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KMS1 and KMS2, two plant endoplasmic reticulum proteins involved in the early secretory pathway
- Wang, Pengwei, Hummel, Eric, Osterrieder, Anne, Meyer, Andreas J., Frigerio, Lorenzo, Sparkes, Imogen, Hawes, Chris
- The plant journal 2011 v.66 no.4 pp. 613-628
- Arabidopsis, RNA interference, electron microscopy, endoplasmic reticulum, hydrophobicity, membrane proteins, protein transport, topology
- We have identified two endoplasmic reticulum (ER)-associated Arabidopsis proteins, KMS1 and KMS2, which are conserved among most species. Fluorescent protein fusions of KMS1 localised to the ER in plant cells, and over-expression induced the formation of a membrane structure, identified as ER whorls by electron microscopy. Hydrophobicity analysis suggested that KMS1 and KMS2 are integral membrane proteins bearing six transmembrane domains. Membrane protein topology was assessed by a redox-based topology assay (ReTA) with redox-sensitive GFP and confirmed by a protease protection assay. A major loop domain between transmembrane domains 2 and 3, plus the N- and C-termini were found on the cytosolic side of the ER. A C-terminal di(tri)-lysine motif is involved in retrieval of KMS1 and deletion led to a reduction of the GFP-KMS1 signal in the ER. Over-expression of KMS1/KMS2 truncations perturbed ER and Golgi morphology and similar effects were also seen when KMS1/KMS2 were knocked-down by RNA interference. Microscopy and biochemical experiments suggested that expression of KMS1/KMS2 truncations inhibited ER to Golgi protein transport.