@misc{polak_keung_2023, title={A molecular assessment of the practical potential of DNA-based computation}, volume={81}, ISSN={["1879-0429"]}, DOI={10.1016/j.copbio.2023.102940}, abstractNote={The immense information density of DNA and its potential for massively parallelized computations, paired with rapidly expanding data production and storage needs, have fueled a renewed interest in DNA-based computation. Since the construction of the first DNA computing systems in the 1990s, the field has grown to encompass a diverse array of configurations. Simple enzymatic and hybridization reactions to solve small combinatorial problems transitioned to synthetic circuits mimicking gene regulatory networks and DNA-only logic circuits based on strand displacement cascades. These have formed the foundations of neural networks and diagnostic tools that aim to bring molecular computation to practical scales and applications. Considering these great leaps in system complexity as well as in the tools and technologies enabling them, a reassessment of the potential of such DNA computing systems is warranted.}, journal={CURRENT OPINION IN BIOTECHNOLOGY}, author={Polak, Rachel E. and Keung, Albert J.}, year={2023}, month={Jun} }
 @article{han_lee_indermaur_keung_2023, title={Chaetocin disrupts the SUV39H1-HP1 interaction independent of SUV39H1 methyltransferase activity}, volume={480}, ISSN={["1470-8728"]}, DOI={10.1042/BCJ20220528}, abstractNote={Chemical tools to control the activities and interactions of chromatin components have broad impact on our understanding of cellular and disease processes. It is important to accurately identify their molecular effects to inform clinical efforts and interpretations of scientific studies. Chaetocin is a widely used chemical that decreases H3K9 methylation in cells. It is frequently attributed as a specific inhibitor of the histone methyltransferase activities of SUV39H1/SU(VAR)3–9, although prior observations showed chaetocin likely inhibits methyltransferase activity through covalent mechanisms involving its epipolythiodixopiperazine disulfide ‘warhead’ functionality. The continued use of chaetocin in scientific studies may derive from the net effect of reduced H3K9 methylation, irrespective of a direct or indirect mechanism. However, there may be other molecular impacts of chaetocin on SUV39H1 besides inhibition of H3K9 methylation levels that could confound the interpretation of past and future experimental studies. Here, we test a new hypothesis that chaetocin may have an additional downstream impact aside from inhibition of methyltransferase activity. Using a combination of truncation mutants, a yeast two-hybrid system, and direct in vitro binding assays, we show that the human SUV39H1 chromodomain (CD) and HP1 chromoshadow domain (CSD) directly interact. Chaetocin inhibits this binding interaction through its disulfide functionality with some specificity by covalently binding with the CD of SUV39H1, whereas the histone H3–HP1 interaction is not inhibited. Given the key role of HP1 dimers in driving a feedback cascade to recruit SUV39H1 and to establish and stabilize constitutive heterochromatin, this additional molecular consequence of chaetocin should be broadly considered.}, number={6}, journal={BIOCHEMICAL JOURNAL}, author={Han, Linna and Lee, Jessica B. and Indermaur, Elaine W. and Keung, Albert J.}, year={2023}, month={Mar}, pages={421–432} }
 @article{volkel_lin_hook_timp_keung_tuck_2023, title={FrameD: framework for DNA-based data storage design, verification, and validation}, volume={39}, ISSN={["1367-4811"]}, url={https://doi.org/10.1093/bioinformatics/btad572}, DOI={10.1093/bioinformatics/btad572}, abstractNote={Abstract}, number={10}, journal={BIOINFORMATICS}, author={Volkel, Kevin D. and Lin, Kevin N. and Hook, Paul W. and Timp, Winston and Keung, Albert J. and Tuck, James M.}, editor={Kelso, JanetEditor}, year={2023}, month={Oct} }
 @article{tam_keung_2023, title={Profiling transcriptomic responses of human stem cell-derived medium spiny neuron-like cells to exogenous phasic and tonic neurotransmitters}, volume={126}, ISSN={["1095-9327"]}, DOI={10.1016/j.mcn.2023.103876}, abstractNote={Transcriptomic responses to neurotransmitters contribute to the complex processes driving memory and addiction. Advances in both measurement methods and experimental models continue to improve our understanding of this regulatory layer. Here we focus on the experimental potential of stem cell derived neurons, currently the only ethical model that can be used in reductionist and experimentally perturbable studies of human cells. Prior work has focused on generating distinct cell types from human stem cells, and has also shown their utility in modeling development and cellular phenotypes related to neurodegeneration. Here we seek an understanding of how stem cell derived neural cultures respond to perturbations experienced during development and disease progression. This work profiles transcriptomic responses of human medium spiny neuron-like cells with three specific goals. We first characterize transcriptomic responses to dopamine and dopamine receptor agonists and antagonists presented in dosing patterns mimicking acute, chronic, and withdrawal regimens. We also assess transcriptomic responses to low and persistent tonic levels of dopamine, acetylcholine, and glutamate to better mimic the in vivo environment. Finally, we identify similar and distinct responses between hMSN-like cells derived from H9 and H1 stem cell lines, providing some context for the extent of variability these types of systems will likely pose for experimentalists. The results here suggest future optimizations of human stem cell derived neurons to increase their in vivo relevance and the biological insights that can be garnered from these models.}, journal={MOLECULAR AND CELLULAR NEUROSCIENCE}, author={Tam, Ryan W. and Keung, Albert J.}, year={2023}, month={Sep} }
