Genome-wide RNAi screen reveals a role for multipass membrane proteins in endosome-to-golgi retrieval.
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SFT2D2, a human paralog of SFT2D3, is required for endosome-to-Golgi retrieval of cargo proteins
"We further demonstrate a role for three multipass membrane proteins, SFT2D2, ZDHHC5, and GRINA, in endosome-to-Golgi retrieval."
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SFT2D2 is partially homologous to yeast Sft2p protein and represents an uncharacterized mammalian homolog of the SFT2 family
"SFT2D2 is partially homologous to the yeast Sft2p protein, a genetic interactor of Sed5p (the yeast syntaxin 5 protein) and affects post-Golgi trafficking (Conchon et al., 1999). However, the mammalian homologs (SFT2D1, SFT2D2, and SFT2D3) are uncharacterized."
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SFT2D2 localizes to both perinuclear membranes and endosomal structures containing the retromer complex
"We find that SFT2D2 localizes to both perinuclear membranes and structures positive for the retromer CSC protein, VPS35. The colocalization of SFT2D2 with VPS35 is especially apparent after treatment with nocodazole to depolymerize microtubules"
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SFT2D2 exhibits specific colocalization with syntaxin 6 SNARE protein, indicating functional interaction with post-Golgi membrane fusion machinery
"We find that SFT2D2 partially colocalizes with syntaxin 5 but exhibits almost complete colocalization with syntaxin 6 (indicated by arrowheads in the immunofluorescence images shown in Figure 3C). Quantitation of the colocalization of SFT2D2 with a number of post-Golgi SNARE proteins is shown graphically in Figure 3C (right panel) and confirms that SFT2D2 resides in a compartment strongly positive for syntaxin 6."
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SFT2D2 depletion alters SNARE protein distribution without affecting their total expression levels
"when SFT2D2 expression is silenced by RNAi, we observed a marked change in the fluorescence intensity of several SNARE proteins, with syntaxin 6 and VAMP3 exhibiting the strongest change (Figure 3D). An example of the altered fluorescence intensity for VAMP3 is shown along with the graph in Figure 3D. Surprisingly, however, changes in the fluorescence intensity are not the result of changes in overall levels of the respective SNARE proteins (Figure 3E)."
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SFT2D2 knockdown causes accumulation of endocytic cargo in peripheral puncta with reduced TGN localization
"Images of anti-CD8 localization obtained in the primary screen show an accumulation of antibody in peripheral puncta in SFT2D2-silenced cells compared to control HeLa cells (Figure 3A) as well as reduced levels of antibody at the TGN."