Research Interests
The overall goal of our laboratory research is to understand the regulatory roles of chromatin modifications in cell growth and development responding environmental stresses
Three major foci in our laboratory research projects: i) the regulatory roles of chromatin modifications in cell growth and development; ii) the molecular mechanism of filamentous growth in fungi; iii) develop a genome-wide survey of ChIP-on-Chip technology to map functional transcription network in cell growth and development.
I. The regulatory roles of chromatin modifications in cell growth and development responding environmental stresses Chromatin is composed of histone octamers (histones H2A, H2B, H3 and H4), which associate with DNA into nucleosomes. It has become increasingly clear that the various histone modifications can have distinct effects at different loci. There appears to be a ¡§cross-talk ¡§ between the modifications. These chromatin modification enzymes act ¡§synergistically¡¨ and the results are consistent with a mechanism in which the histone modifications generate binding pattern effectors that can specifically recognize the chromatin structures and initiate gene transcription or repression. Thus, regulated alterations of chromatin structure are integral to activation and repression of gene expression and it is hypothesized as the ¡§ Histone Code¡¨.
The main interest of my research is to study the molecular mechanism of chromatin modifications that lead to the formation of transcriptionally active or inactive complexes, furthermore, to investigate the affected genes and modifications of genetic material involved in cell development and differentiation. Because the fundamental aspects of gene expression are very similar in yeast, fungi, plants and mammals, we have utilized baker¡¦s yeast, Saccharomyces cerevisiae, Arabidopsis thaliana and mammalian cell cultures as the experimental organisms to investigate the critical issues in the relationship between chromatin metabolism and transcriptional regulation. The major current research projects are listed as following:
1.Studies of the molecular mechanism of co-activator recruitment in gene transcriptional regulation by chromatin remodeling activities and histone modifiers in response to stresses.
2.Purification of IPL1 (Aurora) histone kinase complexes and functional characterization in chromosome segregation during cell cycle progression.
3.Functional characterization of histone demethylase associated complexes (Jumonji family) in chromatin modifications and gene silencing in S. cerevisiae. Figure 1.Model of the establishment of ¡§ordered recruitment¡¨ of hisotone
phosphorylation
, acetylation and TBP in gene activation (from Lo et al. EMBO J. 2005) Figure 2. Structure of the tGCN5 / coA / hisotne H3Pi complex(from Clements et al. Mol.cell, 2003)
Figure 3. The five JmjC domain-containing histone demethylases(HDMTs) in S. cerevisiae. (from Tu et al. JBC, 2007)
II. The molecular mechanism of filamentous growth in fungi -Chromatin modifications in the regulation of FLO genes expression involved in pseudohyphal development in S. cerevisiae.
Systemic fungal infection has become more and more common, especially in immune system compromised patients. Scientists have extensively explored numerous fungal virulence factors to identify potential targets for effective anti-fungal treatments. It has been proposed that the virulence of fungal infection, such as Candida albicans, is dependent on three essential factors: 1) the ability to switch between different morphogenic forms (yeast-like and pseudohyphae or true hyphae); 2) an efficient recognition and adhesion to host cells; and 3) the ability to penetrate into host cells. Among these factors, the formation of pseudohyphae or true hyphae and the fungal adherence to host cells are regarded as the most critical steps in the initial phase of infection. Previously, I identified a gene, FLO11 (flocculin 11), which encoded the first characterized cell surface flocculin required for pseudohyphae formation and invasive growth in budding yeast. Most of FLO genes are repressed in normal growth condition and expressed by environmental stress, such as low glucose and nitrogen starvation. Thus, to understand the mechanisms controlling pseudohyphal development, it is essential to investigate how the expression of FLO genes is regulated and what the function of this class of cell wall adhesive proteins is in yeast dimorphism. Answers to these questions will be critical to understand the molecular mechanism of the host virulence in its pathogenic counterparts. Moreover, results from our studies on this subject may provide insights to discover potential targets to design effective drugs against fungal infection (Fig.4). The major current research projects are listed as following:
1. Studying the role of chromatin modifications in the regulation of FLO genes expression involved in pseudohyphal development in S. cerevisiae.