 @article{volkel_tomek_keung_tuck_2022, title={DINOS: Data INspired Oligo Synthesis for DNA Data Storage}, volume={18}, ISSN={["1550-4840"]}, url={http://dx.doi.org/10.1145/3510853}, DOI={10.1145/3510853}, abstractNote={As interest in DNA-based information storage grows, the costs of synthesis have been identified as a key bottleneck. A potential direction is to tune synthesis for data. Data strands tend to be composed of a small set of recurring code word sequences, and they contain longer sequences of repeated data. To exploit these properties, we propose a new framework called DINOS. DINOS consists of three key parts: (i) The first is a hierarchical strand assembly algorithm, inspired by gene assembly techniques that can assemble arbitrary data strands from a small set of primitive blocks. (ii) The assembly algorithm relies on our novel formulation for how to construct primitive blocks, spanning a variety of useful configurations from a set of code words and overhangs. Each primitive block is a code word flanked by a pair of overhangs that are created by a cyclic pairing process that keeps the number of primitive blocks small. Using these primitive blocks, any data strand of arbitrary length can be assembled, theoretically. We show a minimal system for a binary code with as few as six primitive blocks, and we generalize our processes to support an arbitrary set of overhangs and code words. (iii) We exploit our hierarchical assembly approach to identify redundant sequences and coalesce the reactions that create them to make assembly more efficient.}, number={3}, journal={ACM JOURNAL ON EMERGING TECHNOLOGIES IN COMPUTING SYSTEMS}, publisher={Association for Computing Machinery (ACM)}, author={Volkel, Kevin and Tomek, Kyle J. and Keung, Albert J. and Tuck, James M.}, year={2022}, month={Jul} }
 @article{tam_keung_2022, title={Human Pluripotent Stem Cell-Derived Medium Spiny Neuron-like Cells Exhibit Gene Desensitization}, volume={11}, ISSN={["2073-4409"]}, DOI={10.3390/cells11091411}, abstractNote={Gene desensitization in response to a repeated stimulus is a complex phenotype important across homeostatic and disease processes, including addiction, learning, and memory. These complex phenotypes are being characterized and connected to important physiologically relevant functions in rodent systems but are difficult to capture in human models where even acute responses to important neurotransmitters are understudied. Here through transcriptomic analysis, we map the dynamic responses of human stem cell-derived medium spiny neuron-like cells (hMSN-like cells) to dopamine. Furthermore, we show that these human neurons can reflect and capture cellular desensitization to chronic versus acute administration of dopamine. These human cells are further able to capture complex receptor crosstalk in response to the pharmacological perturbations of distinct dopamine receptor subtypes. This study demonstrates the potential utility and remaining challenges of using human stem cell-derived neurons to capture and study the complex dynamic mechanisms of the brain.}, number={9}, journal={CELLS}, author={Tam, Ryan W. and Keung, Albert J.}, year={2022}, month={May} }
 @article{meanor_keung_rao_2022, title={Modified Histone Peptides Linked to Magnetic Beads Reduce Binding Specificity}, volume={23}, ISSN={["1422-0067"]}, DOI={10.3390/ijms23031691}, abstractNote={Histone post-translational modifications are small chemical changes to the histone protein structure that have cascading effects on diverse cellular functions. Detecting histone modifications and characterizing their binding partners are critical steps in understanding chromatin biochemistry and have been accessed using common reagents such as antibodies, recombinant assays, and FRET-based systems. High-throughput platforms could accelerate work in this field, and also could be used to engineer de novo histone affinity reagents; yet, published studies on their use with histones have been noticeably sparse. Here, we describe specific experimental conditions that affect binding specificities of post-translationally modified histones in classic protein engineering platforms and likely explain the relative difficulty with histone targets in these platforms. We also show that manipulating avidity of binding interactions may improve specificity of binding.}, number={3}, journal={INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES}, author={Meanor, Jenna N. and Keung, Albert J. and Rao, Balaji M.}, year={2022}, month={Feb} }
 @misc{matange_tuck_keung_2021, title={DNA stability: a central design consideration for DNA data storage systems}, volume={12}, ISSN={["2041-1723"]}, url={https://doi.org/10.1038/s41467-021-21587-5}, DOI={10.1038/s41467-021-21587-5}, abstractNote={Abstract}, number={1}, journal={NATURE COMMUNICATIONS}, author={Matange, Karishma and Tuck, James M. and Keung, Albert J.}, year={2021}, month={Mar} }
 @article{sen_voulgaropoulos_keung_2021, title={Effects of early geometric confinement on the transcriptomic profile of human cerebral organoids}, volume={21}, ISSN={["1472-6750"]}, DOI={10.1186/s12896-021-00718-2}, abstractNote={Abstract}, number={1}, journal={BMC BIOTECHNOLOGY}, author={Sen, Dilara and Voulgaropoulos, Alexis and Keung, Albert J.}, year={2021}, month={Oct} }
