Effect of proteasome inhibitors on expression of HLA-G isoforms.
HLA-G primary transcript is alternatively spliced into a number of mRNAs. In addition to full length HLA-G1 protein isoform these mRNAs might also encode truncated HLA-G protein isoforms lacking one or two extracellular domains. Whereas HLA-G1 protein isoform is regularly identified, truncated HLAG protein isoforms are not detected even if all alternative spliced mRNAs are present in cells. The absence of entire domain(s) renders the truncated HLA-G protein isoforms incapable of binding peptide and beta2-microglobulin. These features of truncated HLA-G protein isoforms may result in their rapid degradation by proteasomes. Here we show that despite the presence of all alternatively spliced HLA-G transcripts in JEG-3 cells pretreated with proteasome inhibitors only a full length HLA-G1 protein isoform was regularly detected. Interestingly, immunoblot analysis showed slight increase of HLA-G1 protein in cells pretreated with proteasome inhibitors, although the expression of HLA-G1 transcript was basically not affected. Expression of HLA-G3 transcript increased in JEG-3 cells pre-incubated with LLL, however, neither HLA-G3 nor other HLA-G short protein isoform was regularly detected. In K562 transfectants proteasome inhibitor LLL greatly enhanced expression of the HLA-G1 and -G2 transcripts as well as corresponding protein isoforms. Flow cytometry analysis showed that in cells pre-treated with proteasome inhibitors cell surface expression of HLA-G1 protein decreased but the quantity of intracellularly localized HLA-G antigens increased. Altogether our results suggest that truncated HLA-G proteins isoforms are not detected in JEG-3 cells as a result of their instability and the low translation efficiency of truncated HLA-G transcripts.
Poláková
K
K
Cancer Research Institute, Slovak Academy of Sciences, 833 91 Bratislava, Slovak Republic. exonpola@savba.sk
Bandzuchová
E
E
Bystrická
M
M
Pancuchárová
H
H
Russ
G
G
D002822:Choriocarcinoma / Q000378:metabolism / Q000473:pathology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D006680:HLA Antigens / Q000235:genetics / Q000378:metabolism*; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020014:K562 Cells / Q000378:metabolism / Q000473:pathology; D007976:Leupeptins / Q000494:pharmacology*; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D015853:Cysteine Proteinase Inhibitors; D006680:HLA Antigens; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I; D007976:Leupeptins; D020033:Protein Isoforms; D012333:RNA, Messenger; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde