Prostate-specific membrane antigen and its truncated form PSM'.
BACKGROUND
Prostate specific membrane antigen (PSMA) is a type II transmembrane protein overexpressed in prostate cancer as well as in the neovasculature of several non-prostatic solid tumors. In addition to full-length PSMA, several splice variants exist in prostatic tissue. Notably, the N-terminally truncated PSMA variant, termed PSM', is prevalent in healthy prostate, and the ratio of PSMA/PSM' mRNA has been shown to correlate with cancer progression. The widely accepted hypothesis is that the PSM' protein is a translation product arising from the alternatively spliced PSM' mRNA.


METHODS
Differential ultracentrifugation, cell surface biotinylation, Western blotting, and enzyme activity measurement were used to study the origin and localization of the PSMA/PSM' variants in prostatic (LNCaP; lymph-node carcinoma of the prostate) and non-prostatic (HEK293) cell lines. These experiments were further complemented by analysis of the N-glycosylation patterns of the PSMA/PSM' proteins and by site-directed mutagenesis.


RESULTS
We identified PSM' protein expression in both the LNCaP cell line and a non-cancerous HEK293 human cell line transfected with a plasmid encoding full-length PSMA. Differential centrifugation revealed that PSM' is localized predominantly to the cytosol of both these cell lines and is proteolytically active. Furthermore, the PSM' protein is N-glycosylated by a mixture of high-mannose and complex type oligosaccharides and therefore trafficked beyond the cis-Golgi compartment.


CONCLUSIONS
Our data suggest that the PSM' protein is likely not generated by alternative splicing of the PSMA gene but by different mechanism, probably via an endoproteolytic cleavage of the full-length PSMA.
Mlcochová
Petra
P
Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
Barinka
Cyril
C
Tykvart
Jan
J
Sácha
Pavel
P
Konvalinka
Jan
J
D000230:Adenocarcinoma / Q000378:metabolism* / Q000473:pathology; D002460:Cell Line; D045744:Cell Line, Tumor; D006031:Glycosylation; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D008247:Lysosomes / Q000378:metabolism; D008297:Male; D008861:Microsomes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D017430:Prostate-Specific Antigen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D022821:RNA Splice Sites / Q000235:genetics; D014162:Transfection
D022821:RNA Splice Sites; D017430:Prostate-Specific Antigen