Bachelor of Science, National Taiwan University (1999)
Master of Science, National Taiwan University (2001)
Doctor of Philosophy, National Defense Medical Center (2010)
Chromatin structure is determined by nucleosome positioning, histone modifications, and DNA methylation. How chromatin modifications are coordinately altered under pathological conditions remains elusive. Here we describe a stress-activated mechanism of concerted chromatin modification in the heart. In mice, pathological stress activates cardiomyocytes to express Brg1 (nucleosome-remodeling factor), G9a/Glp (histone methyltransferase), and Dnmt3 (DNA methyltransferase). Once activated, Brg1 recruits G9a and then Dnmt3 to sequentially assemble repressive chromatin-marked by H3K9 and CpG methylation-on a key molecular motor gene (Myh6), thereby silencing Myh6 and impairing cardiac contraction. Disruption of Brg1, G9a or Dnmt3 erases repressive chromatin marks and de-represses Myh6, reducing stress-induced cardiac dysfunction. In human hypertrophic hearts, BRG1-G9a/GLP-DNMT3 complex is also activated; its level correlates with H3K9/CpG methylation, Myh6 repression, and cardiomyopathy. Our studies demonstrate a new mechanism of chromatin assembly in stressed hearts and novel therapeutic targets for restoring Myh6 and ventricular function. The stress-induced Brg1-G9a-Dnmt3 interactions and sequence of repressive chromatin assembly on Myh6 illustrates a molecular mechanism by which the heart epigenetically responds to environmental signals. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.
View details for DOI 10.1016/j.bbamcr.2016.03.002
View details for PubMedID 26952936
Chromosomal rearrangements involving the mixed-lineage leukemia (MLL) gene occur in primary and treatment-related leukemias and confer a poor prognosis. Studies based primarily on mouse models have substantially advanced our understanding of MLL leukemia pathogenesis, but often use supraphysiological oncogene expression with uncertain implications for human leukemia. Genome editing using site-specific nucleases provides a powerful new technology for gene modification to potentially model human disease, however, this approach has not been used to re-create acute leukemia in human cells of origin comparable to disease observed in patients. We applied transcription activator-like effector nuclease-mediated genome editing to generate endogenous MLL-AF9 and MLL-ENL oncogenes through insertional mutagenesis in primary human hematopoietic stem and progenitor cells (HSPCs) derived from human umbilical cord blood. Engineered HSPCs displayed altered in vitro growth potentials and induced acute leukemias following transplantation in immunocompromised mice at a mean latency of 16 weeks. The leukemias displayed phenotypic and morphologic similarities with patient leukemia blasts including a subset with mixed phenotype, a distinctive feature seen in clinical disease. The leukemic blasts expressed an MLL-associated transcriptional program with elevated levels of crucial MLL target genes, displayed heightened sensitivity to DOT1L inhibition, and demonstrated increased oncogenic potential ex vivo and in secondary transplant assays. Thus, genome editing to create endogenous MLL oncogenes in primary human HSPCs faithfully models acute MLL-rearranged leukemia and provides an experimental platform for prospective studies of leukemia initiation and stem cell biology in a genetic subtype of poor prognosis leukemia.
View details for DOI 10.1182/blood-2015-05-646398
View details for Web of Science ID 000365451100009
View details for PubMedID 26311362
Acute lymphoblastic leukemia (ALL) is the most common childhood cancer; however, its genetic diversity limits investigation into the molecular pathogenesis of disease and development of therapeutic strategies. Here, we engineered mice that conditionally express the E2A-PBX1 fusion oncogene, which results from chromosomal translocation t(1;19) and is present in 5% to 7% of pediatric ALL cases. The incidence of leukemia in these mice varied from 5% to 50%, dependent on the Cre-driving promoter (Cd19, Mb1, or Mx1) used to induce E2A-PBX1 expression. Two distinct but highly similar subtypes of B cell precursor ALLs that differed by their pre-B cell receptor (pre-BCR) status were induced and displayed maturation arrest at the pro-B/large pre-B II stages of differentiation, similar to human E2A-PBX1 ALL. Somatic activation of E2A-PBX1 in B cell progenitors enhanced self-renewal and led to acquisition of multiple secondary genomic aberrations, including prominent spontaneous loss of Pax5. In preleukemic mice, conditional Pax5 deletion cooperated with E2A-PBX1 to expand progenitor B cell subpopulations, increasing penetrance and shortening leukemia latency. Recurrent secondary activating mutations were detected in key signaling pathways, most notably JAK/STAT, that leukemia cells require for proliferation. These data support conditional E2A-PBX1 mice as a model of human ALL and suggest targeting pre-BCR signaling and JAK kinases as potential therapeutic strategies.