 @article{sen_drobna_keung_2021, title={Evaluation of UBE3A antibodies in mice and human cerebral organoids}, volume={11}, ISSN={["2045-2322"]}, DOI={10.1038/s41598-021-85923-x}, abstractNote={Abstract}, number={1}, journal={SCIENTIFIC REPORTS}, author={Sen, Dilara and Drobna, Zuzana and Keung, Albert J.}, year={2021}, month={Mar} }
 @article{lee_caywood_lo_levering_keung_2021, title={Mapping the dynamic transfer functions of eukaryotic gene regulation}, volume={12}, ISSN={["2405-4720"]}, DOI={10.1016/j.cels.2021.08.003}, abstractNote={Biological information can be encoded within the dynamics of signaling components, which has been implicated in a broad range of physiological processes including stress response, oncogenesis, and stem cell differentiation. To study the complexity of information transfer across the eukaryotic promoter, we screened 119 dynamic conditions-modulating the pulse frequency, amplitude, and pulse width of light-regulating the binding of an epigenome editor to a fluorescent reporter. This system revealed tunable gene expression and filtering behaviors and provided a quantification of the limit to the amount of information that can be reliably transferred across a single promoter as ∼1.7 bits. Using a library of over 100 orthogonal chromatin regulators, we further determined that chromatin state could be used to tune mutual information and expression levels, as well as completely alter the input-output transfer function of the promoter. This system unlocks the information-rich content of eukaryotic gene regulation.}, number={11}, journal={CELL SYSTEMS}, author={Lee, Jessica B. and Caywood, Leandra M. and Lo, Jennifer Y. and Levering, Nicholas and Keung, Albert J.}, year={2021}, month={Nov}, pages={1079-+} }
 @article{waldman_rao_keung_2021, title={Mapping the residue specificities of epigenome enzymes by yeast surface display}, volume={28}, ISSN={["2451-9448"]}, DOI={10.1016/j.chembiol.2021.05.022}, abstractNote={Histone proteins are decorated with a combinatorially and numerically diverse set of biochemical modifications. Here, we describe a versatile and scalable approach which enables efficient characterization of histone modifications without the need for recombinant protein production. As proof-of-concept, we first use this system to rapidly profile the histone H3 and H4 residue writing specificities of the human histone acetyltransferase, p300. Subsequently, a large panel of commercially available anti-acetylation antibodies are screened for their specificities, identifying many suitable and unsuitable reagents. Furthermore, this approach enables efficient mapping of the large binary crosstalk space between acetylated residues on histones H3 and H4 and uncovers residue interdependencies affecting p300 activity. These results show that using yeast surface display to study histone modifications is a useful tool that can advance our understanding of chromatin biology by enabling efficient interrogation of the complexity of epigenome modifications.}, number={12}, journal={CELL CHEMICAL BIOLOGY}, author={Waldman, Alison C. and Rao, Balaji M. and Keung, Albert J.}, year={2021}, month={Dec}, pages={1772-+} }
 @article{tomek_volkel_indermaur_tuck_keung_2021, title={Promiscuous molecules for smarter file operations in DNA-based data storage}, volume={12}, ISSN={["2041-1723"]}, url={https://doi.org/10.1038/s41467-021-23669-w}, DOI={10.1038/s41467-021-23669-w}, abstractNote={Abstract}, number={1}, journal={NATURE COMMUNICATIONS}, author={Tomek, Kyle J. and Volkel, Kevin and Indermaur, Elaine W. and Tuck, James M. and Keung, Albert J.}, year={2021}, month={Jun} }
 @article{annabi_baker_boettiger_chakraborty_chen_corbett_correia_dahlman_oliveira_ertuerk_et al._2021, title={Voices of biotech research}, volume={39}, ISSN={["1546-1696"]}, DOI={10.1038/s41587-021-00847-1}, abstractNote={Nature Biotechnology asks a selection of faculty about the most exciting frontier in their field and the most needed technologies for advancing knowledge and applications.}, number={3}, journal={NATURE BIOTECHNOLOGY}, author={Annabi, Nasim and Baker, Matthew and Boettiger, Alistair and Chakraborty, Debojyoti and Chen, Yvonne and Corbett, Kizzmekia S. and Correia, Bruno and Dahlman, James and Oliveira, Tulio and Ertuerk, Ali and et al.}, year={2021}, month={Mar}, pages={281–286} }
 @misc{sen_keung_2020, title={Capturing complex epigenetic phenomena through human multicellular systems}, volume={16}, ISSN={["2468-4511"]}, DOI={10.1016/j.cobme.2020.05.008}, abstractNote={Epigenetic states inherently define a wide range of complex biological phenotypes and processes in development and disease. Accurate cellular modeling would ideally capture the epigenetic complexity of these processes as well as the accompanying molecular changes in chromatin biochemistry including in DNA and histone modifications. Here we highlight recent work that demonstrate how multicellular systems provide a natural approach to capture complex epigenetic phenomena. They accomplish this through more closely matching the in vivo environment and through the intrinsic nature of multicellular systems being able to generate and model multiple distinct cellular states, all within one system. We also discuss challenges and limitations of such systems, efforts to tune and modulate epigenetics directly in multicellular systems, and how molecular interventional approaches could advance and improve the utility of these models.}, journal={CURRENT OPINION IN BIOMEDICAL ENGINEERING}, author={Sen, Dilara and Keung, Albert J.}, year={2020}, month={Dec}, pages={34–41} }
 @article{lin_volkel_tuck_keung_2020, title={Dynamic and scalable DNA-based information storage}, volume={11}, ISSN={["2041-1723"]}, url={https://doi.org/10.1038/s41467-020-16797-2}, DOI={10.1038/s41467-020-16797-2}, abstractNote={Abstract}, number={1}, journal={NATURE COMMUNICATIONS}, publisher={Springer Science and Business Media LLC}, author={Lin, Kevin N. and Volkel, Kevin and Tuck, James M. and Keung, Albert J.}, year={2020}, month={Jun} }
 @article{sen_voulgaropoulos_drobna_keung_2020, title={Human Cerebral Organoids Reveal Early Spatiotemporal Dynamics and Pharmacological Responses of UBE3A}, volume={15}, ISSN={["2213-6711"]}, DOI={10.1016/j.stemcr.2020.08.006}, abstractNote={Angelman syndrome is a complex neurodevelopmental disorder characterized by delayed development, intellectual disability, speech impairment, and ataxia. It results from the loss of UBE3A protein, an E3 ubiquitin ligase, in neurons of the brain. Despite the dynamic spatiotemporal expression of UBE3A observed in rodents and the potential clinical importance of when and where it is expressed, its expression pattern in humans remains unknown. This reflects a common challenge of studying human neurodevelopment: prenatal periods are hard to access experimentally. In this work, human cerebral organoids reveal a change from weak to strong UBE3A in neuronal nuclei within 3 weeks of culture. Angelman syndrome human induced pluripotent stem cell-derived organoids also exhibit early silencing of paternal UBE3A, with topoisomerase inhibitors partially rescuing UBE3A levels and calcium transient phenotypes. This work establishes human cerebral organoids as an important model for studying UBE3A and motivates their broader use in understanding complex neurodevelopmental disorders.}, number={4}, journal={STEM CELL REPORTS}, author={Sen, Dilara and Voulgaropoulos, Alexis and Drobna, Zuzana and Keung, Albert J.}, year={2020}, month={Oct}, pages={845–854} }
 @article{tomek_volkel_simpson_hass_indermaur_tuck_keung_2019, title={Driving the Scalability of DNA-Based Information Storage Systems}, volume={8}, ISSN={["2161-5063"]}, DOI={10.1021/acssynbio.9b00100}, abstractNote={The extreme density of DNA presents a compelling advantage over current storage media; however, to reach practical capacities, new systems for organizing and accessing information are needed. Here, we use chemical handles to selectively extract unique files from a complex database of DNA mimicking 5 TB of data and design and implement a nested file address system that increases the theoretical maximum capacity of DNA storage systems by five orders of magnitude. These advancements enable the development and future scaling of DNA-based data storage systems with modern capacities and file access capabilities.}, number={6}, journal={ACS SYNTHETIC BIOLOGY}, author={Tomek, Kyle J. and Volkel, Kevin and Simpson, Alexander and Hass, Austin G. and Indermaur, Elaine W. and Tuck, James M. and Keung, Albert J.}, year={2019}, month={Jun}, pages={1241–1248} }
 @article{park_patel_keung_khalil_2019, title={Engineering Epigenetic Regulation Using Synthetic Read-Write Modules}, volume={176}, ISSN={["1097-4172"]}, DOI={10.1016/j.cell.2018.11.002}, abstractNote={Chemical modifications to DNA and histone proteins are involved in epigenetic programs underlying cellular differentiation and development. Regulatory networks involving molecular writers and readers of chromatin marks are thought to control these programs. Guided by this common principle, we established an orthogonal epigenetic regulatory system in mammalian cells using N6-methyladenine (m6A), a DNA modification not commonly found in metazoan epigenomes. Our system utilizes synthetic factors that write and read m6A and consequently recruit transcriptional regulators to control reporter loci. Inspired by models of chromatin spreading and epigenetic inheritance, we used our system and mathematical models to construct regulatory circuits that induce m6A-dependent transcriptional states, promote their spatial propagation, and maintain epigenetic memory of the states. These minimal circuits were able to program epigenetic functions de novo, conceptually validating "read-write" architectures. This work provides a toolkit for investigating models of epigenetic regulation and encoding additional layers of epigenetic information in cells.}, number={1-2}, journal={CELL}, author={Park, Minhee and Patel, Nikit and Keung, Albert J. and Khalil, Ahmad S.}, year={2019}, month={Jan}, pages={227-+} }
 @article{liao_ttofali_slotkowski_denny_cecil_leenay_keung_beisel_2019, title={Modular one-pot assembly of CRISPR arrays enables library generation and reveals factors influencing crRNA biogenesis}, volume={10}, ISSN={["2041-1723"]}, DOI={10.1038/s41467-019-10747-3}, abstractNote={Abstract}, journal={NATURE COMMUNICATIONS}, author={Liao, Chunyu and Ttofali, Fani and Slotkowski, Rebecca A. and Denny, Steven R. and Cecil, Taylor D. and Leenay, Ryan T. and Keung, Albert J. and Beisel, Chase L.}, year={2019}, month={Jul} }
 @article{keung_khalil_2016, title={A unifying model of epigenetic regulation}, volume={351}, ISSN={["1095-9203"]}, DOI={10.1126/science.aaf1647}, abstractNote={
            Single-cell tracking reveals a common “algorithm” of operation used by chromatin regulators
            
              [Also see Report by
              
                Bintu
                et al.
