Figure S2.
CCDC134 is an ER-resident protein. (A) Multiple sequence alignment of CCDC134 protein across species. UniProt entry names: CC134_HUMAN, G7N3Z8_MACMU, CC134_MOUSE, E1BVM7_CHICK, CC134_XENTR, and A0A8M6Z583_DANRE. Boxes above the alignment indicate consensus prediction from JPred4. Red: helix; blue lines: deleted regions of deletion mutant constructs used in this study (1: Δ2–25, 2: Δ25–57, 3: Δ57–91, 4: Δ91–133, 5: Δ133–156, 6: Δ156–178, 7: Δ178–229); pink line: predicted N-linked glycosylation on NQT sequon; green lines: signal peptide (1–22) or ER retention signal (226–229). (B and C) Representative confocal images microscopy of HeLa cells transfected as indicated. Green: anti-CCDC134; red: anti-Calreticulin; blue: DAPI. Scale bar: 10 μm. Ev: empty vector. (D) Immunoblots of proteins precipitated from supernatant (SN) and whole-cell extracts (WCE) of HEK293T cells transfected as indicated. (E and F) Immunoblots of lysate from indicated CAL-1 cells untreated (E) or treated with EndoH (H) or PNGase F (F) (F). Left and right panels of immunoblots in F are presented with two different exposures. Ev: empty vector; SH: strep-HA tag; short exp.: short exposure. (G–J) Immunoblots of indicated CAL-1 cells stimulated with R848 (5 µg/ml, for 0–1 h) (G) or untreated (H–J). F: full-length; C: cleaved form. Short exp.: short exposure. In D–J, data are representative of two independent experiments. Source data are available for this figure: SourceData FS2.

CCDC134 is an ER-resident protein. (A) Multiple sequence alignment of CCDC134 protein across species. UniProt entry names: CC134_HUMAN, G7N3Z8_MACMU, CC134_MOUSE, E1BVM7_CHICK, CC134_XENTR, and A0A8M6Z583_DANRE. Boxes above the alignment indicate consensus prediction from JPred4. Red: helix; blue lines: deleted regions of deletion mutant constructs used in this study (1: Δ2–25, 2: Δ25–57, 3: Δ57–91, 4: Δ91–133, 5: Δ133–156, 6: Δ156–178, 7: Δ178–229); pink line: predicted N-linked glycosylation on NQT sequon; green lines: signal peptide (1–22) or ER retention signal (226–229). (B and C) Representative confocal images microscopy of HeLa cells transfected as indicated. Green: anti-CCDC134; red: anti-Calreticulin; blue: DAPI. Scale bar: 10 μm. Ev: empty vector. (D) Immunoblots of proteins precipitated from supernatant (SN) and whole-cell extracts (WCE) of HEK293T cells transfected as indicated. (E and F) Immunoblots of lysate from indicated CAL-1 cells untreated (E) or treated with EndoH (H) or PNGase F (F) (F). Left and right panels of immunoblots in F are presented with two different exposures. Ev: empty vector; SH: strep-HA tag; short exp.: short exposure. (G–J) Immunoblots of indicated CAL-1 cells stimulated with R848 (5 µg/ml, for 0–1 h) (G) or untreated (H–J). F: full-length; C: cleaved form. Short exp.: short exposure. In D–J, data are representative of two independent experiments. Source data are available for this figure: SourceData FS2.

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