View details for DOI 10.1172/JCI81158
View details for Web of Science ID 000362303600039
View details for PubMedID 26301816
MLL fusion proteins in leukemia induce aberrant transcriptional elongation and associated chromatin perturbations; however, the upstream signaling pathways and activators that recruit or retain MLL oncoproteins at initiated promoters are unknown. Through functional and comparative genomic studies, we identified an essential role for NF-κB signaling in MLL leukemia. Suppression of NF-κB led to robust antileukemia effects that phenocopied loss of functional MLL oncoprotein or associated epigenetic cofactors. The NF-κB subunit RELA occupies promoter regions of crucial MLL target genes and sustains the MLL-dependent leukemia stem cell program. IKK/NF-κB signaling is required for wild-type and fusion MLL protein retention and maintenance of associated histone modifications, providing a molecular rationale for enhanced efficacy in therapeutic targeting of this pathway in MLL leukemias.
View details for DOI 10.1016/j.ccr.2013.08.019
View details for Web of Science ID 000326198500006
View details for PubMedID 24054986
Small ubiquitin-like modifier (SUMO) conjugation and interaction are increasingly associated with various cellular processes. However, little is known about the cellular signaling mechanisms that regulate proteins for distinct SUMO paralog conjugation and interactions. Using the transcriptional coregulator Daxx as a model, we show that SUMO paralog-selective binding and conjugation are regulated by phosphorylation of the Daxx SUMO-interacting motif (SIM). NMR structural studies show that Daxx (732)E-I-I-V-L-S-D-S-D(740) is a bona fide SIM that binds to SUMO-1 in a parallel orientation. Daxx-SIM is phosphorylated by CK2 kinase at residues S737 and S739. Phosphorylation promotes Daxx-SIM binding affinity toward SUMO-1 over SUMO-2/3, causing Daxx preference for SUMO-1 conjugation and interaction with SUMO-1-modified factors. Furthermore, Daxx-SIM phosphorylation enhances Daxx to sensitize stress-induced cell apoptosis via antiapoptotic gene repression. Our findings provide structural insights into the Daxx-SIM:SUMO-1 complex, a model of SIM phosphorylation-enhanced SUMO paralog-selective modification and interaction, and phosphorylation-regulated Daxx function in apoptosis.
View details for DOI 10.1016/j.molcel.2011.02.022
View details for Web of Science ID 000289500500008
View details for PubMedID 21474068
Mitotic spindles play essential roles in chromosome congression and segregation during mitosis. Aurora A regulates spindle assembly in part via phosphorylating human TACC3 on S558, which triggers TACC3 relocalization to mitotic spindles and stabilizes microtubules (MTs). In this study, we identified clathrin heavy chain (CHC) as an adaptor protein to recruit S558-phosphorylated TACC3 onto the spindle during mitosis for MT stabilization. CHC binds phospho-S558 TACC3 via its linker domain and first CHC repeat. CHC depletion or mutation on phospho-TACC3 binding abrogates TACC3 spindle relocalization. Depletion of either or both CHC and TACC3 yields similar defective phenotypes: loss of ch-TOG on spindles, disorganized spindles, and chromosome misalignment with comparable mitotic delay. Our findings elucidate the association between aurora A phosphorylation and spindle apparatus and demonstrate that regulation from aurora A is mediated by CHC in recruiting phospho-TACC3 and subsequently ch-TOG to mitotic spindles.
