Zheng Fan, Ph.D.
My research interest is to understand the function and structure of ion channels.
One area of my research is studying inwardly rectifying potassium channels, specifically their structure-function relationships, gating mechanisms, and regulation under physiological conditions. This is a long-term research direction of this laboratory. Our present interest is on ATP-sensitive potassium (KATP) channels, a subfamily of inwardly rectifying potassium channels. ATP plays a pivotal role in the energy economy of the cell. Additionally, in many cell types such as muscle cells and pancreatic islet beta-cells, ATP has important roles in transducing cellular signals that involve a particular type of potassium channel: the KATP channel. In islet beta-cells, for example, insulin release in response to hyperglycemia is mediated by a remarkable inhibition of the KATP channel. My laboratory has been working on research concerned with the molecular basis for the biophysical activity of KATP channels. Among the goals of this research is to increase our understanding of how KATP channels couple energy metabolism to cellular processes under physiological and pathophysiological circumstances. Another goal of this research is to elucidate the roles of membrane phospholipids in regulating KATP channels and other inwardly rectifying potassium channels. This part of the research project builds on our previous pioneering studies in this area. Currently, we are interested in the cellular and molecular processes associated with this regulatory mechanism and their physiological relevance.
The second area of my research deals with the gating process of a bacterial potassium channel, KcsA. KcsA is the first ion channel of known structure, and its X-ray structures are available for several conformations. A great amount of evidence has indicated that the pore region of this channel, which contains a selectivity filter for potassium ions, has a structure similar to that of most mammalian potassium channels. KcsA is therefore used as a prototypic model to study the structural basis of ion channel function. In 2005 we reported for the first time an inactivation process occurring at the selectivity filter of the KcsA channel. Ion channel inactivation describes a channel closing process that follows channel activation. Activation-coupled inactivation allows ion channels to block ion flow in the presence of a stimulating signal, whereby ion channels control fundamental physiological processes such as formation of activation potentials in excitable cells. Two major mechanisms of such inactivation have been identified. Among them, N-type inactivation has been analyzed in unprecedented, definitive structural detail. Prior to our study, the understanding of another type of inactivation, known as C-type inactivation, was based on functional studies. These studies implicated that C-type inactivation was related to the selectivity filter. However, although structural data obtained from KcsA showed an inactivated conformation at the selectivity filter, activation-coupled inactivation had not been observed in this channel. This is inconsistent with the selectivity filter hypothesis, thereby puzzling researchers. Our 2005 paper demonstrates that the KcsA channel has an activation process and an associated inactivation process. Furthermore, the inactivation of KcsA exhibits many similarities to C-type inactivation of mammalian channels. These findings provide a rationale that links the structural data available for KcsA to the C-type inactivation, and have justified KcsA to be a simplified but compelling model for deciphering the structural basis of channel inactivation.
Most recently, our work has also focused on the pathogenesis of maternally inherited dilated cardiomyopathy (DCM) and arrhythmias associated with sodium channel mutations. This is a new area of research that was brought to this laboratory through an extramural collaborative research activity. Our colleagues at the Shanghai Institute of Cardiovascular Diseases have identified the disease in a patient family, and the genetic study has localized a mutant gene locus that encodes the cardiac sodium channel SCN5A. We performed functional studies on the corresponding mutant sodium channel and determined the phenotypic changes caused by this mutation. A major finding of our study is that both DMC and arrhythmias in the patients of the family are causally associated with the sodium channel mutation. Previously, it was believed that cardiac sodium channel mutations primarily caused abnormalities of electrical activity in heart, such as Long-QT Syndrome, Brugada Syndrome, and conductance block. Our study suggests a possible pathogenic link between progressive DCM and sodium channel mutations. Two recent papers also reported an association between SCN5A mutations and cardiomyopathy. Our study has expanded the repertoire of SCN5A mutations associated with DCM. We are currently studying why and how some SCN5A mutations cause structural damage to the heart muscle.
