We are an interdisciplinary team using the tools of biophysics, biochemistry, genetics, molecular and structural biology to elucidate the structure and function of chromatin, the native state of the genome in association with proteins and RNAs. This ‘epigenome’ carries the blueprint of gene expression programs directing cell growth, homeostasis and differentiation throughout plant and animal life. Our basic studies are highly relevant to medicine as genome mutations and dysfunctions of the epigenome underlie many human diseases.
We are most interested in the role of nucleosome organization and dynamics in controlling gene transcription. Genomes are packaged as arrays of nucleosomes, fundamental histone-DNA particles that are inaccessible to RNA polymerase enzymes that transcribe DNA into RNA, the first step of information flow from genes to proteins. Initiation of transcription requires alterations in nucleosome and chromatin architecture at promoter and enhancer regulatory DNAs, allowing RNA polymerase recruitment and activity at transcription start sites. We seek to elucidate molecular mechanisms underlying this basic process. How do sequence-specific transcription factors operate together with ATP-dependent chromatin remodeling and histone modification enzymes to control transcription on chromatin? We seek answers at the single-molecule level where individual proteins are visualized in real time to decipher their functional behaviors.
What is the spatiotemporal distribution of epigenetic regulators within the nucleus? What are the timescales by which factors navigate the nuclear environment and target genomic sites in a background of nonspecific interactions? To address these questions, we systematically apply super-resolution fluorescence microscopy to image and track key chromatin proteins in living cells at the single-molecule level. We quantify diffusion constants, chromatin-bound and free fractions, search and residence times, factor clustering, and frequencies of site occupancy, using these values to model the transcription reaction on chromatin. Causal relationships between chromatin organization and transcription factor dynamics are examined by live imaging of conditional mutants.
Single molecules of histone variant H2A.Z imaged in live yeast cells. The endogenous H2A.Z was tagged with Halo and molecules were fluorescently labelled with Janelia Fluor 552. Time is indicated in seconds and rough outline of nuclear regions are marked in yellow.
Single molecules of Swr1 (the catalytic subunit of the SWR1 complex) imaged in live yeast cells. The endogenous Swr1 was tagged with Halo and molecules were fluorescently labelled with Janelia Fluor 646. Time is indicated in seconds and rough outline of nuclear regions are marked in yellow.
Multi-component ATP-dependent enzymes of the ISWI family mobilize nucleosomal DNA to reposition nucleosome core histones, thereby exposing or occluding DNA elements. A distinctive form of nucleosome remodeling by the related 14-component SWR1 complex evicts nucleosomal H2A histone, replacing it with the histone H2A.Z variant at gene promoters without changing nucleosome position. We are currently studying the mechanism of SWR1-catalyzed histone exchange. In collaboration with Prof. Taekjip Ha’s laboratory, we use single-molecule fluorescence resonance energy transfer (smFRET) to characterize dynamic remodeling intermediates and measure their lifetimes in vitro. The molecular architecture of the SWR1 complex is being investigated by cryo-electron microscopy. Using smFRET, we also investigate mechanisms by which a pioneer transcription factor cooperates with chromatin remodelers to create a hypersensitive site on chromatin.
Studying Chromatin Remodelers with Single-Molecule FRET
Our lab has used single-molecule FRET to observe chromatin remodelers in action. By placing fluorophores strategically on nucleosomal DNA and histone proteins we can directly observe chromatin remodelers acting on nucleosomes in real time. Shown on the right is a nucleosome construct labeled with Cy3 on DNA, Cy5 on H2A and Cy7 on H2A.Z that was created enzymatically using the chromatin remodeler SWR1. We image single nucleosomes, shown as white dots in the top image, using sensitive fluorescence microscopes. By measuring the FRET efficiency between all the fluorophores in the image, as shown in the bottom plot, we can identify the location of nucleosomal DNA and histone proteins to nanometer precision. This powerful assay allowed us to look at how DNA and histones are manipulated by chromatin remodelers in real time and identify new reaction intermediates that help us understand how nucleosome remodelers work.
How does condensation of nucleosome arrays into distinct 3D configurations influence transcription initiation and elongation? On artificial sequences, reconstituted nucleosome arrays have been proposed to compact into specific higher order conformations, but the supra-nucleosome architecture of a natural gene has been elusive. We plan to isolate individual genes in the native state from chromatin to visualize their state of compaction by electron microscopy. To gain mechanistic insights on the multi-step transcription process under physiological conditions in vitro, we will study transcription reaction intermediates at the single-molecule level by TIRF microscopy.
Undergraduate students participate in our research through a spring semester intensive laboratory course on single-molecule live imaging of chromatin factors. AS.020.361: Advanced Research Lab in Cell and Molecular Biology.
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Principal Scientist
Associate Scientist
Postdoctoral Fellow Research Scientist
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Graduate Student (shared with TJ Ha Lab)
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Graduate Student (MD-PhD) (shared with Alexis Battle lab)
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2023
Welcome Sergei Rudnizky (postdoc, shared Taekjip Ha lab from 2022) and Sonia Weiner (undergraduate student).
Welcome summer students Kemi Abiodun, Kelly Chen, Annie Huang, Sean Kao, Chloe Liang, Kayra Ozturan, Sonia Weiner.
