Characterization of kinesin-like proteins in silkworm posterior silkgland cells

BmCREC Is an Endoplasmic Reticulum (ER) Resident Protein and Required for ER/Golgi Morphology

“…Using the PSG, silkworm kinesins were cloned, some cargo molecules of BmKinesin-1 were identified (14), and the roles of the COPI complex in Golgi morphology and PSG tube expansion were unraveled (4), suggesting that the silkworm PSG is a model for studying intracellular trafficking and key developmental processes (14,40). Here, we identify BmCREC as a novel cargo protein of COPI vesicles driven by BmKinesin-1 in PSG and reveal its role in ER/Golgi morphology maintenance and PSG tube expansion.…”

“…Pulldown Analysis and Immunoprecipitation-Pulldown assay with silkworm PSG homogenates was performed as reported previously (14). For immunoprecipitation in PSG, the homogenates were incubated with the affinity-purified antibodies in lysis buffer (20 mM HEPES, 320 mM sucrose, 120 mM NaCl, 2 mM EGTA, 1 mM MgCl 2 , pH 6.8).…”

“…Anti-BmCREC and anti-EGFP rabbit polyclonal antibodies were produced and purified according to a previous report (14). Anti-silkworm ␣/␤Ј/␥-COP (4), anti-BmKinesin-1 (14), anticalnexin (Abcam), anti-EGFP (MBL), anti-Drosophila GM130 (Abcam), anti-KDELR (Abcam), anti-lamin (20), and anti-tubulin (Sigma) antibodies were also used.…”

“…Anti-silkworm ␣/␤Ј/␥-COP (4), anti-BmKinesin-1 (14), anticalnexin (Abcam), anti-EGFP (MBL), anti-Drosophila GM130 (Abcam), anti-KDELR (Abcam), anti-lamin (20), and anti-tubulin (Sigma) antibodies were also used.…”

“…Cell Transfection, RNAi, Immunostaining, and Fluorescence Resonance Energy Transfer (FRET) Assay-Cell transfection and immunostaining were performed as reported previously (14).…”