Jianhua Fu, Ph.D.
Assistant Professor
Dr. Fu received his Ph.D. degree in Protein Crystallography from University of Pittsburgh in 1994. He spent a postdoctoral period in the laboratory of Roger Kornberg at Stanford University School of Medicine, where he cracked the phase problem in determining the crystallographic structure of RNA Polymerase II. Dr. Fu received fellowship awards from Universitywide AIDS Research Program (University of California) and American Cancer Society. Since 2000, his laboratory has been funded by the NIH to study the structural mechanism of RNA polymerase II machinery. Dr. Fu joined the faculty at Medical College of Wisconsin in 2007.
Contact Information
Dr. Fu welcomes MCW graduate students to discuss prospects of thesis research in the area of Structural Biology. Potential postdoctoral scientists are encouraged to communicate their interests as well.
Phone: (414) 456-5849
Fax: (414) 456-6510
E-mail: jfu@mcw.edu
Current Topics
Current research in the laboratory is focused on the structural mechanisms of gene transcription and regulation in eukaryotic cells. Transcription is the first step in the expression of genetic information from a cell's chromosomal DNA. In eukaryotic cells, three RNA polymerases, named Pol I, II and III, carry out transcription. Among them, Pol II is responsible for synthesizing protein-coding RNAs, the messenger RNAs (or mRNAs) that direct the synthesis of cellular proteins. Transcription by Pol II is one of the fundamental processes that underlie development, oncogenesis and viral pathogenesis (e.g. HIV-AIDS).
The Pol II transcription system forms a multi-protein machine with the RNA polymerase at its heart and other accessory protein factors around it. Many components of this gigantic apparatus have been discovered over the past 40 years, but understanding is still missing as to how this machine controls its initiation frequency and its RNA synthesis rate, in response to external signals. To gain insights into its mechanism, detailed knowledge of the 3-dimensional (3-D) workings of the machinery is required. Owing to the breakthrough in determining the structure of the free Pol II (Fu et al., Cell, 98: 799-810, 1999; Cramer et al., Sci., 288: 640-649, 2000), it is now feasible to determine the 3-D structure of the apparatus in more extensive fashions. Since Pol II by itself is dormant, we are taking steps to work out Pol II-factor(s) complex structures that correspond to its active functions in vivo.
We are particularly interested in post-initiation steps in the transcription cycle (Figure). Intermediate Pol II-factor complexes involved in this stage of transcription have been shown to be targets of viral activators, restrictor of estrogen alpha-dependent growth of breast epithelial cells, and regulators of gene activities in response to environmental cues. We are working on cocrystal structures of complexes formed between Pol II and factors known to be involved in this process.
Figure: A simplistic model for Pol II initiation, early elongation and reinitiation. Pol II and TBP-TFIIB-TATA promoter complex are each rendered as solvent accessible surfaces. PIC stands for the pre-initiation complex. Cet1/Ceg1 represents the pre-mRNA capping enzyme. DSIF is the negative elongation factor and, pTEFb, the positive elongation factor. Protein-protein interactions within the early elongation complex (EEC) are indicated by two-way arrows. The CTD domain of the largest subunit of Pol II is represented by the zigzag line. Fcp1 is a CTD phosphatase, plays a key role in generating initiation-competent Pol II for reinitiation.
Research Methodology
We take a multidisciplinary approach to analyze 3-D structures of large complexes. We use Protein Biochemistry and Molecular Cloning to obtain pure materials and assessing their activities. We then apply X-ray Crystallography to determine their 3-D structures. X-ray crystallography (figures below) is the primary method for determining atomic structures of biological macro-molecules. Computational techniques are being continually improved in the lab to cope with the complexity of large molecular systems such as complexes of RNA polymerase II. Recent developments in X-ray detector technology and synchrotron instrumentation have greatly facilitated our structural work.
Research Instrumentation
The department houses state-of-the-art instrumentation dedicated to Structural Biology research. The facility includes chromatographic systems for protein purification, an in-house X-ray diffraction core and an automated crystallization machine for high-throughput screening and optimization. High-end computer workstations have been set up for 3-D graphic visualization and fast-speed crystallographic analysis.
Research Training
Graduate students and other research members of the lab receive systematic training in Structural Biology in general, and X-ray Crystallography and Protein Biochemistry in particular. This lab believes in logic, rigor and the validation of scientific results. Training in this lab usually entails instruction, discussion and exploration: from conceiving ideas, designing approaches for testing hypotheses, analyzing complex observations and experimental data, integrating information, to gaining scientific insights.
Selected Publications
"Phasing RNA polymerase II using intrinsically bound Zn atoms: an updated structural model." PA Meyer, P Ye, M Zhang, M-H Suh and J Fu, Structure, 14: 973-982 (2006) .
"Fcp1 directly recognizes the CTD and also interacts with a site on RNA polymerase II distinct from the CTD." M-H Suh, P Ye, M Zhang, S Hausmann, S Shuman, Al Gnatt and J Fu, PNAS, 102: 17314-17319 (2005).
"An agarose-acrylamide composite native gel system suitable for separating ultra-large protein complexes." M-H Suh, P Ye, AB Datta, M Zhang and J Fu, Anal. Biochem., 343: 166-175 (2005).
"CTD-dependent termination of RNA polymerase II transcription by the pre-mRNA 3' end processing factor Pcf11." Z Zhang, J Fu and DS Gilmour, Genes & Dev., 19: 1572-1580 (2005).
"Single molecule imaging of RNA polymerase II using atomic force microscopy." T Rhodin, J Fu, K Umemura, M Gad, S Jarvis, M Ishikawa, Appl. Surf. Sci., 210: 105-111 (2003).
"Exploratory study of RNA polymerase II using dynamic atomic force microscopy." T Rhodin, K Umemura, M Gad, S Jarvis, M Ishikawa and J Fu, Appl. Surf. Sci., 188: 486-488 (2002).
"Structural basis of Transcription: an RNA polymerase II elongation complex at 3.3 Å resolution." AL Gnatt, P Cramer, J Fu, DA Bushnell and RD Kornberg, Science, 292: 1876-1881 (2001).
"Architecture of RNA polymerase II and implications for the transcription mechanism." P Cramer, DA Bushnell, J Fu, AL Gnatt, B Maier-Davis, NE Thompson, RR Burgess, AM Edwards, PR David and RD Kornberg, Science, 288: 640-649 (2000).
"Yeast RNA polymerase II at 5 Å resolution." J Fu, AL Gnatt, DA Bushnell, GJ Jensen, NE Thompson, RR Burgess, PR David and RD Kornberg, Cell, 98: 799-810 (1999).
"Electron crystal structure of an RNA polymerase II transcription elongation complex." CL Poglitsch, GD Meredith, AL Gnatt, GJ Jensen, W-H Chang, J Fu and RD Kornberg, Cell, 98: 791-798 (1999).
"Formation and crystallization of yeast RNA polymerase II elongation complexes." A Gnatt, J Fu and RD Kornberg, J. Biol. Chem., 272: 30799-30805 (1997).