[Effect of KLF6 and its splice variant KLF6V on proliferation and differentiation of human hepatocellular carcinoma HepG2 cells].
OBJECTIVE
To investigate the roles of Kruppel-like factor 6 (KLF6) and its splice variant KLF6V on suppressing growth and inducing differentiation of human hepatocellular carcinoma hepG2 cells.


METHODS
KLF6V cDNA was amplificated by RT-PCR from human hepatocellular carcinoma (HCC) tissue and then sequenced. The recombinant vectors expressing KLF6 variant (KLF6V) were constructed using molecular clone technology based on established plasmid pcDNA3.1A(-)/wtKLF6. KLF6V or KLF6-transfected HepG2 cells were established after being screened with G418. Growth activity of HepG2/KLF6 or HepG2/KLF6V cells was detected by in vitro MTT assay. Expression of p21WAF1 or cyclin D1 protein was detected by Western blot, and expressions of AFP or ALB protein were measured by radioimmunoassay.


RESULTS
A novel alternatively spliced transcript of the human KLF6 gene was found and its sequencing revealed that the variant form of KLF6 lacked 126nt and its encoded protein products had a deletion of 42 aa near the COOH-terminal amino acid in comparison with full-length KLF6. Although KLF6 alternative splicing was present in both normal and cancerous tissues, expression of the KLF6 splice variants seemed to be up-regulated in HCCs tissues. The isoform of KLF6 proteins antagonized the ability of wild-type KLF6 to up-regulate p21 expression or down-regulate cyclin D1 expression and suppress HepG2 cell proliferation. KLF6 gene increased albumin production and decreased alpha fetoprotein production of the cells.


CONCLUSIONS
The isoform of KLF6 protein, present in HCC tissue, antagonizes the ability of wild-type KLF6 to suppress cell proliferation and induce cellular differentiation.
Pan
Xiu-cheng
XC
Department of Infectious Diseases, Affiliated Hospital of Xuzhou Medical College, Xuzhou 221002, China. xzpxc68@126.com
Chen
Zhi
Z
Ji
Fang
F
Guo
Zhong-sheng
ZS
Chen
Min
M
Fu
Juan-juan
JJ
D002454:Cell Differentiation; D049109:Cell Proliferation; D018076:DNA, Complementary; D015972:Gene Expression Regulation, Neoplastic; D056945:Hep G2 Cells; D006801:Humans; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors / Q000235:genetics*; D020033:Protein Isoforms / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D014162:Transfection
D018076:DNA, Complementary; C105465:KLF6 protein, human; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins