Grant Award Details
To support pioneers in the stem cell field who move their labs to California. Dr. Darzacq moved his group to UC Berkeley from France with this award. His group has focused its activity on three 3 goals: establishing new technologies for imaging molecular processes in differentiating tissues, studying the cellular reprograming of the pre-initiation complex concomitant to differentiation events during tissue regeneration and establishing a research program to study genome architecture and the proteins establishing it during differentiation in ES cells.
Grant Application Details
- Single Molecule Biophysics and Biology of Cellular Identity
One of your earliest childhood biology lessons probably occurred when your body demonstrated to you that your skin is an organ that is able to self-regenerate. Indeed wound healing is a fascinating process in which cells carry out a precise and complex choreography that includes cellular differentiation and regulation of gene expression.
Our lab studies a particular cell type called dermal fibroblasts. If a wound occurs, they migrate to the site of injury, change into muscle like cells (myofibroblasts) that contract to help with wound closure, and, once the wound has healed, enter programed cell death to clear the work area. Disruption to this process can result in chronic ulcers or keloid scarring. A major goal of our studies is to understand how the fibroblast to myofibroblast transition is regulated, so that therapeutic strategies can be devised to prevent and treat this pervasive problem.
In addition to our motivation to understand wound healing in order to learn how to control it and cure its pathologies, wound healing is an accessible system to study more general differentiation events involved in tissue regeneration. By studying the changes that fibroblasts undergo during wound healing, we revealed an important mechanism of gene regulation that could help to explain more generally how cells maintain a particular identity and how they can be driven to a different state. The molecules we identified are known to control general gene activity as well as the spatial organization of genes within the cell’s nucleus. The studies proposed here will further investigate those findings.
One of the ways in which our laboratory studies how cells control gene activity is by directly visualizing gene expression. Using highly specialized microscopy, biochemistry and computer analyses, we are able to observe the behavior of individual gene regulatory molecules within individual living cells. We will continue to use and improve these methods to better understand how genes are controlled. We believe that these studies will open the door to new strategies in cellular reprogramming and potentially to new strategies for modifying cells for therapeutic use.
Our research program includes clinically relevant and translational studies of wound healing, development and transfer of new imaging technologies to enable investigators to visualize the behavior of individual molecules within living cells, and basic biological studies seeking to understand fundamental mechanisms of gene regulation that control stem cell pluripotency and differentiation. These endeavors will benefit the State of California in various different and overlapping ways.
Successful development of stem cell-based therapies hinges on the ability to control cell identity and cell fate. By contributing to the understanding of fundamental gene regulatory mechanisms controlling pluripotency and tissue-specific differentiation, the research proposed here has the potential to influence and positively impact a wide range of regenerative medicine studies ongoing in the state.
Chronic skin ulcers are a pervasive and dangerous human medical problem. Current treatments can be slow and painful, and are too often ineffective, resulting in the necessity of limb amputation. The true economic and personal cost of chronic ulcers is difficult to quantify because public health research most frequently includes it as a symptom of systemic disease. However, in 2009 chronic ulcers were estimated to affect 6.5 million patients in the Unites States, with treatment costs in excess of 25 billion dollars. Our identification of specific factors that control the fibroblast to myofibroblast transition suggests new approaches to differential diagnosis and to treatment of chronic wounds using fibroblasts and myofibroblasts as direct targets or therapeutic agents.
The ability to detect, track and manipulate individual gene regulatory molecules in living human cells is a disruptive technology that already is having a major impact in developmental biology and stem cell research. Technologies for imaging individual molecules in live cells are evolving so rapidly that they are difficult to commercialize and are therefore available only to a relatively small number of laboratories that are able to build these systems from component parts. Our lab will serve as a center to help develop these technologies, and to help other laboratories adopt these powerful new research tools. California also has a rich industrial base in microscopy development and engineering. The development and implementation of new super resolution live cell imaging technologies such as we propose here will offer many opportunities for collaboration between industry and academia, opening new markets and enabling the spread of these innovations throughout academic and bio-pharmaceutical laboratories. This synergism, in turn, will accelerate research in regenerative therapeutics.
- Mol Cell (2020) Resolving the 3D Landscape of Transcription-Linked Mammalian Chromatin Folding. (PubMed: 32213323)
- Nat Chem Biol (2020) Guided nuclear exploration increases CTCF target search efficiency. (PubMed: 31792445)
- Genome Res (2019) CTCF sites display cell cycle-dependent dynamics in factor binding and nucleosome positioning. (PubMed: 30655336)
- Elife (2019) Determining cellular CTCF and cohesin abundances to constrain 3D genome models. (PubMed: 31205001)
- Mol Cell (2019) Distinct Classes of Chromatin Loops Revealed by Deletion of an RNA-Binding Region in CTCF. (PubMed: 31522987)
- Genes Dev (2019) Evaluating phase separation in live cells: diagnosis, caveats, and functional consequences. (PubMed: 31594803)
- Elife (2019) Evidence for DNA-mediated nuclear compartmentalization distinct from phase separation. (PubMed: 31038454)
- Science (2018) Imaging dynamic and selective low-complexity domain interactions that control gene transcription. (PubMed: 29930090)
- Elife (2018) A stable mode of bookmarking by TBP recruits RNA polymerase II to mitotic chromosomes. (PubMed: 29939130)
- Nat Struct Mol Biol (2018) RNA polymerase II clustering through carboxy-terminal domain phase separation. (PubMed: 30127355)
- Elife (2018) Dynamic multifactor hubs interact transiently with sites of active transcription in Drosophila embryos. (PubMed: 30589412)
- Biochem Soc Trans (2018) Protein motion in the nucleus: from anomalous diffusion to weak interactions. (PubMed: 30065106)
- Methods Mol Biol (2018) Single Molecule Imaging in Live Embryos Using Lattice Light-Sheet Microscopy. (PubMed: 29956254)
- Nucleus (2018) Recent evidence that TADs and chromatin loops are dynamic structures. (PubMed: 29077530)
- Elife (2018) Robust model-based analysis of single-particle tracking experiments with Spot-On. (PubMed: 29300163)
- Opt Express (2017) Faster and less phototoxic 3D fluorescence microscopy using a versatile compressed sensing scheme. (PubMed: 28788909)
- Genes Dev (2017) Dense Bicoid hubs accentuate binding along the morphogen gradient. (PubMed: 28982761)
- Mol Biol Cell (2017) Multi-scale tracking reveals scale-dependent chromatin dynamics after DNA damage. (PubMed: 28794266)
- Genes Dev (2017) A dynamic interplay of enhancer elements regulates Klf4 expression in naive pluripotency. (PubMed: 28982762)
- Nature (2017) Phase separation drives heterochromatin domain formation. (PubMed: 28636597)
- Elife (2017) CTCF and cohesin regulate chromatin loop stability with distinct dynamics. (PubMed: 28467304)
- Nat Commun (2017) Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9. (PubMed: 28290446)
- Nat Commun (2016) A single-molecule view of transcription reveals convoys of RNA polymerases and multi-scale bursting. (PubMed: 27461529)
- Biomed Opt Express (2016) Multifocus microscopy with precise color multi-phase diffractive optics applied in functional neuronal imaging. (PubMed: 27231594)
- Elife (2016) A dynamic mode of mitotic bookmarking by transcription factors. (PubMed: 27855781)