Abstract
BackgroundWe previously examined expression of Long Interspersed Element-1 (LINE-1) in a variety of prostate cancer cells including hormone-dependent LNCaP cells. These studies demonstrated expression and sub-cellular localization of LINE-1 proteins, ORF1p, with antibody 4H1, and ORF2p, with antibody chA1-L1.ResultsHere we conduct immunoprecipitation/mass spectrometry analysis using chA1-L1 antibody against ORF2p in LNCaP cells. Our results indicate that antigens detected by the antibody include the transcriptional regulator BCLAF1. We show that chA1-L1 recognizes BCLAF1 using siRNA knockdown and overexpression of a tagged BCLAF1. We also show that chA1-L1 antibody recognizes ORF2p in HEK293 cells overexpressing LINE-1. Further, analysis of ORF2p (chA1-L1) and BCLAF1 foci using immunofluorescence in LNCaP cells showed significant colocalization.ConclusionsOverall, our findings indicate that chA1-L1 antibody recognizes both BCLAF1 and ORF2p but the majority of antigen recognized in LNCaP cells is BCLAF1.
Highlights
We previously examined expression of Long Interspersed Element-1 (LINE-1) in a variety of prostate cancer cells including hormone-dependent LNCaP cells
Overall, our findings indicate that chA1-L1 antibody recognizes both BCL2 Associated Transcription Factor 1 (BCLAF1) and ORF2p but the majority of antigen recognized in LNCaP cells is BCLAF1
We demonstrated robust ORF2p expression in many prostate cancer cell lines, most of which was expressed as punctate foci localized in the nucleus
Summary
We previously examined expression of Long Interspersed Element-1 (LINE-1) in a variety of prostate cancer cells including hormone-dependent LNCaP cells. We characterized LINE-1 expression, retrotransposition frequency, and protein localization in prostate cancer cells [13]. Our findings demonstrated most prostate cancer cell lines expressed ORF1p, which was primarily localized in the cytoplasm. We utilized a newly available ORF2p antibody, chA1-L1, to characterize endogenous ORF2p expression in prostate cancer cells [14].
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