Min-Hao Kuo
Associate Professor
- Research Associate, 1998-1999, University of Virginia
- Post-doctoral fellow, 1996-1998, University of Rochester
- Ph.D., 1995, University of Rochester
- B.S., 1986, Department of Medical Technology, National Taiwan University
401 Biochemistry Building
Michigan State University
East Lansing, MI 48824-1319
Office: 517-355-0163
Lab: 517-432-8786
FAX: 517-353-9334
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Min-Hao Kuo Research Interests
The research in our lab revolves around studies of the molecular roles of post-translational modifications of proteins, and more importantly, the mechanisms employed by these modifications to exert their biological functions. Current research projects fall into one of three areas:
1. Transcriptional regulation and other chromatin functions regulated by histone modifications, including acetylation, phosphorylation, and methylation.
2. Protein-protein interactions that are controlled by specific post-translational modifications. The tumor suppressor protein p53 is one of the models used in our research.
3. Extension and further applications of the tethered catalysis/yeast two-hybrid system developed in our lab.
Post-translational modifications play pivotal roles in numerous cellular functions. A variety of modifications can be covalently conjugated to selective amino acid residues of the target proteins to regulate the biochemical, enzymatic, conformational, and biophysical characteristics of the underlying acceptor proteins. The best known chemical modifications include phosphorylation, acetylation, methylation, glycosylation, etc. Furthermore, small peptides, such as ubiquitin and SUMO, are conjugated via iso-peptide bonds to regulate, among other effects, the stability and intracellular localization of the modified proteins. Just as there are many modifications each protein may receive, there are also multiple molecular mechanisms how these modifications regulate the behaviors of the underlying proteins. We use several model systems to study the “what” and “how” of functions of protein modifications. The following are some of our current projects.
1. Histone acetylation and transcriptional regulation
Histone acetyltransferases play important roles in transcriptional regulation, DNA damage repair, recombination, and DNA replication. Misregulation and mutations of several such acetyltransferases result in devastating consequences in animals, including embryonic death and cancer. Using a prototypic histone acetyltransferase, Gcn5, from the budding yeast Saccharomyces cerevisiae as the model, we try to understand how acetylation of histones facilitates transcription at the molecular level. Toward this end, genetic suppressor screens have been performed to identify extragenic mutations that correct the gcn5- transcriptional defects. These Bypass of Gcn5 Requirement, or BGR, mutations may unveil key steps conveying Gcn5 signaling or acting in parallel with Gcn5 in transcription. We have identified several such BGR alleles in two genome-wide mutagenesis screens. In addition, a targeted histone mutagenesis project also revealed novel Gcn5-INdependent, or GIN mutant alleles of histones that support efficient transcription without acetylation. Biophysical studies of nucleosomal arrays composed of these mutant histones may help uncover the structure-function relationship of acetylation and transcription.
2. Tethered catalysis/yeast two-hybrid system to identify novel acetylated histone-binding
proteins
A fast-increasing body of evidence shows that one of the recurring, but to this day largely unexplored, functions of post-translational modifications is to regulate protein-protein interactions. It is common that a given modification exerts multiple functions via recruiting different proteins. Identifying and studying these proteins will allow us to dissect the corresponding molecular functions of the recruiting modification, as well as how this modification cross-talks to other cellular signaling pathways. Targeted intervention of some of these interactions may be the underlying rationale for drug design. To understand the effects of post-translational modifications on protein-protein interactions, we have developed a tethered catalysis/yeast two-hybrid (TC/Y2H) system.
In TC/Y2H, the protein of interest (X) is fused to the cognate modifying enzyme in such a way that the physical linkage forces an efficient modification of X. As a control, a catalytically inactive mutant enzyme is fused to X in parallel. Because of this inactivating mutation, protein X remains unmodified. Using these two otherwise identical fusion proteins, yeast two-hybrid screen is then conducted to identify proteins that interact specifically with the protein X possessing the modification. In theory, proteins that only interact with X in the absence of the modification can also be identified.
The TC/Y2H has been used in our lab in several screens for modification-dependent protein-protein interactions. We have identified novel acetylated histone-binding proteins with different chromatin-related functions such as transcription, chromatin assembly, centromere, and sporulation. Biochemical and proteomic applications of TC/Y2H are being tested.
3. Example of TC/Y2H application: Acetylation of tumor suppressor p53 recruits multiple regulators
In the era of “omics”, protein interaction networks (a.k.a. “interactomes”) have quickly become information-rich databases that warrant invaluable knowledge as to how cellular proteins and pathways are integrated for intricate responses to environmental stimuli. However, most of the existing databases severely bias toward modification-independent interactions. To explore the largely unchartered territory of interactomes instigated by post-translational modifications, we have been using the TC/Y2H method on several important proteins whose modifications are known to play critical roles in different biological functions.
p53 is the guardian of genome integrity. Loss-of-function mutations of p53 have been associated with 50% of all tumors. A variety of post-translational modifications are triggered by DNA damages which in turn activate p53 for functions in cell cycle arrest, transcriptional regulation of multiple genes, and apoptosis if the damages are too severe. To begin to understand how these p53 activating signals are relayed to the corresponding interacting proteins that modulate and coordinate different molecular functions of p53, we use the TC/Y2H system to screen for mammalian proteins that interact preferentially with an acetylated p53. Thus far, we have identified a slew of functionally diversified proteins in this screen. Functional studies of these novel p53 binding proteins and further applications of the TC/Y2H on other modified p53 are in progress.