mapping; D010750:Phosphoproteins / Q000032:analysis / Q000378:metabolism; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D000097682:Lamin B Receptor
D002843:Chromatin; D004268:DNA-Binding Proteins; D005982:Glutathione Transferase; D008565:Membrane Proteins; D010750:Phosphoproteins; D010770:Phosphotransferases; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; D000097682:Lamin B Receptor; C093413:lamina-associated polypeptide 2