Identification and characterization of a novel human cDNA encoding a 21 kDa pRb-associated protein

Gene. 2001 Jan 24;263(1-2):85-92. doi: 10.1016/s0378-1119(00)00585-0.

Abstract

The retinoblastoma protein (pRb) functions as a critical master regulator in cell cycle regulation, which is an important cell-regulatory process, through its interaction with various cellular proteins. Using the C-terminus of human pRb and the yeast two-hybrid system, a novel protein named RBP21 that contains 187 amino acid residues with a calculated molecular weight of 21 kDa was identified as a pRb-binding protein. Sequence analysis indicates that RBP21 shares homology with other retinoblastoma-binding proteins in the pRb-binding motif LxCxE at the C-terminal region. In vitro specific interaction between pRb and RBP21 was confirmed using in vitro translation products. When overexpressed in COS-7 cells, RBP21 could co-immunoprecipitate with pRb. This interaction requires the LxCxE motif of RBP21 and the entire pocket region of pRb. Each point mutation of the conserved amino acid residues in pRb-binding motif of RBP21 abolished its specific interaction with pRb. RH mapping result showed that this novel gene was mapped to chromosome region 15q21.1-21.3. Northern blot analysis suggested that RBP21 was widely expressed in various human tissues and cancer cell lines. When expressed in HeLa cells as a green fluorescent protein fusion, RBP21 was distributed throughout the cell.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenovirus E1A Proteins*
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Binding Sites
  • Blotting, Northern
  • COS Cells
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism
  • Cell Cycle Proteins
  • Chromosome Mapping
  • Chromosomes, Human, Pair 15 / genetics
  • Cloning, Molecular
  • DNA, Complementary / chemistry
  • DNA, Complementary / genetics*
  • Female
  • Gene Expression
  • Green Fluorescent Proteins
  • HL-60 Cells
  • HeLa Cells
  • Humans
  • K562 Cells
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Male
  • Microscopy, Fluorescence
  • Molecular Sequence Data
  • Molecular Weight
  • Nuclear Proteins
  • Protein Binding
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Repressor Proteins
  • Retinoblastoma Protein / chemistry
  • Retinoblastoma Protein / metabolism*
  • Sequence Analysis, DNA
  • Tissue Distribution
  • Tumor Cells, Cultured
  • Two-Hybrid System Techniques

Substances

  • Adenovirus E1A Proteins
  • Carrier Proteins
  • Cell Cycle Proteins
  • DNA, Complementary
  • EID1 protein, human
  • Luminescent Proteins
  • Nuclear Proteins
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • Retinoblastoma Protein
  • Green Fluorescent Proteins

Associated data

  • GENBANK/AF109873