BACKGROUND: Phospholamban (PLN) is an effective inhibitor of the 
sarco(endo)plasmic reticulum Ca(2+)-ATPase, which transports Ca(2+) into the SR 
lumen, leading to muscle relaxation. A mutation of PLN in which one of the 
di-arginine residues at positions 13 and 14 was deleted led to a severe, early 
onset dilated cardiomyopathy. Here we were interested in determining the 
cellular mechanisms involved in this disease-causing mutation.
METHODOLOGY/PRINCIPAL FINDING: Mutations deleting codons for either or both 
Arg13 or Arg14 resulted in the mislocalization of PLN from the ER. Our data show 
that PLN is recycled via the retrograde Golgi to ER membrane traffic pathway 
involving COP-I vesicles, since co-immunoprecipitation assays determined that 
COP I interactions are dependent on an intact di-arginine motif as PLN RDelta14 
did not co-precipitate with COP I containing vesicles. Bioinformatic analysis 
determined that the di-arginine motif is present in the first 25 residues in a 
large number of all ER/SR Gene Ontology (GO) annotated proteins. Mutations in 
the di-arginine motif of the Sigma 1-type opioid receptor, the beta-subunit of 
the signal recognition particle receptor, and Sterol-O-acyltransferase, three 
proteins identified in our bioinformatic screen also caused mislocalization of 
these known ER-resident proteins.
CONCLUSION: We conclude that PLN is enriched in the ER due to COP I-mediated 
transport that is dependent on its intact di-arginine motif and that the 
N-terminal di-arginine motif may act as a general ER retrieval sequence.