Localization and characterization of the novel protein encoded by C20orf3

Aysegul Ilhan , Wolfgang Gartner , Anastasiya Nabokikh , Teodora Daneva , Otto Majdic , Gerald Cohen , Georg A. Böhmig , Wolfgang Base , Walter H. Hörl , Ludwig Wagner (1)

1 Department of Nephrology and Dialysis

In this study, we characterized the gene product of open reading frame 3 encoded at human chromosome 20 (C20orf3), which represents a member of the lactonohydrolase super family. Multiple tissue Northern showed ubiquitous expression of the 2.4 kb transcript coding for 416 amino acids (aas), with highest levels in human liver, placenta and the kidney. After recombinant production of protein variants in E.coli and insect cells, antibodies directed against different epitopes within the C20orf3 gene product were generated. Using these immunoreagents the protein expression was demonstrated in the liver, glomerula and tubular structures of the kidney as well as in endothelial cells and arterial wall. Positive staining was also observed at the pancreatic islet of Langerhans. Via immunoblotting we identified three size variants. In line with the results of in silico analysis demonstrating a single transmembran spanning sequence (aas 40-61) at the N-terminus of the full length protein, FACS cell surface staining confirmed a mainly extra cellular localization of the full length protein. Sucrose gradient cell fractionation revealed membrane association of the dominant 50 kDa variant in Hep G2 and Rin-5F cells. The finding of a strong arylesterase activity with β-napthyl acetate and phenyl acetate of the C20orf3 protein containing fractions suggests potential involvement of this protein in enzymatic processes. C20orf3 promoter driven reporter assays, which were verified by gene-specific RT-qPCR showed a strong inhibitory effect of human serum on transcription using the human embryonic kidney cell line Hek 293. In conclusion, we characterized the structure and expression pattern of the C20orf3 gene product. According to a series of analogies with PON family members, we speculate that the C20orf3 gene product represents a new member of this important protein family present at cellular level.

Mots clés: Life Sciences

Aysegul Ilhan, Wolfgang Gartner, Anastasiya Nabokikh, Teodora Daneva, Otto Majdic, et al.. Localization and characterization of the novel protein encoded by C20orf3. Biochemical Journal, 2008, 414 (3), pp.485-495. ⟨10.1042/BJ20080503⟩. ⟨hal-00478994⟩