Congratulations to Dr. Ashlee Xinyu Feng, graduate of the CMDB program and recipient of the Biology Department Roseman Award for outstanding research in biochemistry!
Congratulations to Xiaona and Taibo for the collaborative paper with Mike Levine’s lab on GAF-dependent chromatin looping!
Our lab receives a 5-year MIRA grant award from NIGMS to investigate “Kinetic Mechanisms of Chromatin Remodeling and Transcription.” Congratulations to everyone who made it possible. Awesome effort.
Welcome Muhammad Bennani, Maggie Wang, Sophie Yao, Chloe Liang to the Spring Semester Advanced Cell Biology Lab Course in Single-Molecule Imaging.
Alice Chen joins the lab as a graduate student from the BCMB program. Welcome Alice!
2022
Congratulations to Dr. Jee Min Kim, graduate of the CMDB program and recipient of the Biology Department Roseman Award for outstanding research in biochemistry! All the best for a successful postdoc at NIH!
Congratulations to Xiaona Tang et al. on the publication of the NSMB paper on live imaging of GAGA factor! Great piece of work.
Big congratulations to Assistant Professor Vu Nguyen, new faculty member at University of California, San Diego. We will miss you!
Giho Park returns to the lab as an MD-PhD student, to continue CryoEM with Robert. Welcome back Giho!
Maryam Yamadi joins the lab as a new graduate student. Welcome Maryam!
August, 2021
Congratulation to Vu and Jee Min for their respective publications in Molecular Cell and eLife, and the collaborative venture with Francois Robert’s lab, also out in Molecular Cell.
July, 2021
Zelin Wei joins the lab as our new graduate student. Welcome!
June, 2021
Farewell to Ejlal and Giho, our gap year students on their way to MD-PhDs
December, 2020
Vu Nguyen submits his magnum opus to bioRxiv “Spatio-Temporal Coordination of Transcription Preinitiation Complex Assembly in Live Cells.” Way to go, Vu!
November, 2020
We are most pleased to be part of the NIH 4D nucleome program. Thanks to Xiaona Tang, lab lead for the funded Ha-Johnson-Wu consortium U01 proposal “Chromatin Function During Transcription and DNA Repair at Single-Molecule Resolution in Living Cells.”
August, 2020
Congratulations to Taibo for passing his oral exams!
July, 2020
We have a lab website on-line! Thanks to Jenna Louder.
A warm welcome to Nathan Jones, Ph.D. Biophysics, joining us from Ohio State!
June, 2020
Phase I reopening after a 3-month shut-down, and we are relieved to get back to work, cautiously.
May, 2020
Congratulations to Sun-Jay on his graduation from the Computer Science program, and many thanks for contributions to Sojourner, our single-molecule computation pipeline!
Congratulations to Eve (Ejlal) on her graduation from the MCB program and continuing as research technologist for her gap year!
Taibo Li, MD. Ph.D. student joins the lab, co-mentored with Prof. Alexis Battle, Dept. of Biomedical Engineering. Welcome!
Congratulations to Anand and the team for their eLIFE publication “Live-cell single particle imaging reveals the role of RNA polymerase II in histone H2A.Z eviction.” A milestone for the lab’s entry into single-molecule imaging.
April 2020
Congratulations to Hin for his Croucher Foundation post-doctoral fellowship award!
March, 2020
The lab and Hopkins are shut down due to COVID19.
December, 2019
Welcome Hin Ling, PhD. Biochemistry, to the lab. From Hong Kong to snowy Baltimore!
July 2019
Congratulations to Robert for his NRSA post-doctoral fellowship award!
June, 2019
Giho Park graduates from the MCB program and re-joins the lab as research technologist.
June, 2019
Congratulations to Vivian, Pat, and Kai-Yu our undergrad researchers on their graduation from the MCB program at Hopkins!
January and April, 2019
Sheng and Qin, respectively, return to Canada, on the way to new positions in China. Many thanks for essential roles in setting up our Hopkins operation and our very best wishes!
July 2018
Congratulations to Matt for his NRSA post-doctoral fellowship award!
June, 2018
Ashlee Feng joins the lab as our second graduate student from CMDB, jointly mentored by TJ Ha.
March, 2018
Robert Louder, Ph.D. structural biology, arrives to his native Maryland from sunny Berkeley.
June, 2017
Xiaona Tang, Ph.D. Molecular Genetics, moves with Drosophila strains from our previous NIH lab to continue her postdoc at Hopkins.
April, 2017
Matt Poyton, Ph.D. Chemistry, joins the lab, a joint postdoc with TJ Ha. Welcome!
June, 2017
Jee-Min Kim joins the lab as our very first graduate student, from the CMDB program!
January, 2017
Gaku, Anand, and Vu from HHMI-Janelia join Carl at Homewood.
September, 2016
Sheng Liu, Ph.D. Molecular Genetics, and Qin Liu, B.Sc. join the lab from Vancouver, Canada.
July, 2016
Carl moves from HHMI Janelia’s Transcription Imaging Consortium to start the new lab at the Hopkins Homewood campus, Baltimore.
Johns Hopkins University
Krieger School of Arts & Sciences
Department of Biology