              
              ]
            
          }, number={6274}, journal={SCIENCE}, author={Keung, Albert J. and Khalil, Ahmad S.}, year={2016}, month={Feb}, pages={661–662} }
 @misc{park_keung_khalil_2016, title={The epigenome: the next substrate for engineering}, volume={17}, ISSN={["1474-760X"]}, DOI={10.1186/s13059-016-1046-5}, abstractNote={We are entering an era of epigenome engineering. The precision manipulation of chromatin and epigenetic modifications provides new ways to interrogate their influence on genome and cell function and to harness these changes for applications. We review the design and state of epigenome editing tools, highlighting the unique regulatory properties afforded by these systems.}, journal={GENOME BIOLOGY}, author={Park, Minhee and Keung, Albert J. and Khalil, Ahmad S.}, year={2016}, month={Aug} }
 @article{keung_joung_khalil_collins_2015, title={Chromatin regulation at the frontier of synthetic biology}, volume={16}, ISSN={1471-0056 1471-0064}, url={http://dx.doi.org/10.1038/NRG3900}, DOI={10.1038/NRG3900}, abstractNote={Synthetic biology approaches to characterize gene regulation have largely used transcription factor circuits in bacteria. However, the multilayered regulation of genes by chromatin in eukaryotes provides opportunities for more sophisticated control of gene expression. This Review describes diverse approaches for engineering eukaryotic chromatin states, the insights gained into physiological gene regulation principles, and the broad potential applications throughout biomedical research and industry. As synthetic biology approaches are extended to diverse applications throughout medicine, biotechnology and basic biological research, there is an increasing need to engineer yeast, plant and mammalian cells. Eukaryotic genomes are regulated by the diverse biochemical and biophysical states of chromatin, which brings distinct challenges, as well as opportunities, over applications in bacteria. Recent synthetic approaches, including 'epigenome editing', have allowed the direct and functional dissection of many aspects of physiological chromatin regulation. These studies lay the foundation for biomedical and biotechnological engineering applications that could take advantage of the unique combinatorial and spatiotemporal layers of chromatin regulation to create synthetic systems of unprecedented sophistication.}, number={3}, journal={Nature Reviews Genetics}, publisher={Springer Science and Business Media LLC}, author={Keung, Albert J. and Joung, J. Keith and Khalil, Ahmad S. and Collins, James J.}, year={2015}, month={Feb}, pages={159–171} }
 @article{keung_bashor_kiriakov_collins_khalil_2014, title={Using Targeted Chromatin Regulators to Engineer Combinatorial and Spatial Transcriptional Regulation}, volume={158}, ISSN={0092-8674}, url={http://dx.doi.org/10.1016/J.CELL.2014.04.047}, DOI={10.1016/J.CELL.2014.04.047}, abstractNote={The transcription of genomic information in eukaryotes is regulated in large part by chromatin. How a diverse array of chromatin regulator (CR) proteins with different functions and genomic localization patterns coordinates chromatin activity to control transcription remains unclear. Here, we take a synthetic biology approach to decipher the complexity of chromatin regulation by studying emergent transcriptional behaviors from engineered combinatorial, spatial, and temporal patterns of individual CRs. We fuse 223 yeast CRs to programmable zinc finger proteins. Site-specific and combinatorial recruitment of CRs to distinct intralocus locations reveals a range of transcriptional logic and behaviors, including synergistic activation, long-range and spatial regulation, and gene expression memory. Comparing these transcriptional behaviors with annotated CR complex and function terms provides design principles for the engineering of transcriptional regulation. This work presents a bottom-up approach to investigating chromatin-mediated transcriptional regulation and introduces chromatin-based components and systems for synthetic biology and cellular engineering.}, number={1}, journal={Cell}, publisher={Elsevier BV}, author={Keung, Albert J. and Bashor, Caleb J. and Kiriakov, Szilvia and Collins, James J. and Khalil, Ahmad S.}, year={2014}, month={Jul}, pages={110–120} }
 @article{keung_dong_schaffer_kumar_2013, title={Pan-neuronal maturation but not neuronal subtype differentiation of adult neural stem cells is mechanosensitive}, volume={3}, ISSN={2045-2322}, url={http://dx.doi.org/10.1038/SREP01817}, DOI={10.1038/SREP01817}, abstractNote={Most past studies of the biophysical regulation of stem cell differentiation have focused on initial lineage commitment or proximal differentiation events. It would be valuable to understand whether biophysical inputs also influence distal endpoints more closely associated with physiological function, such as subtype specification in neuronal differentiation. To explore this question, we cultured adult neural stem cells (NSCs) on variable stiffness ECMs under conditions that promote neuronal fate commitment for extended time periods to allow neuronal subtype differentiation. We find that ECM stiffness does not modulate the expression of NeuroD1 and TrkA/B/C or the percentages of pan-neuronal, GABAergic, or glutamatergic neuronal subtypes. Interestingly, however, an ECM stiffness of 700 Pa maximizes expression of pan-neuronal markers. These results suggest that a wide range of stiffnesses fully permit pan-neuronal NSC differentiation, that an intermediate stiffness optimizes expression of pan-neuronal genes, and that stiffness does not impact commitment to particular neuronal subtypes.}, number={1}, journal={Scientific Reports}, publisher={Springer Science and Business Media LLC}, author={Keung, Albert J. and Dong, Meimei and Schaffer, David V. and Kumar, Sanjay}, year={2013}, month={May} }
 @article{mackay_keung_kumar_2012, title={A Genetic Strategy for the Dynamic and Graded Control of Cell Mechanics, Motility, and Matrix Remodeling}, volume={102}, ISSN={0006-3495}, url={http://dx.doi.org/10.1016/j.bpj.2011.12.048}, DOI={10.1016/j.bpj.2011.12.048}, abstractNote={Cellular mechanical properties have emerged as central regulators of many critical cell behaviors, including proliferation, motility, and differentiation. Although investigators have developed numerous techniques to influence these properties indirectly by engineering the extracellular matrix (ECM), relatively few tools are available to directly engineer the cells themselves. Here we present a genetic strategy for obtaining graded, dynamic control over cellular mechanical properties by regulating the expression of mutant mechanotransductive proteins from a single copy of a gene placed under a repressible promoter. With the use of constitutively active mutants of RhoA GTPase and myosin light chain kinase, we show that varying the expression level of either protein produces graded changes in stress fiber assembly, traction force generation, cellular stiffness, and migration speed. Using this approach, we demonstrate that soft ECMs render cells maximally sensitive to changes in RhoA activity, and that by modulating the ability of cells to engage and contract soft ECMs, we can dynamically control cell spreading, migration, and matrix remodeling. Thus, in addition to providing quantitative relationships between mechanotransductive signaling, cellular mechanical properties, and dynamic cell behaviors, this strategy enables us to control the physical interactions between cells and the ECM and thereby dictate how cells respond to matrix properties.}, number={3}, journal={Biophysical Journal}, publisher={Elsevier BV}, author={MacKay, Joanna L. and Keung, Albert J. and Kumar, Sanjay}, year={2012}, month={Feb}, pages={434–442} }
 @article{keung_asuri_kumar_schaffer_2012, title={Soft microenvironments promote the early neurogenic differentiation but not self-renewal of human pluripotent stem cells}, volume={4}, ISSN={1757-9708}, url={http://dx.doi.org/10.1039/c2ib20083j}, DOI={10.1039/c2ib20083j}, abstractNote={Human pluripotent stem cells (hPSCs) are of great interest in biology and medicine due to their ability to self-renew and differentiate into any adult or fetal cell type. Important efforts have identified biochemical factors, signaling pathways, and transcriptional networks that regulate hPSC biology. However, recent work investigating the effect of biophysical cues on mammalian cells and adult stem cells suggests that the mechanical properties of the microenvironment, such as stiffness, may also regulate hPSC behavior. While several studies have explored this mechanoregulation in mouse embryonic stem cells (mESCs), it has been challenging to extrapolate these findings and thereby explore their biomedical implications in hPSCs. For example, it remains unclear whether hPSCs can be driven down a given tissue lineage by providing tissue-mimetic stiffness cues. Here we address this open question by investigating the regulation of hPSC neurogenesis by microenvironmental stiffness. We find that increasing extracellular matrix (ECM) stiffness in vitro increases hPSC cell and colony spread area but does not alter self-renewal, in contrast to past studies with mESCs. However, softer ECMs with stiffnesses similar to that of neural tissue promote the generation of early neural ectoderm. This mechanosensitive increase in neural ectoderm requires only a short 5-day soft stiffness "pulse", which translates into downstream increases in both total neurons as well as therapeutically relevant dopaminergic neurons. These findings further highlight important differences between mESCs and hPSCs and have implications for both the design of future biomaterials as well as our understanding of early embryonic development.}, number={9}, journal={Integrative Biology}, publisher={Oxford University Press (OUP)}, author={Keung, Albert J. and Asuri, Prashanth and Kumar, Sanjay and Schaffer, David V.}, year={2012}, month={Aug}, pages={1049–1058} }
