title: Hint1 Up-Regulates IκBα by Targeting the β-TrCP Subunit of SCF E3 Ligase in Human Hepatocellular Carcinoma Cells.
abstract: OBJECTIVE
  There is increasing evidence that histidine triad nucleotide-binding protein 1 (HINT1) is a novel tumor suppressor. In the present study, we investigated the mechanism by which HINT1 promotes the stability of inhibitor of NF-κB α (IκBα) in the cytoplasm of hepatocellular carcinoma (HCC) cells, which was observed in our previous study (Wang et al. in Int J Cancer 124:1526-1534, 2009).
  
  
  METHODS
  We examined HINT1 and IκBα expression in HCC cell lines and determined the effect of HINT1 overexpression and knockdown on IκBα protein and mRNA expression in these cell lines. Then, ubiquitination assays were performed to investigate the effects of HINT1 expression plasmid transfection on IκBα ubiquitination. Next, the interaction between HINT1 and β-TrCP was investigated in immunoprecipitation and immunofluorescence assays.
  
  
  RESULTS
  Our data showed that increased HINT1 expression in HepG2 and SMMC7702 cells markedly increased IκBα protein levels, while decreased HINT1 expression markedly decreased them. Overexpression or knockdown of HINT1 did not alter the transcription of IκBα, but HINT1 inhibited proteasomal IκBα degradation and reduced its ubiquitination levels. This inhibition might occur because HINT1 is a component of the SCF(β-TrCP) E3 ligase, which is responsible for IκBα ubiquitination and degradation.
  
  
  CONCLUSIONS
  This study provides new evidence that HINT1 is a regulator of IκBα through SCF(β-TrCP) E3 ligase. These findings help to clarify the mechanism underlying the anticancer effects of HINT1.
authors.affiliation: Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming City, 650101, Yunnan Province, China.
authors.forename: Zhitian
authors.initials: Z
authors.lastname: Shi
authors.affiliation: Department of Gastroenterological Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming City, 650101, Yunnan Province, China.
authors.forename: Xuesong
authors.initials: X
authors.lastname: Wu
authors.affiliation: Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming City, 650101, Yunnan Province, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Ke
authors.affiliation: Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming City, 650101, Yunnan Province, China. linwang0705@126.com.|, No. 1168 Chunrongxi Road, Chenggong District, Kunming City, 650500, Yunnan Province, China. linwang0705@126.com.
authors.forename: Lin
authors.identifier: http://orcid.org/0000-0001-7383-934X
authors.initials: L
authors.lastname: Wang
mesh_terms: D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D057809:HEK293 Cells; D056945:Hep G2 Cells; D006801:Humans; D020672:I-kappa B Proteins / Q000235:genetics* / Q000378:metabolism; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044843:SKP Cullin F-Box Protein Ligases / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation; D054875:Ubiquitination; D015854:Up-Regulation; D044844:beta-Transducin Repeat-Containing Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: C471644:BTRC protein, human; C063306:HINT1 protein, human; D020672:I-kappa B Proteins; C000606204:NFKBIA protein, human; D009419:Nerve Tissue Proteins; D044844:beta-Transducin Repeat-Containing Proteins; D000072000:NF-KappaB Inhibitor alpha; D044843:SKP Cullin F-Box Protein Ligases
keywords: Beta transducin repeat-containing proteins; HINT1 protein, human; Hepatocellular carcinoma; I-kappa B proteins; NF-kappa B; SKP Cullin F-box protein ligases