View details for DOI 10.1083/jcb.200911120
View details for Web of Science ID 000279188400008
View details for PubMedID 20566684
Transforming acidic coiled-coil protein 3 (TACC3) was reported to be important for regulating mitotic spindle assembly and chromosome segregation. While the protein level of TACC3 was shown to be altered during cell cycle progression, the molecular mechanism in controlling TACC3 level is unclear. Here, we show that TACC3 protein level can be regulated by Cdh1, a well known activator of anaphase-promoting complex/cyclosome. We identified Cdh1 as an interacting partner of TACC3 by a yeast array screen. Both in vitro and in vivo binding studies indicated that TACC3 can form complexes with Cdh1. Depletion of endogenous Cdh1 prolonged TACC3 protein level during mitotic exit. Alteration of Cdh1 level by ectopic overexpression or siRNA knockdown correlated well with an increase or decrease of ubiquitinated TACC3, respectively. Furthermore, the domain mapping studies of TACC3 revealed that multiple domains are involved in Cdh1-regulated degradation of TACC3. Altogether, our findings suggest that Cdh1 controls TACC3 protein stability during mitotic exit.
View details for PubMedID 19823035
Bamboo is distinguished by its rapid growth. To investigate sucrose metabolism in this plant, we cloned the cDNAs encoding sucrose synthase (SuS) from Bambusa oldhamii and investigated their expression in growing shoots and leaves. Four cDNA clones, BoSus1, BoSus2, BoSus3 and BoSus4, were isolated by screening a cDNA library from etiolated bamboo shoots. Recombinant BoSuS proteins were produced in Escherichia coli and purified by immobilized metal affinity chromatography and ultrafiltration. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was used to determine the abundance of the transcript of each gene. BoSus1 and BoSus3 may be duplicate or homeologous genes, the sequences of which show high identity. Similarly, BoSus2 shows high identity with BoSus4. Kinetic analysis showed that the two BoSuS isoforms of each type had similar michaelis constant (Km) values for sucrose, but different values for UDP. The four genes were expressed in various bamboo organs but were differentially regulated. The increase in the abundance of their mRNA paralleled the growth rate of the bamboo. The results suggest that, in bamboo, SuS is encoded by at least four genes, each with a specific role in providing substrates for the polysaccharide biosynthesis and/or energy production necessary to support the rapid growth of this species.
View details for DOI 10.1111/j.1469-8137.2005.01638.x
View details for Web of Science ID 000235731300008
View details for PubMedID 16539603
ADP-ribosylation factors (ARFs) are ubiquitous regulators of virtually every step of vesicular membrane traffic. Yeast Arf3p, which is most similar to mammalian ARF6, is not essential for cell viability and not required for endoplasmic reticulum-to-Golgi protein transport. Although mammalian ARF6 has been implicated in the regulation of early endocytic transport, we found that Arf3p was not required for fluid-phase, membrane internalization, or mating-type receptor-mediated endocytosis. Arf3p was partially localized to the cell periphery, but was not detected on endocytic structures. The nucleotide-binding, N-terminal region, and N-terminal myristate of Arf3p are important for its proper localization. C-Terminally green fluorescent protein-tagged Arf3, expressed from the endogenous promoter, exhibited a polarized localization to the cell periphery and buds, in a cell cycle-dependent manner. Arf3-GFP achieved its proper localization during polarity growth through an actin-independent pathway. Both haploid and homologous diploid arf3 mutants exhibit a random budding defect, and the overexpression of the GTP-bound form Arf3p(Q71L) or GDP-binding defective Arf3p(T31N) mutant interfered with budding-site selection. We conclude that the GTPase cycle of Arf3p is likely to be important for the function of Arf3p in polarizing growth of the emerging bud and/or an unidentified vesicular trafficking pathway.
View details for DOI 10.1091/mbc.E03-01-0013
View details for Web of Science ID 000185501600027
View details for PubMedID 12972567