NF-kappaB activates transcription of the RNA-binding factor HuR, via PI3K-AKT signaling, to promote gastric tumorigenesis.
OBJECTIVE
HuR is a RNA-binding factor whose expression is commonly upregulated in some human tumor types. We explored the molecular mechanism underlying HuR elevation and its role in gastric cancer tumorigenesis.


METHODS
HuR expression and subcellular localization were determined by polymerase chain reaction, immunoblot, and immunohistochemical analyses. Its effect on tumor growth was characterized using flow cytometry, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling, and soft agar analyses. Luciferase reporter, chromatin immunoprecipitation, and electrophoretic mobility shift assays were used to measure transcriptional activation by nuclear factor kappaB (NF-kappaB) signaling.


RESULTS
Compared with normal gastric tissues, HuR was expressed at higher levels in gastric tumors, particularly in advanced versus early tumors; this increase was associated with enhanced cytoplasmic translocation of HuR. HuR overexpression increased proliferation of tumor cells, activating the G(1) to S transition of the cell cycle, DNA synthesis, and anchorage-independent growth. Small interfering RNA-mediated knockdown of HuR expression reduced tumor cell proliferation and response to apoptotic stimuli. No genetic or epigenetic alterations of HuR were observed in gastric tumor cell lines or primary tumors; overexpression depended on phosphatidylinositol 3-kinase/AKT signaling and NF-kappaB activity. AKT activation increased p65/RelA binding to a putative NF-kappaB binding site in the HuR promoter, the stability of HuR target transcripts, and the cytoplasmic import of HuR.


CONCLUSIONS
HuR is a direct transcription target of NF-kappaB; its activation in gastric cancer cell lines depends on phosphatidylinositol 3-kinase/AKT signaling. HuR activation by this pathway has proliferative and antiapoptotic effects on gastric cancer cells.
Kang
Min-Ju
MJ
School of Life Sciences and Biotechnology, Korea University, Seoul, Korea.
Ryu
Byung-Kyu
BK
Lee
Min-Goo
MG
Han
Jikhyon
J
Lee
Jin-Hee
JH
Ha
Tae-Kyu
TK
Byun
Do-Sun
DS
Chae
Kwon-Seok
KS
Lee
Bong-Hee
BH
Chun
Hyang Sook
HS
Lee
Kil Yeon
KY
Kim
Hyo-Jong
HJ
Chi
Sung-Gil
SG
D000954:Antigens, Surface / Q000235:genetics*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D016328:NF-kappa B / Q000235:genetics* / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000235:genetics*; D016601:RNA-Binding Proteins / Q000235:genetics*; D013274:Stomach Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D014158:Transcription, Genetic
D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D016328:NF-kappa B; D012334:RNA, Neoplasm; D016601:RNA-Binding Proteins; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt