MOLECULAR STUDIES OF VOLTAGE-SENSITTVE POTASSIUM CHANNELS 
Lily Y. Jan, Ph.D., Investigator 
Voltage-sensitive potassium channels represent a 
diverse group of ion channels found in most cell 
types studied in the animal and the plant king- 
doms. Different combinations of potassium chan- 
nels are found in different cells and are involved in 
a variety of cell functions. In the nervous system, 
they control excitability and modulate the strength 
of synaptic inputs; some of the potassium channels 
have been implicated in the processes of learning 
and memory. 
To understand better how potassium channels 
work and how the tremendous diversity of these 
channels is generated, one would like to study 
these channels biochemically. Although the sparsity 
of these channels has presented a serious problem 
in their purification, the first potassium channel has 
been cloned in Dr. Jan's laboratory by taking advan- 
tage of Drosophila genetics. The initial molecular 
characterizations have provided some clues to 
questions concerning diversity and structural ele- 
ments involved in different channel functions. 
L Alternative Splicing at the Shaker Locus Gener- 
ates Potassium Channel Diversity. 
The Shaker locus in Drosophila gives rise to a 
number of protein products by alternative splicing. 
These Shaker proteins have different amino- and/or 
carboxyl-terminal regions but have the same core 
region, including most of the putative membrane- 
spanning sequences. When in vitro transcribed 
RNA from each Shaker cDNA is injected into 
Xenopus oocytes, it induces inactivating potassium 
channels (A channels) of distinct kinetic properties. 
Moreover, the different Shaker proteins show dif- 
ferent distributions in the central nervous system. 
Thus the different Shaker products are likely to 
form different subtypes of potassium channels; 
these subtypes may have different tissue distribu- 
tions as well as different physiological properties. 
IL Structure-Function Analysis of the Potassium 
Channels Encoded by the Shaker Locus. 
Like other voltage-gated cation channels, the po- 
tassium channels encoded by the Shaker locus con- 
tain intrinsic voltage sensors that detect the electri- 
cal potential across the membrane. These sensors 
are thought to be displaced by depolarization of 
the membrane, triggering conformation changes 
that open the channel. After channel opening, the 
Shaker channels are closed by a specialized inacti- 
vation gate that remains shut as long as depolariza- 
tion persists. To study the molecular basis of chan- 
nel opening and inactivation, Dr. Jan's colleagues 
have altered the primary structure of these potas- 
sium channel components by site-directed muta- 
genesis, expressed mutant channels in Xenopus oo- 
cytes, and characterized the potassium currents 
electrophysiologically. 
The intrinsic voltage sensors are expected to cor- 
respond to charged or polarizable amino acids of 
the ion channel that are located within the mem- 
brane field. It has been proposed that the basic 
amino acids of the S4 sequence, which is found in 
sodium and calcium channels as well as potassium 
channels, may function as voltage sensors. Each of 
these basic residues has been substituted one at a 
time by either a different basic residue or the neu- 
tral residue glutamine. Some of these mutants 
show altered voltage sensitivity of macroscopic cur- 
rent activation and inactivation, without having ob- 
vious effects on other functional properties of the 
channel, such as potassium selectivity or the rate of 
recovery from inactivation. 
In contrast to the voltage sensors, the inactiva- 
tion gate may be outside of the membrane, as indi- 
cated by previous biophysical studies on sodium 
channels and potassium channels. For this reason, 
the effects of mutations of the hydrophilic terminal 
regions are being tested. Preliminary results indi- 
cate that some of these mutations alter the kinetic 
properties of inactivation. 
III. Search for Other Channel Genes. 
Comparative studies can often provide valuable 
clues to structure-function analysis. Using Shaker 
sequences, several groups have obtained other 
channel sequences from mammals, amphibians, 
and snails. Most of these sequences are fairly sim- 
ilar (—70% amino acid identity). They reveal se- 
quences that are highly conserved, as well as those, 
including regularly spaced hydrophobic residues 
within some of the putative membrane-spanning 
regions, that seem to tolerate changes readily. Such 
information is useful in formulating structural mod- 
els for experimental tests. Detailed comparison of 
channel function, similar to what is being carried 
out for the different Shaker proteins, may also be 
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