Ching Kung
Vilas Professor of Genetics and Molecular Biology
http://www.molbio.wisc.edu/kung
Microbial ion channels and sensory transductions detail in web page http://www.molbio.wisc.edu/kung/
Our laboratory studies sensory transductions in microbes. We have pioneered the study of ion channels in Paramecium, yeast, and E. coli. We have devised methods to electrically record from these microbes to follow their ion-channel activities. See 1 2 for reviews. We produced giant E. coli cells and applied patch-clamp electrodes on their membranes. Our survey revealed the activities of ion channels that respond directly to membrane stretch. We went on to clone the gene of one such channel 3, the crystal structures of which were then solved by the Rees laboratory. The most surprising finding is that purified channel protein reconstituted into a lipid bilayer retains mechanosensitivity 4. This finding indicates that the lipid bilayer transmit force to the protein. Today, these bacterial channels are the best biochemical and biophysical models to study mechanosensitivity 5 2. The force-from-lipid concept is currently being applied to other channels, including Shaker. Patch-clamp survey of the plasma membrane of Saccharomyces cerevisiae revealed a mechanosensitive channel and a K + -specific channel. The latter, an 8-TM 2-pore-domain channel, has been subjected to genetic dissections. Gain-of-function mutations with constitutive currents were found to be at the base of S6 and S8 6 7, anticipating the later identification of channel gate by crystallography. We also found a mechanosensitive cation channel, TRPY1, in the vacuolar membrane of yeast 8. It is a member of TRPs, the “transient-receptor-potential” channel superfamily that plays key roles in animal sensations. TRPY1 gain-of-function (GOF) mutants revealed the importance of aromatic amino acids in channel gating 9. One such GOFs is a phenylalanine located at the base of S5 10. Mutations at the same point have yielded GOF phenotypes in Drosophila TRPC1 and rat TRPV1. GOF mutations in mouse TRPML3 (varitint-waddler) and human TRPV4 (autosomal dominant brachyomia) are nearby. These coincidences indicate a common gating mechanism among all TRP channels 11. Most recently, we have expressed rat TRPV1 and V4 in yeast. These animal channels retain their molecular characteristics: V1 responding to heat but not to hypotonic swelling; V4 to swelling but not heat. Swelling has been modeled to activate enzyme(s), producing polyunsaturated fatty acids (PUFAs) to open TRPV4 in mammalian cells. This model relegates mechanosensitivity to the enzyme and not the channel. Yeast has only a single Δ9 fatty-acid monodesaturase and cannot make PUFAs, suggesting an alternative mechanism for TRPV4 activation 12. However, rat TRPV4 is expressed in internal membranes of yeast, hampering patch-clamp examination. Ion channels govern its ciliary motion and therefore its swimming behavior of Paramecium as in animals. We have isolated paramecium behavioral mutants, characterized their electrophysiological defects. We have invented a method to clone genes based on Paramecium phenotypes. A class of over-excitable mutants in Paramecium have mutations at the C-lobe of calmodulin (CaM), while a class of under-excitable mutants at the N-lobe 13. We showed by patch clamp that CaM is a detachable subunit of a Ca 2+-activated channel. That CaM is often an ion-channel subunit and has a functional bipartition has now been widely observed among animal ion channels 14. The Paramecium genome has been completely sequenced 15. It contains 298 K +-channel genes! (Only 91 in human, 1 in budding yeast, 1 in E. coli).
Ph.D., University of Pennsylvania, 1968
Postdoctoral Research: Indiana University and UCLA
Research Interests
Research Description
2 Martinac, B., Saimi, Y. and Kung, C. Ion channels in microbes. Physiol Rev. 88 (4), 1449 (2008).
3 Sukharev, S. I. et al. A large-conductance mechanosensitive channel in E. coli encoded by mscL alone. Nature. 368 (6468), 265 (1994).
4 Sukharev, S. I., Blount, P., Martinac, B. and Kung, C. Mechanosensitive channels of Escherichia coli: the MscL gene, protein, and activities. [Review] [94 refs]. Annual Review of Physiology. 59, 633 (1997).
5 Anishkin, A. and Kung, C. Microbial mechanosensation. Curr Opin Neurobiol. 15 (4), 397 (2005).
6Loukin, S. H. et al. Random Mutagenesis Reveals a Region Important for Gating of the Yeast K+ Channel, Ykc1. EMBO. 16, 4817 (1997).
7 Loukin, S. H. and Saimi, Y. K+-Dependent Composite Gating of the Yeast K+ Channel, Tok1. Biophysical J. 77 (6), 3060 (1999).
8 Zhou, X. L. et al. The transient receptor potential channel on the yeast vacuole is mechanosensitive. Proc Natl Acad Sci U S A. 100 (12), 7105 (2003).
9 Zhou, X., Z. Su, A. Anishkin, W.J. Haynes, E.M. Friske, S.H. Loukin, C. Kung and Y. Saimi. Yeast screen show aromatic residues at the end of the sixth helix anchor TRP-channel gate. Proc. Natl. Acad. Sci. 104 (39), 15555 (2007).
10 Su, Z. et al. Yeast gain-of-function mutations reveal structure function relationships conserved among different subfamilies of transient receptor potential channels. Proc Natl Acad Sci U S A. (2007).
11 Myers, B. R., Saimi, Y., Julius, D. and Kung, C. Multiple unbiased prospective screens identify TRP channels and their conserved gating elements. Journal of General Physiology. 132, 481 (2008).
12 Loukin, S. H., Su, Z., and Kung, C. Hypotonic shocks activate rat TRPV4 in yeast in the absence of polyunsaturated fatty acids. FEBS Lett. 583 (4), 754 (2009).
13 Kink, J. A. et al. Mutations in paramecium calmodulin indicate functional differences between the C-terminal and N-terminal lobes in vivo. Cell. 62 (1), 165 (1990).
14 Saimi, Y. and Kung, C. Calmodulin as an ion channel subunit. Annu Rev Physiol. 64, 289 (2002).
15 Aury, J. M. et al. Global trends of whole-genome duplications revealed by the ciliate Paramecium tetraurelia. Nature 444 (7116), 171 (2006).
Representative Publications
- Kung, C. (2005) A possible unifying principle for mechanosensation. Nature. 436: 647-654.
- Su, Z.-W., et al. (2007) Yeast gain-of-function mutations reveal structure-function relationships conserved among different subfamilies of transient receptor potential channels. Proc. Natl. Acad. Sci. 104: 19607-1961.
- Martinac, B. et al. (2008) Ion channels in microbes. Phys. Reviews. 88: 1449-1490.
- Myers, B.R., et al. (2008) Multiple unbiased prospective screens identify TRP channels and their conserved gating elements. J. Gen. Physiol. 132: 481-486.
- Loukin, S.H. et al. (2009) Hypotonic shock activates rat TRPV4 in yeast in the absence of polyunsaturated fatty acids. FEBS Lett. 583: 754-758.