Chris Hsiung, M.D., Ph.D., is a physician-scientist developing methods to study the combinatorial functions of genes and genomic regulatory elements in specifying cell states.
- Coagulation laboratory
Residency: Stanford University Department of Pathology (2020) CA
Medical Education: University of Pennsylvania Registrar's Office (2017) PA
M.D., Perelman School of Medicine, University of Pennsylvania, Medicine (2017)
Ph.D., Perelman School of Medicine, University of Pennsylvania, Cell and Molecular Biology (2016)
B.A., University of California, Berkeley, Molecular and Cell Biology (2009)
A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition
GENES & DEVELOPMENT
2016; 30 (12): 1423–39
During mitosis, RNA polymerase II (Pol II) and many transcription factors dissociate from chromatin, and transcription ceases globally. Transcription is known to restart in bulk by telophase, but whether de novo transcription at the mitosis-G1 transition is in any way distinct from later in interphase remains unknown. We tracked Pol II occupancy genome-wide in mammalian cells progressing from mitosis through late G1. Unexpectedly, during the earliest rounds of transcription at the mitosis-G1 transition, ∼50% of active genes and distal enhancers exhibit a spike in transcription, exceeding levels observed later in G1 phase. Enhancer-promoter chromatin contacts are depleted during mitosis and restored rapidly upon G1 entry but do not spike. Of the chromatin-associated features examined, histone H3 Lys27 acetylation levels at individual loci in mitosis best predict the mitosis-G1 transcriptional spike. Single-molecule RNA imaging supports that the mitosis-G1 transcriptional spike can constitute the maximum transcriptional activity per DNA copy throughout the cell division cycle. The transcriptional spike occurs heterogeneously and propagates to cell-to-cell differences in mature mRNA expression. Our results raise the possibility that passage through the mitosis-G1 transition might predispose cells to diverge in gene expression states.
View details for DOI 10.1101/gad.280859.116
View details for Web of Science ID 000378533000006
View details for PubMedID 27340175
View details for PubMedCentralID PMC4926865
Genome accessibility is widely preserved and locally modulated during mitosis
2015; 25 (2): 213–25
Mitosis entails global alterations to chromosome structure and nuclear architecture, concomitant with transient silencing of transcription. How cells transmit transcriptional states through mitosis remains incompletely understood. While many nuclear factors dissociate from mitotic chromosomes, the observation that certain nuclear factors and chromatin features remain associated with individual loci during mitosis originated the hypothesis that such mitotically retained molecular signatures could provide transcriptional memory through mitosis. To understand the role of chromatin structure in mitotic memory, we performed the first genome-wide comparison of DNase I sensitivity of chromatin in mitosis and interphase, using a murine erythroblast model. Despite chromosome condensation during mitosis visible by microscopy, the landscape of chromatin accessibility at the macromolecular level is largely unaltered. However, mitotic chromatin accessibility is locally dynamic, with individual loci maintaining none, some, or all of their interphase accessibility. Mitotic reduction in accessibility occurs primarily within narrow, highly DNase hypersensitive sites that frequently coincide with transcription factor binding sites, whereas broader domains of moderate accessibility tend to be more stable. In mitosis, proximal promoters generally maintain their accessibility more strongly, whereas distal regulatory elements tend to lose accessibility. Large domains of DNA hypomethylation mark a subset of promoters that retain accessibility during mitosis and across many cell types in interphase. Erythroid transcription factor GATA1 exerts site-specific changes in interphase accessibility that are most pronounced at distal regulatory elements, but has little influence on mitotic accessibility. We conclude that features of open chromatin are remarkably stable through mitosis, but are modulated at the level of individual genes and regulatory elements.
