Related Experiment Video
Updated: May 22, 2026
Separation of Spermatogenic Cell Types Using STA-PUT Velocity Sedimentation
Published on: October 9, 2013
The threonine protease activity of testes-specific protease 50 (TSP50) is essential for its function in cell
Yu-Yin Li1, Yong-Li Bao, Zhen-Bo Song
1National Engineering Laboratory for Druggable Gene and Protein Screening, Northeast Normal University, Changchun, China.
Background:
Testes-specific protease 50 (TSP50), a newly discovered threonine enzyme, has similar amino acid sequences and enzymatic structures to those of many serine proteases. It may be an oncogene. TSP50 is up-regulated in breast cancer epithelial cells, and ectopic expression of TSP50 in TSP50-deficient Chinese hamster ovary (CHO) cells has been found to promote cell proliferation. However, the mechanisms by which TSP50 exerts its growth-promoting effects are not yet fully understood.
Methodology/Principal Findings:
To delineate whether the threonine protease activity of TSP50 is essential to its function in cell proliferation, we constructed and characterized a mutant TSP50, called TSP50 T310A, which was identified as a protease-dead mutant of TSP50. By a series of proliferation analyses, colony formation assays and apoptosis analyses, we showed that T310A mutation significantly depresses TSP50-induced cell proliferation in vitro. Next, the CHO stable cell line expressing either wild-type or T310A mutant TSP50 was injected subcutaneously into nude mice. We found that the T310A mutation could abolish the tumorigenicity of TSP50 in vivo. A mechanism investigation revealed that the T310A mutation prevented interaction between TSP50 and the NF-κBIκBα complex, which is necessary for TSP50 to perform its function in cell proliferation.
Conclusion:
Our data highlight the importance of threonine 310, the most critical protease catalytic site in TSP50, to TSP50-induced cell proliferation and tumor formation.
