Identification of over-expressed genes in human renal cell carcinoma by combining suppression subtractive hybridization and cDNA library array.
To isolate the over-expressed genes in human renal cell carcinoma (RCC) and analyze its molecular basis of carcinogenesis, we used the mRNA from human RCC tissues as tester and that from the matched normal kidney tissues as driver to construct the suppression subtractive hybridization library. 379 of the subtracted clones were arrayed onto a nylon membrane and the over-expressed genes were then screened by hybridizing the filter with radioactively labeled cDNA from RCC and matched normal kidney tissues. 67 clones over-expressed in RCC by a factor of 6 or more were sequenced and its identities were analyzed in GenBank database. 4 clones were previously unknown fragments and 2 clones represent KIAA genes. The rest clones were the known genes and some of them were RCC-related, including vascular endothelial growth factor, vimentin and tissue factor. Most of the known genes were the RCC-related genes previously unknown, including zinc ribbon domain-containing 1 protein (ZNRD1), pituitary tumor transforming gene1 (PTTG1). Northern blot and semi-quantitative RT-PCR confirmed that the mRNA levels of the 3 novel fragments and 1 KIAA and 3 known genes were significantly higher in RCC than in the matched normal kidney tissues. Immunohistochemical and Western blot analysis for PTTG1 and ZNRD1 revealed increased protein level in RCC. The over-expressed genes in RCC are the potential molecular targets for diagnosis and therapy and it is very important to understand the molecular mechanism of RCC through the profile of over-expressed genes.
Ai
Junkui
J
Institute of Urology, Peking University & Department of Urology, First Hospital, Peking University, Beijing 100034, China.
Zhang
Zhiwen
Z
Xin
Dianqi
D
Zhu
Hongjian
H
Yan
Quanjian
Q
Xin
Zhongcheng
Z
Na
Yanqun
Y
Guo
Yinglu
Y
D001483:Base Sequence; D002292:Carcinoma, Renal Cell / Q000235:genetics*; D017931:DNA Primers; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D015723:Gene Library; D006801:Humans; D007680:Kidney Neoplasms / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D009363:Neoplasm Proteins / Q000235:genetics; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012333:RNA, Messenger / Q000235:genetics; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D009363:Neoplasm Proteins; D012333:RNA, Messenger