The sarco(endo)plasmic reticulum calcium ATPase (SERCA) and its regulatory 
partner phospholamban (PLN) are essential for myocardial contractility. Arg(9) → 
Cys (R9C) and Arg(14) deletion (R14del) mutations in PLN are associated with 
lethal dilated cardiomyopathy in humans. To better understand these mutations, 
we made a series of amino acid substitutions in the cytoplasmic domain of PLN 
and tested their ability to inhibit SERCA. R9C is a complete loss-of-function 
mutant of PLN, whereas R14del is a mild loss-of-function mutant. When combined 
with wild-type PLN to simulate heterozygous conditions, the mutants had a 
dominant negative effect on SERCA function. A series of targeted mutations in 
this region of the PLN cytoplasmic domain ((8)TRSAIRR(14)) demonstrated the 
importance of hydrophobic balance in proper PLN regulation of SERCA. We found 
that Arg(9) → Leu and Thr(8) → Cys substitutions mimicked the behavior of the 
R9C mutant, and an Arg(14) → Ala substitution mimicked the behavior of the 
R14del mutant. The results reveal that the change in hydrophobicity resulting 
from the R9C and R14del mutations is sufficient to explain the loss of function 
and persistent interaction with SERCA. Hydrophobic imbalance in the cytoplasmic 
domain of PLN appears to be a predictor for the development and progression of 
dilated cardiomyopathy.