 @article{jang_koerber_kim_asuri_vazin_bartel_keung_kwon_park_schaffer_2011, title={An Evolved Adeno-associated Viral Variant Enhances Gene Delivery and Gene Targeting in Neural Stem Cells}, volume={19}, ISSN={1525-0016}, url={http://dx.doi.org/10.1038/mt.2010.287}, DOI={10.1038/mt.2010.287}, abstractNote={Gene delivery to, and gene targeting in, stem cells would be a highly enabling technology for basic science and biomedical application. Adeno-associated viral (AAV) vectors have demonstrated the capacity for efficient delivery to numerous cells, but their application to stem cells has been limited by low transduction efficiency. Due to their considerable advantages, however, engineering AAV delivery systems to enhance gene delivery to stem cells may have an impact in stem cell biology and therapy. Therefore, using several diverse AAV capsid libraries-including randomly mutagenized, DNA shuffled, and random peptide insertion variants-we applied directed evolution to create a "designer" AAV vector with enhanced delivery efficiency for neural stem cells (NSCs). A novel AAV variant, carrying an insertion of a selected peptide sequence on the surface of the threefold spike within the heparin-binding site, emerged from this evolution. Importantly, this evolved AAV variant mediated efficient gene delivery to rat, mouse, and human NSCs, as well as efficient gene targeting within adult NSCs, and it is thus promising for applications ranging from basic stem cell biology to clinical translation.}, number={4}, journal={Molecular Therapy}, publisher={Elsevier BV}, author={Jang, Jae-Hyung and Koerber, James T and Kim, Jung-Suk and Asuri, Prashanth and Vazin, Tandis and Bartel, Melissa and Keung, Albert and Kwon, Inchan and Park, Kook In and Schaffer, David V}, year={2011}, month={Apr}, pages={667–675} }
 @article{keung_de juan-pardo_schaffer_kumar_2011, title={Rho GTPases Mediate the Mechanosensitive Lineage Commitment of Neural Stem Cells}, volume={29}, ISSN={1066-5099}, url={http://dx.doi.org/10.1002/stem.746}, DOI={10.1002/stem.746}, abstractNote={Abstract}, number={11}, journal={STEM CELLS}, publisher={Wiley}, author={Keung, Albert J. and de Juan-Pardo, Elena M. and Schaffer, David V. and Kumar, Sanjay}, year={2011}, month={Oct}, pages={1886–1897} }
 @article{mackay_keung_kumar_2010, title={A Genetic Strategy for Graded and Dynamic Control of Cell-Matrix Mechanobiology}, volume={98}, ISSN={0006-3495}, url={http://dx.doi.org/10.1016/j.bpj.2009.12.4004}, DOI={10.1016/j.bpj.2009.12.4004}, abstractNote={Mechanical interactions between cells and the surrounding extracellular matrix, such as adhesion, contraction, and force transduction, play a central role in many fundamental cell behaviors, including proliferation, cell death, and motility. The ability to precisely manipulate the intracellular machinery that regulates these interactions could therefore provide a powerful tool for controlling the mechanical properties of living cells and could also allow us to re-engineer how cells sense and respond to mechanical stimuli in their microenvironment, which would be particularly useful for tissue engineering and cellular technologies where cells are interfaced with synthetic microenvironments. Towards this goal, we have genetically engineered stable cell lines in which we can precisely and dynamically alter the mechanobiological behavior of living cells by varying the activity of signal transduction proteins, such as RhoA GTPase, using constitutively active and dominant negative mutants under the control of a tetracycline-repressible promoter. Through a variety of imaging and biophysical techniques, including atomic force microscopy and traction force microscopy, we have demonstrated graded and dynamic control over cytoskeletal architecture, cell shape and spreading, contractility, and cellular stiffness. In addition, using glioblastoma multiforme as a model system, we show how these cell lines can be used to study the effects of altered cellular mechanical properties on cancer cell motility and invasion.}, number={3}, journal={Biophysical Journal}, publisher={Elsevier BV}, author={MacKay, Joanna L. and Keung, Albert and Kumar, Sanjay}, year={2010}, month={Jan}, pages={730a} }
 @article{keung_healy_kumar_schaffer_2010, title={Biophysics and dynamics of natural and engineered stem cell microenvironments}, volume={2}, ISSN={1939-5094 1939-005X}, url={http://dx.doi.org/10.1002/wsbm.46}, DOI={10.1002/wsbm.46}, abstractNote={Abstract}, number={1}, journal={Wiley Interdisciplinary Reviews: Systems Biology and Medicine}, publisher={Wiley}, author={Keung, Albert J. and Healy, Kevin E. and Kumar, Sanjay and Schaffer, David V.}, year={2010}, month={Jan}, pages={49–64} }
 @article{keung_de juan-pardo_schaffer_kumar_2010, title={Mechanotransductive Engineering of Neural Stem Cell Behavior}, volume={98}, ISSN={0006-3495}, url={http://dx.doi.org/10.1016/j.bpj.2009.12.2181}, DOI={10.1016/j.bpj.2009.12.2181}, abstractNote={Neural stem cells (NSCs) play important roles in learning and memory in the adult mammalian brain and may also serve as a source of cells in cell replacement therapies to treat neurodegenerative diseases. Therefore, investigating how NSC behavior is regulated is crucial to understanding the fundamental biology of the brain as well as in engineering biomedical therapies. Towards these ends, an increasing wealth of knowledge in the NSC field describes a complex picture of biochemical and genetic regulation of NSC self-renewal and differentiation. However, little is known about the biophysical control of NSC behavior by the extracellular matrix (ECM). Here we demonstrate