2. Developing the potential targets to design effective drugs against fungal infection by chemical genetics screening. Figure 4.FLO11 is required for psedohyphal formation and invasive growth. (A). The morpholorgy of pseudohyphe in budding yeast S. cerevisiae. FLO11 is required for both invasive growth (B) and filamentous growth (C). (from Lo & Dranginis, Mol. Bio. Cell. 1998)
III. Develop a genome-wide survey of ¡§ChIP-on-Chip¡¨ technology to map functional transcription network in cell growth and development The histone code remains a hot topic because of the influence of chromatin modification in regulation of eukaryotic transcription. However, the true extent of the diversity of ¡¥transcriptional network¡¨ is remains largely unknown, until now, the recent ¡§ChIP-on-chip¡¨ technology exists to address this issue. ¡§ChIP-on-chip¡¨ is a genome-wide chromatin immunoprecipitation (ChIP) experiment aimed to identify unknown targets by hybridization of immunoprecipitated DNA to a microarray (chip). The classical approaches for finding target-gene are relied on interpreting altered mRNA levels and therefore cannot distinguish direct versus indirect target genes. This new ¡§ChIP-on-chip¡¨ strategy avoid the disadvantage of classical approached and provides the exciting opportunity to address questions concerning the identity of the genes that have the potential to be directly regulated by an individual transcription factor. These new ChIP-based approaches have been developed over the past few years, first in yeast and then in the more complex human system. Several promoter-enriched microarraies have been used to search for target genes are commercial available, recently. Our laboratory is developing a new modified method to ultimate this powerful genome-wide survey of ChIP-on-Chip technology in order to map the functional transcription network of variable cell systems in response to environmental stresses. Figure 5. Schematic of modified ¡§ChIP-on-chip¡¨ technique.
Tu, S., Bulloch, E.M.M., Yang, L., Ren, C., Huang, W.-C., Hsu, P.-H., Chen, C.-H., Liao, C.-L., Yu, H.-M., Lo*,W.-S., Freitas*, M.A. and Tsai*, M. D. (2007) Identification of histone demethylases in Sacchromyces cerevisiae. J. Biol. Chem.282(19):14262-71 (*Corresponding authors )
Nayak, V., Zhao, K., Wyce, A., Schwartz, M.F., Lo,W.-S., Berger, S.L. and Marmorstein, R. (2006) Structure and dimerization of the kinase domain from yeast Snf1, a member of the Snf1/AMPK protein family. Structure 14(3): 477-485.
Lo,W.-S., Gamache, E.R., Harry, K.W., Yang, D., Pillus, L. and Berger, S.L. (2005) Activator-targeted Snf1-mediated histone H3 phosphorylation promotes TBP recruitment through distinct promoter-specific mechanisms. EMBO J. 24:997-1008
Lo,W.-S., Henry, K.W., Schwartz, M.F. and Berger, S.L. (2004) Histone modification patterns during gene activation. Method in Enzymology 377:130-153
Clements, A., Poux, A.N., Lo,W.-S., Pillus, L., Berger, S.L. and Marmorstein R. (2003) Structural basis for histone and phospho-histone binding by the GCN5 histone acetyltransferase. Molecular Cell, 12(2): 461-473
Henry, K.W., Wyce, A., Lo,W.-S., Duggan, L.J., Emre, N.C.T., Kao, C.-F., Pillus, L., Osley, M. A. and Berger, S. L. (2003) H2B ubiquitylation and deubiquitylation through the SAGA-associated hydrolase Ubp8 are both required for transcriptional activation. Gene & Dev., 17(21):2648-2663
Lo,W.-S., Duggan, L., Belotserkovskya, R., Emre, N.C.T., Lane, W., Shiekhattar, R. and Berger, S.L. (2001) Snf1 is a histone kinase which works in concert with the histone acetyltransferase Gcn5 to regulate transcription.Science, 293:1142-1146.
Lo,W.-S., Trievel, R.C., Rojas, J.R., Duggan, L., Hsu, J.-Y., Allis, C.D., Marmorstein, R. and Berger, S.L. (2000) Phosphorylation of Serine 10 in Histone H3 Is Functionally Linked In Vitro and In Vivo to Gcn5-Mediated Acetylation at Lysine 14. Molecular Cell 5(6):917-926.
Lo,W.-S. and Dranginis, A.M. (1999) The regulation of FLO11, a cell surface flocculin required for pseudohyphae formation and invasion, by STE12, TEC1 and FLO8. Current Genetics 35:439
Lo,W.-S. and. Dranginis A.M (1998) The cell surface flocculin Flo11p required for pseudohyphae formation and invasion by Saccharomyces cerevisiae. Molecular Biology of the Cell, 9(1): 161-172
Lo,W.-S., Raiteses, E.I. and Dranginis, A.M. (1997) Development of pseudohyphae by embedded haploid and diploid yeast.Current Genetics, 32: 197-202.
Lo,W.-S. and Dranginis, A.M. (1996) FLO11, a yeast gene related to the STA genes, encodes a novel cell surface flocculin. Journal of Bacteriology, 178(24): 7144-7151.3.
Lo,W.-S. and Bartelt, D.C. (1996) Construction of an Aspergillus nidulans cDNA library suitable for use in yeast two hybrid screen. Fungal Genetics Newsletter , 43:39-40.
