IDENTIFICATION OF THE HUMAN PROSTATIC-CARCINOMA ONCOGENE PTI-1 BY RAPID EXPRESSION CLONING AND DIFFERENTIAL RNA DISPLAY

被引:99
作者
SHEN, RQ
SU, ZZ
OLSSON, CA
FISHER, PB
机构
[1] COLUMBIA UNIV,COLL PHYS & SURG,INST CANC RES,CTR COMPREHENS CANC,DEPT PATHOL,NEW YORK,NY 10032
[2] COLUMBIA UNIV,COLL PHYS & SURG,INST CANC RES,CTR COMPREHENS CANC,DEPT UROL,NEW YORK,NY 10032
关键词
DNA COTRANSFECTION; CREF-TRANS; 6; REVERSE TANSCRIPTION-PCR; IN VITRO TRANSLATION; MUTATED EF-1-ALPHA;
D O I
10.1073/pnas.92.15.6778
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Elucidating the relevant genomic changes mediating development and evolution of prostate cancer is paramount for effective diagnosis and therapy. A putative dominant-acting nude mouse prostatic carcinoma tumor-inducing gene, PTI-1, has been cloned that is expressed in patient-derived human prostatic carcinomas but not in benign prostatic hypertrophy or normal prostate tissue, PTI-1 was detected by cotransfecting human prostate carcinoma DNA into CREF-Trans 6 cells, inducing tumors in nude mice, and isolating genes displaying increased expression in tumor-derived cells by using differential RNA display (DD), Screening a human prostatic carcinoma (LNCaP) cDNA library with a 214 bp DNA fragment found by DD permitted the cloning of a full-length 2.0-kb PTI-1 cDNA. Sequence analysis indicates that PTI-1 is a gene containing a 630-bp 5' sequence and a 3' sequence homologous to a truncated and mutated form of human elongation factor 1 alpha. In vitro translation demonstrates that the PTI-1 cDNA encodes a predominant approximate to 46-kDa protein. Probing Northern blots with a DNA fragment corresponding to the 5' region of PTI-1 identifies multiple PTI-1 transcripts in RNAs from human carcinoma cell Lines derived from the prostate, lung, breast, and colon. In contrast, PTI-1 RNA is not detected in human melanoma, neuroblastoma, osteosarcoma, normal cerebellum, or glioblastoma multiforme cell lines. By using a pair of primers recognizing a 280-bp region within the 630-bp 5' PTI-1 sequence, reverse transcription-PCR detects PTI-1 expression in patient-derived prostate carcinomas but not in normal prostate or benign hypertrophic prostate tissue. In contrast, reverse transcription-PCR detects prostate-specific antigen expression in all of the prostate tissues. These results indicate that PTI-1 may be a member of a class of oncogenes that could affect protein translation and contribute to carcinoma development in human prostate and other tissues. The approaches used, rapid expression cloning with the CREF-Trans 6 system and the DD strategy, should prove widely applicable for identifying and cloning additional human oncogenes.
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页码:6778 / 6782
页数:5
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