Related Experiment Videos
The mUBC9 murine ubiquitin conjugating enzyme interacts with the E2A transcription factors
D A Loveys1, M B Streiff, T S Schaefer
1Division of Pediatric Hematology, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Abstract:
The ubiquitin-mediated degradation of cellular proteins requires the sequential activity of E1, E2 and, in some cases, E3 enzymes. Using the yeast two-hybrid system, we have cloned 1.0- and 2.5-kb cDNAs encoding the identical murine E2, or ubiquitin conjugating enzyme by virtue of its interaction with the E2A transcription factor. This cDNA encodes the 158-amino-acid protein, mUBC9, which has considerable sequence homology to UBC9 from Saccharomyces cerevisiae and HUS5 from Schizosaccharomyces pombe and is identical to the human UBC9 protein. HUS5 is essential for DNA damage repair, whereas UBC9 is necessary for G2/M progression. The human protein has been shown to correct the UBC9 defect in yeast. Antisera raised against bacterially expressed mUBC9 fusion protein recognize a murine cellular protein of approximately 18 kDa, corresponding to the predicted mobility. Unlike E2A, the mUBC9 protein level is not regulated by serum growth factors. The activity of the apparent homologues UBC9 and HUS5 suggests that mUBC9 may be involved in the degradation of key nuclear proteins that regulate cell cycle progression.
Insights
Researchers identified a murine ubiquitin conjugating enzyme (mUBC9) crucial for cell cycle progression. This enzyme is homologous to yeast and fission yeast proteins involved in DNA repair and cell division.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Ubiquitin-mediated protein degradation is essential for cellular function, involving E1, E2, and E3 enzymes.
- E2 enzymes, or ubiquitin-conjugating enzymes, play a critical role in the ubiquitination pathway.
Purpose of the Study:
- To clone and characterize the murine homolog of the ubiquitin-conjugating enzyme UBC9.
- To investigate the potential role of mUBC9 in cellular processes like cell cycle progression.
Main Methods:
- Yeast two-hybrid system was employed to identify interacting proteins and clone cDNAs.
- cDNA encoding the murine UBC9 (mUBC9) was isolated and sequenced.
- Antibodies were generated against mUBC9 for protein detection and characterization.
Main Results:
- Identified and cloned 1.0- and 2.5-kb cDNAs encoding the murine UBC9 (mUBC9) protein.
- mUBC9 shares significant sequence homology with yeast UBC9 and fission yeast HUS5, and is identical to human UBC9.
- mUBC9 protein is approximately 18 kDa and its expression is not regulated by serum growth factors.
Conclusions:
- The murine UBC9 (mUBC9) is a conserved ubiquitin-conjugating enzyme with potential roles in cell cycle regulation.
- Homology to yeast UBC9 and fission yeast HUS5 suggests mUBC9 may be involved in degrading key nuclear proteins regulating cell cycle progression.