title: The threonine protease activity of testes-specific protease 50 (TSP50) is essential for its function in cell proliferation.
abstract: 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.
  
  
  RESULTS
  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.
  
  
  CONCLUSIONS
  Our data highlight the importance of threonine 310, the most critical protease catalytic site in TSP50, to TSP50-induced cell proliferation and tumor formation.
authors.affiliation: National Engineering Laboratory for Druggable Gene and Protein Screening, Northeast Normal University, Changchun, China.
authors.forename: Yu-Yin
authors.initials: YY
authors.lastname: Li
authors.forename: Yong-Li
authors.initials: YL
authors.lastname: Bao
authors.forename: Zhen-Bo
authors.initials: ZB
authors.lastname: Song
authors.forename: Lu-Guo
authors.initials: LG
authors.lastname: Sun
authors.forename: Ping
authors.initials: P
authors.lastname: Wu
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.forename: Cong
authors.initials: C
authors.lastname: Fan
authors.forename: Yan-Xin
authors.initials: YX
authors.lastname: Huang
authors.forename: Yin
authors.initials: Y
authors.lastname: Wu
authors.forename: Chun-Lei
authors.initials: CL
authors.lastname: Yu
authors.forename: Ying
authors.initials: Y
authors.lastname: Sun
authors.forename: Li-Hua
authors.initials: LH
authors.lastname: Zheng
authors.forename: Guan-Nan
authors.initials: GN
authors.lastname: Wang
authors.forename: Yu-Xin
authors.initials: YX
authors.lastname: Li
mesh_terms: D000483:Alleles; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D016466:CHO Cells; D020134:Catalytic Domain; D049109:Cell Proliferation / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D016328:NF-kappa B / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D017354:Point Mutation; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013912:Threonine
chemical_list: D016328:NF-kappa B; D013912:Threonine; D004317:Doxorubicin; D051550:I-kappa B Kinase; D012697:Serine Endopeptidases; C446379:testis-specific protease 50