View details for DOI 10.1101/gr.180646.114
View details for Web of Science ID 000348974500006
View details for PubMedID 25373146
View details for PubMedCentralID PMC4315295
Comparative analysis of mitosis-specific antibodies for bulk purification of mitotic populations by fluorescence-activated cell sorting
2014; 56 (2): 90-+
Mitosis entails complex chromatin changes that have garnered increasing interest from biologists who study genome structure and regulation-fields that are being advanced by high-throughput sequencing (Seq) technologies. The application of these technologies to study the mitotic genome requires large numbers of highly pure mitotic cells, with minimal contamination from interphase cells, to ensure accurate measurement of phenomena specific to mitosis. Here, we optimized a fluorescence-activated cell sorting (FACS)-based method for isolating formaldehyde-fixed mitotic cells--at virtually 100% mitotic purity and in quantities sufficient for high-throughput genomic studies. We compared several commercially available antibodies that react with mitosis-specific epitopes over a range of concentrations and cell numbers, finding antibody MPM2 to be the most robust and cost-effective.
View details for DOI 10.2144/000114137
View details for Web of Science ID 000331491800007
View details for PubMedID 24502799
View details for PubMedCentralID PMC4090248
Effects of sheared chromatin length on ChIP-seq quality and sensitivity.
G3 (Bethesda, Md.)
Chromatin immunoprecipitation followed by massively parallel, high throughput sequencing (ChIP-seq) is the method of choice for genome-wide identification of DNA segments bound by specific transcription factors or in chromatin with particular histone modifications. However, the quality of ChIP-seq datasets varies widely, with a substantial fraction being of intermediate to poor quality. Thus, it is important to discern and control the factors that contribute to variation in ChIP-seq. In this study, we focused on sonication, a user-controlled variable, to produce sheared chromatin. We systematically varied the amount of shearing of fixed chromatin from a mouse erythroid cell line, carefully measuring the distribution of resultant fragment lengths prior to ChIP-seq. This systematic study was complemented with a retrospective analysis of additional experiments. We found that the level of sonication had a pronounced impact on the quality of ChIP-seq signals. Over-sonication consistently reduced quality, while the impact of under-sonication differed among transcription factors, with no impact on sites bound by CTCF but frequently leading to the loss of sites occupied by TAL1 or bound by POL2. The bound sites not observed in low quality datasets were inferred to be a mix of both direct and indirect binding. We leveraged these findings to produce a set of CTCF ChIP-seq datasets in rare, primary hematopoietic progenitor cells. Our observation that the amount of chromatin sonication is a key variable in success of ChIP-seq experiments indicates that monitoring the level of sonication can improve ChIP-seq quality and reproducibility and facilitate ChIP-seq in rare cell types.
View details for DOI 10.1093/g3journal/jkab101
View details for PubMedID 33788948
Interrogating Histone Acetylation and BRD4 as Mitotic Bookmarks of Transcription.
2019; 27 (2): 400–415.e5
Global changes in chromatin organization and the cessation of transcription during mitosis are thought to challenge the resumption of appropriate transcription patterns after mitosis. The acetyl-lysine binding protein BRD4 has been previously suggested to function as a transcriptional "bookmark" on mitotic chromatin. Here, genome-wide location analysis of BRD4 in erythroid cells, combined with data normalization and peak characterization approaches, reveals that BRD4 widely occupies mitotic chromatin. However, removal of BRD4 from mitotic chromatin does not impair post-mitotic activation of transcription. Additionally, histone mass spectrometry reveals global preservation of most posttranslational modifications (PTMs) during mitosis. In particular, H3K14ac, H3K27ac, H3K122ac, and H4K16ac widely mark mitotic chromatin, especially at lineage-specific genes, and predict BRD4 mitotic binding genome wide. Therefore, BRD4 is likely not a mitotic bookmark but only a "passenger." Instead, mitotic histone acetylation patterns may constitute the actual bookmarks that restore lineage-specific transcription patterns after mitosis.
View details for PubMedID 30970245
A new bookmark of the mitotic genome in embryonic stem cells
NATURE CELL BIOLOGY
2016; 18 (11): 1124–25
Embryonic stem cells maintain pluripotency through countless mitoses. A recent report shows that the transcription factor Esrrb remains bound to chromatin during mitosis, including at regulatory regions that support pluripotency. Mitotic chromatin occupancy by Esrrb might stabilize the defining transcriptional programmes of embryonic stem cells through cell division.