that ECM-derived mechanical signals can act with Rho GTPases to regulate NSC stiffness and differentiation. Culturing NSCs on increasingly stiff ECMs suppresses neurogenesis and enhances gliogenesis, even in the absence of exogenous differentiating agents. This shift is accompanied by enhanced RhoA and Cdc42 activation and increased cellular stiffness. Direct manipulation of RhoA and Cdc42 activity disrupts the ability of NSCs to sense ECM stiffness and tips the balance between neurogenesis and gliogenesis in the presence and absence of exogenous differentiation cues. Inhibitors of a downstream effector of RhoA, Rho kinase, as well as inhibition of myosin II contractility rescues neuronal differentiation of NSCs cultured on stiff substrates as well as for NSCs expressing CA RhoA and CA Cdc42, suggesting that NSC stiffness/contractility regulates NSC differentiation. These results establish Rho GTPase-based mechanotransduction and cellular stiffness as novel regulators of NSC behavior.}, number={3}, journal={Biophysical Journal}, publisher={Elsevier BV}, author={Keung, Albert J. and de Juan-Pardo, Elena M. and Schaffer, David V. and Kumar, Sanjay}, year={2010}, month={Jan}, pages={404a–405a} }
 @article{keung_filler_bent_2007, title={Thermal Control of Amide Product Distributions at the Ge(100)-2×1 Surface}, volume={111}, ISSN={1932-7447 1932-7455}, url={http://dx.doi.org/10.1021/jp065278d}, DOI={10.1021/jp065278d}, abstractNote={We have investigated the adsorption of a series of primary, secondary, and tertiary amides including formamide, N-methylformamide, N-methylacetamide, and N,N-dimethylacetamide on the Ge(100)-2×1 surface using multiple internal reflection Fourier transform infrared spectroscopy and density functional theory. At 310 K, primary and secondary amides were observed to form thermodynamically favored N−H dissociation structures with interdimer interactions as well as kinetically favored oxygen dative-bonded structures. The relative surface product distributions could be controlled thermally. Dative-bonded adducts were isolated by exposing the amides to the surface at 240 K, whereas N−H dissociation products were formed by annealing to 450 K. While the acetamides could potentially form “-ene” type products, such products were not observed and instead the acetamides exhibited the same reactivity pattern as the formamides in this study.}, number={1}, journal={The Journal of Physical Chemistry C}, publisher={American Chemical Society (ACS)}, author={Keung, Albert J. and Filler, Michael A. and Bent, Stacey F.}, year={2007}, month={Jan}, pages={411–419} }
 @article{filler_van deventer_keung_bent_2006, title={Carboxylic Acid Chemistry at the Ge(100)-2 × 1 Interface:  Bidentate Bridging Structure Formation on a Semiconductor Surface}, volume={128}, ISSN={0002-7863 1520-5126}, url={http://dx.doi.org/10.1021/ja0549502}, DOI={10.1021/ja0549502}, abstractNote={The reactions of acetic acid, acetic-d3 acid-d, and formic acid with the Ge(100)-2 x 1 surface have been investigated using multiple internal reflection Fourier transform infrared (MIR-FTIR) spectroscopy, X-ray photoelectron spectroscopy (XPS), and density functional theory (DFT) calculations. The infrared and photoelectron data provide experimental evidence for an O-H dissociation product at 310 K. DFT calculations indicate that the O-H dissociation pathway is significantly favored, both kinetically and thermodynamically, over other potential reaction pathways. All of the carboxylic acids studied exhibit unexpected vibrational modes between 1400 and 1525 cm(-1), which are attributed to the presence of a bidentate bridging structure where both oxygen atoms interact directly with the surface.}, number={3}, journal={Journal of the American Chemical Society}, publisher={American Chemical Society (ACS)}, author={Filler, Michael A. and Van Deventer, James A. and Keung, Albert J. and Bent, Stacey F.}, year={2006}, month={Jan}, pages={770–779} }
 @article{keung_filler_porter_bent_2005, title={Tertiary amide chemistry at the Ge(100)-2×1 surface}, volume={599}, ISSN={0039-6028}, url={http://dx.doi.org/10.1016/j.susc.2005.09.035}, DOI={10.1016/j.susc.2005.09.035}, abstractNote={We have investigated the adsorption of several tertiary amides, including N,N-dimethylformamide, N,N-dimethylformamide-d7, 1-methyl-2-pyrrolidinone, and N-methylcaprolactam, on Ge(1 0 0)-2 × 1 using multiple internal reflection Fourier transform infrared spectroscopy and density functional theory. At 310 K, all four tertiary amides were observed to selectively form a dative bond to the germanium surface through the oxygen atom. While previous work has shown that oxygen dative bonds are unstable near room temperature, tertiary amides exhibit delocalization of electron density from nitrogen to oxygen, which appears to increase the stability of the oxygen dative-bonded state. Partial desorption of these surface adducts on the timescale of minutes indicates weakly bound surface adducts with coverage dependent binding energies.}, number={1-3}, journal={Surface Science}, publisher={Elsevier BV}, author={Keung, Albert J. and Filler, Michael A. and Porter, David W. and Bent, Stacey F.}, year={2005}, month={Dec}, pages={41–54} }