KMS1 and KMS2, two plant endoplasmic reticulum proteins involved in the early secretory pathway

Pengwei Wang, Eric Hummel, Anne Osterrieder, Andreas J Meyer, Lorenzo Frigerio, Imogen Sparkes, Chris Hawes

Research output: Contribution to journalArticle (Academic Journal)peer-review

24 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)613-28
Number of pages16
JournalPlant Journal
Volume66
Issue number4
DOIs
Publication statusPublished - May 2011

Keywords

  • Amino Acid Sequence
  • Arabidopsis
  • Arabidopsis Proteins
  • Cloning, Molecular
  • Endoplasmic Reticulum
  • Gene Knockdown Techniques
  • Green Fluorescent Proteins
  • Hydrophobic and Hydrophilic Interactions
  • Molecular Sequence Data
  • Protein Transport
  • RNA Interference
  • Recombinant Fusion Proteins
  • SNARE Proteins
  • Secretory Pathway
  • Sequence Alignment
  • Sequence Analysis, Protein
  • Tobacco
  • Journal Article
  • Research Support, Non-U.S. Gov't

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