View details for DOI 10.1038/ncb3432
View details for Web of Science ID 000387165600006
View details for PubMedID 27784905
View details for PubMedCentralID PMC5742480
Enhancer Regulation of Transcriptional Bursting Parameters Revealed by Forced Chromatin Looping
2016; 62 (2): 237–47
Mammalian genes transcribe RNA not continuously, but in bursts. Transcriptional output can be modulated by altering burst fraction or burst size, but how regulatory elements control bursting parameters remains unclear. Single-molecule RNA FISH experiments revealed that the β-globin enhancer (LCR) predominantly augments transcriptional burst fraction of the β-globin gene with modest stimulation of burst size. To specifically measure the impact of long-range chromatin contacts on transcriptional bursting, we forced an LCR-β-globin promoter chromatin loop. We observed that raising contact frequencies increases burst fraction but not burst size. In cells in which two developmentally distinct LCR-regulated globin genes are cotranscribed in cis, burst sizes of both genes are comparable. However, allelic co-transcription of both genes is statistically disfavored, suggesting mutually exclusive LCR-gene contacts. These results are consistent with competition between the β-type globin genes for LCR contacts and suggest that LCR-promoter loops are formed and released with rapid kinetics.
View details for DOI 10.1016/j.molcel.2016.03.007
View details for Web of Science ID 000374643900009
View details for PubMedID 27067601
View details for PubMedCentralID PMC4842148
Dynamic enhancer-gene body contacts during transcription elongation
GENES & DEVELOPMENT
2015; 29 (19): 1992–97
Enhancers govern transcription through multiple mechanisms, including the regulation of elongation by RNA polymerase II (RNAPII). We characterized the dynamics of looped enhancer contacts during synchronous transcription elongation. We found that many distal enhancers form stable contacts with their target promoters during the entire interval of elongation. Notably, we detected additional dynamic enhancer contacts throughout the gene bodies that track with elongating RNAPII and the leading edge of RNA synthesis. These results support a model in which the gene body changes its position relative to a stable enhancer-promoter complex, which has broad ramifications for enhancer function and architectural models of transcriptional elongation.
View details for DOI 10.1101/gad.255265.114
View details for Web of Science ID 000363002700002
View details for PubMedID 26443845
View details for PubMedCentralID PMC4604340
Native cysteine residues are dispensable for the structure and function of all five yeast mitotic septins
PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS
2013; 81 (11): 1964–79
Budding yeast septins assemble into hetero-octamers and filaments required for cytokinesis. Solvent-exposed cysteine (Cys) residues provide sites for attaching substituents useful in assessing assembly kinetics and protein interactions. To introduce Cys at defined locations, site-directed mutagenesis was used, first, to replace the native Cys residues in Cdc3 (C124 C253 C279), Cdc10 (C266), Cdc11 (C43 C137 C138), Cdc12 (C40 C278), and Shs1 (C29 C148) with Ala, Ser, Val, or Phe. When plasmid-expressed, each Cys-less septin mutant rescued the cytokinesis defects caused by absence of the corresponding chromosomal gene. When integrated and expressed from its endogenous promoter, the same mutants were fully functional, except Cys-less Cdc12 mutants (which were viable, but exhibited slow growth and aberrant morphology) and Cdc3(C124V C253V C279V) (which was inviable). No adverse phenotypes were observed when certain pairs of Cys-less septins were co-expressed as the sole source of these proteins. Cells grew less well when three Cys-less septins were co-expressed, suggesting some reduction in fitness. Nonetheless, cells chromosomally expressing Cys-less Cdc10, Cdc11, and Cdc12, and expressing Cys-less Cdc3 from a plasmid, grew well at 30°C. Moreover, recombinant Cys-less septins--or where one of the Cys-less septins contained a single Cys introduced at a new site--displayed assembly properties in vitro indistinguishable from wild-type.
View details for DOI 10.1002/prot.24345
View details for Web of Science ID 000325980300010
View details for PubMedID 23775754
View details for PubMedCentralID PMC3880206