hERG Potassium Channels in Drug Discovery and Development - Ion Channels and Their Inhibitors - page 156

conductance are described [ 16 - 18 ]. The genetic mutations reduce hERG channel
function, which could have a deleterious effect on cardiac electrical activity. These
mutations will not only contribute to a better understanding of channels function-
ing, but also form the basis of developing therapies aimed at correcting hERG
channel dysfunction. The magnitude of hERG current in a cardiac myocyte is
mainly determined by total number of channels on the plasma membrane, proba-
bility that any given channel is open and conductance of a single channel.
1.4 hERG Protein Trafficking
hERG channel biogenesis involves synthesis of a core-glycosylated monomer in the
endoplasmic reticulum (ER) (135 kD band on western blot) followed by co-
assembly into a tetramer that is subsequently transported to the Golgi. In the
Golgi-complex glycosylation occurs resulting in the addition of sugar moieties to
each subunit [ 19 ]. Therefore, western blot analysis of hERG channels expressed
in mammalian cells gives two bands at 135 kD (ER) and 155 kD (Golgi and
plasma membrane) if the channels are properly assembled and trafficked to
the plasma membrane but only one band (135 kD) appear if they do not traffick
from the ER. Mutations that affect either subunit assembly in the ER or trafficking
from the ER to the plasma membrane will result in a trafficking failure. To date
there are very few studies that have addressed the molecular basis of subunit
assembly and domain interactions in hERG [ 20 , 21 ].
Various approaches have been used to restore normal hERG trafficking in vitro
and in vivo under different physiological and pathological conditions. Especially
the trafficking-deficient mutations have become a focus of interest recently, since
most of these mutations give rise to channel proteins capable of conducting IK r
current when intracellular trafficking is restored in vitro [ 22 ]. Mutations that affect
either subunit assembly in ER or trafficking from ER to plasma membrane will
result in a trafficking failure. hERG channel proteins are synthesized in the ER then
it is transported to the cell surface of the Golgi apparatus. During this process,
hERG proteins undergo two crucial steps of glycosylation. Initially newly
synthesized immature hERG channels undergo asparagine (N)-linked core glyco-
sylation in the ER. Consecutively, immature proteins are transported to Golgi
apparatus, where complex glycosylation occurs. Finally, the fully glycosylated
form of the hERG wild-type (WT) channel is inserted into the cell membrane
[ 23 , 24 ]. Misfolded and incompletely assembled proteins are common side products
of protein synthesis in the ER. Analysis of the hERG sequence showed a putative
ER retention signal present in the C terminus of hERG . Despite its presence in wild-
type hERG proteins, these channels are not trafficking deficient. The C-terminal
104 amino acids mask and inactivate this retention signal in wild-type channels
and that truncation of the C terminus leads to exposure of the R-G-R sequence,
resulting in mistrafficking [ 25 , 26 ]. In mammalian cells, another of quality control
mechanisms consist of ER-associated and cytoplasmic chaperones. Identified two