Identification and characterization of 9D7, a novel human protein overexpressed in renal cell carcinoma.
With the objective of discovering novel tumor-associated antigens of the cancer/testis type, we compared the transcriptional profiles of renal cell carcinoma (RCC) and non-tumorous kidney and further screened for genes expressed in RCC and testis, but not other normal tissues. In a first step, a representational difference analysis library consisting of approximately 1,900 RCC cDNA clones was generated. Clones were then spotted onto filters and hybridized with cDNA probes derived from a testis-specific cDNA library, a pool of RCCs and a pool of 10 healthy normal tissues, respectively. Based on strong hybridization signals with both RCC and testis, but not normal tissue probes, 185 clones were sequenced and annotated. After EST-database comparison, 35 clones were selected for experimental analysis, including conventional and quantitative RT-PCR as well as Northern blotting. Clone 9D7 showed strong mRNA expression in RCC as well as in several other major tumor types. In normal tissues there was little or no mRNA expression with the exception of heart. 9D7 was cloned to full-size and found to represent a novel human gene containing 5 exons residing on chromosome 14. Alternative splicing within exon 1 generates 2 open-reading-frames consisting of 717 or 435 bp corresponding to predicted proteins of 239 or 145 amino acids. 9D7 shows high homology (227/239 amino acids or 95% identity) to a growth factor-inducible gene of Rattus norvegicus involved in apoptosis. In situ hybridization as well as immunohistochemical analysis using 9D7-specific antisera confirmed overexpression of 9D7 in RCCs as compared to normal kidney tissue.
Klade
Christoph S
CS
Boehringer Ingelheim Austria GmbH, Research and Development, Vienna, Austria. cklade@intercell.co.at
Dohnal
Alexander
A
Fürst
Walter
W
Sommergruber
Wolfgang
W
Heider
Karl-Heinz
KH
Gharwan
Helen
H
Ratschek
Manfred
M
Adolf
Günther R
GR
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D002292:Carcinoma, Renal Cell / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D020869:Gene Expression Profiling; D006801:Humans; D017403:In Situ Hybridization; D007680:Kidney Neoplasms / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000951:Antigens, Neoplasm; D017931:DNA Primers; D009363:Neoplasm Proteins; D012333:RNA, Messenger