Current Techniques Utilized
- Methods of Electrophysiology: Patch-clamp recordings (whole-cell/single-channel configurations), single-channel recording in planar lipid bilayer, fluorescent measurement of cross-membrane ion flex
- Methods of Molecular Biology: Site-directed mutagenesis, cloning of cDNA, analysis of mRNA expression using Northern blotting/quantitative RT-PCR/RNAase protection
- Methods of Protein Biochemistry: Chromatography purification of membrane proteins, detection of proteins using immunoblotting/immunoprecipitation, reconstitution of membrane proteins into planar lipid bilayer/protoliposomes.
- Methods of Lipid Biochemistry: Thin layer chromatography
- Methods of Cell Biology: Cell culture, transfection/infection of mammalian cells, immunostaining and confocal visualization of protein distribution in cultured cells
Selected Recent Publications
- Teng S., Gao L., Paajanen V., Pu J., Fan Z. (2009) Readthrough of nonsense mutation W822X in the SCN5A gene can effectively restore expression of cardiac Na+channels.Cardiovascular Res. doi:10.1093/cvr/cvp116 (Note: This paper was published with an accompanying editorial article)
- Ge J., Sun A., Paajanen V., Wang S., Su C., Yang Z., Li Y., Wang S., Jia J., Wang K, Zou Y, Gao L., Wang K., Fan Z. (2008) Molecular and clinical characterization of a novel SCN5A mutation associated with atrioventricular block and dilated cardiomyopathy. Circulation: Arrhythmia and Electrophysiology 1:83-92. (Note: This paper was published with an accompanying editorial article; both were selected as the only two “Editor’s Picks” in the same issue of the journal)
- Vaithianathan T., Liu P., Asuncion-Chin M., Fan Z., Dopico A.M. (2008) Direct regulation of BK channels by phosphatidylinositol 4, 5-bisphosphate as a novel signaling mechanism. J. Gen. Physiol. 132:13-28. (Note: This paper was published with an accompanying commentary article)
- Nishimura H., Yang Y., Lau K., Kuykindoll R., Fan Z., Yamaguchi K., and Yamamoto T. (2007) Aquaporin-2 water channel in developing quail kidney: Possible role in programming adult fluid homeostasis. Am. J. Physiol. (Regul Integr Comp Physiol) doi:10.1152/ajpregu.00163
- Yang Y., Cui Y., Fan Z., Cook G., and Nishimura H. (2006) Two distinct aquaporint-4 cDNAs isolated from medullary cone of quail kidney. Comp. Biochem. Physiol. 147:84-93
- Gao L., Mi X., Paajanen V., Wang K., and Fan Z. (2005) From the Cover: Activation-coupled inactivation in the bacterial potassium channel KcsA. Proc Natl Acad Sci U S A. (via track II) 102:17630-17635. (Note: This paper was featured with four others on the cover of Dec 6, 2005 issue of the journal)
- Gosmanov A.R., Fan Z., Mi X., Schneider E.G., and Thomason D.B. (2004) ATP-sensitive potassium channels mediate hyperosmotic stimulation of NKCC in slow-twitch muscle. Am. J. Physiol (Cell Physiol) 286:C586-595
- Yang Y., Cui Y., Wang W., Zhang L., Bufford L., Sasaki S., Fan Z., Nishimura H. (2004) Molecular and functional characterization of a vasotocin-sensitive aquaporin water channel in quail kidney. Am. J. Physiol. (Regul Integr Comp Physiol) 287:R915-924
- Fan Z., Gao L., and Wang W. (2003) Phosphatidic acid stimulates cardiac KATP channels like phosphatidylinositols, but with novel gating kinetics. Am. J. Physiol (Cell Physiol). 284: C94-C102
- Wang C., Wang K., Wang W., Cui Y., and Fan Z. (2002) Compromised ATP binding as a mechanism of phosphoinositide modulation of ATP-sensitive K+channels. FEBS Lett. 532:177-182
- Cui Y., and Fan Z. Mechanism of Kir6.2 channel inhibition by sulfhydryl modification: pore block or allosteric gating? (2002) J. Physiol. 540: 731-741
- Cui Y., Wang W., and Fan Z. (2002) Cytoplasmic vestibule of the weak inward rectifier Kir6.2 potassium channel. J. Bio. Chem. 277: 10523-10530