Publications

Motivated by the question of how biological systems maintain homeostasis in changing environments, Shinar and Feinberg introduced in 2010 the concept of absolute concentration robustness (ACR). A biochemical system exhibits ACR in some species if the steady-state value of that species does not depend on initial conditions. Thus, a system with ACR can maintain a constant level of one species even as the initial condition changes. Despite a great deal of interest in ACR in recent years, the following basic question remains open: How can we determine quickly whether a given biochemical system has ACR? Although various approaches to this problem have been proposed, we show that they are incomplete. Accordingly, we present new methods for deciding ACR, which harness computational algebra. We illustrate our results on several biochemical signaling networks.

Tight junctions play an essential role in sealing tissues, by forming belts of adhesion strands around cellular perimeters. Recent work has shown that the condensation of ZO1 scaffold proteins is required for tight junction assembly. However, the mechanisms by which junctional condensates initiate at cell-cell contacts and elongate around cell perimeters remain unknown. Combining biochemical reconstitutions and live-cell imaging of MDCKII tissue, we found that tight junction belt formation is driven by adhesion receptor-mediated ZO1 surface condensation coupled to local actin polymerization. Adhesion receptor oligomerization provides the signal for surface binding and local condensation of ZO1 at the cell membrane. Condensation produces a molecular scaffold that selectively enriches junctional proteins. Finally, ZO1 condensates directly facilitate local actin polymerization and filament bundling, driving the elongation into a continuous tight junction belt. More broadly, our work identifies how cells couple surface condensation with cytoskeleton organization to assemble and structure adhesion complexes.

Biomolecular condensates are thought to create subcellular microenvironments that regulate specific biochemical activities. Extensive in vitro work has helped link condensate formation to a wide range of cellular processes, including gene expression, nuclear transport, signalling and stress responses. However, testing the relationship between condensate formation and function in cells is more challenging. In particular, the extent to which the cellular functions of condensates depend on the nature of the molecular interactions through which the condensates form is a major outstanding question. Here, we review results from recent genetic complementation experiments in cells, and highlight how genetic complementation provides important insights into cellular functions and functional specificity of biomolecular condensates. Combined with observations from human genetic disease, these experiments suggest that diverse condensate-promoting regions within cellular proteins confer different condensate compositions, biophysical properties and functions.

Increasing insights into how sequence motifs in intrinsically disordered regions (IDRs) provide functions underscore the need for systematic motif detection. Contrary to structured regions where motifs can be readily identified from sequence alignments, the rapid evolution of IDRs limits the usage of alignment-based tools in reliably detecting motifs within. Here, we developed SHARK-capture, an alignment-free motif detection tool designed for difficult-to-align regions. SHARK-capture innovates on word-based methods by flexibly incorporating amino acid physicochemistry to assess motif similarity without requiring rigid definitions of equivalency groups. SHARK-capture offers consistently strong performance in a systematic benchmark, with superior residue-level performance. SHARK-capture identified known functional motifs across orthologs of the microtubule-associated zinc finger protein BuGZ. We also identified a short motif in the IDR of S. cerevisiae RNA helicase Ded1p, which we experimentally verified to be capable of promoting ATPase activity. Our improved performance allows us to systematically calculate 10,889 motifs for 2695 yeast IDRs and provide it as a resource. SHARK-capture offers the most precise tool yet for the systematic identification of conserved regions in IDRs and is freely available as a Python package (https://pypi.org/project/bio-shark/) and on https://git.mpi-cbg.de/tothpetroczylab/shark.

Kinesin-1-powered microtubules have emerged as versatile components in biocomputing and biosensing technologies. However, the inability to identify and track individual microtubules has constrained their applications to ensemble behaviors, limiting their potential for single-entity-based nanotechnologies. To address this challenge, we present a novel method for encoding digital information directly onto individual microtubules using photobleaching patterns. Binary numbers (1 to 15) were encoded within ∼12 μm segments of moving microtubules by photobleaching with a stationary pulsed laser, creating spatial frequency patterns corresponding to distinct bits of information. Fourier analysis enabled the accurate retrieval of the encoded data, demonstrating the feasibility of direct information storage and retrieval on macromolecular structures. This approach offers a transformative solution for recording microtubule trajectories within nanotechnological devices by encoding path information directly onto microtubules at branch points, obviating the need for video-based tracking. We anticipate that this innovation will advance the development of individualized microtubule-based technologies.

We review empirical methods that can be used to provide physical descriptions of dynamic cellular processes during development and disease. Our focus will be nonspatial descriptions and the inference of underlying interaction networks including cell-state lineages, gene regulatory networks, and molecular interactions in living cells. Our overarching questions are: How much can we learn from just observing? To what degree is it possible to infer causal and/or precise mathematical relationships from observations? We restrict ourselves to data sets arising from only observations, or experiments in which minimal perturbations have taken place to facilitate observation of the systems as they naturally occur. We discuss analysis perspectives in order from those offering the least descriptive power but requiring the least assumptions such as statistical associations. We end with those that are most descriptive, but require stricter assumptions and more previous knowledge of the systems such as causal inference and dynamical systems approaches. We hope to provide and encourage the use of a wide array of options for quantitative cell biologists to learn as much as possible from their observations at all stages of understanding of their system of interest. Finally, we provide our own recipe of how to empirically determine quantitative relationships and growth laws from live-cell microscopy data, the resultant predictions of which can then be verified with perturbation experiments. We also include an extended supplement that describes further inference algorithms and theory for the interested reader.

Common genetic variants in a conserved cis-regulatory element (CRE) at histone deacetylase (HDAC)9 are a major risk factor for cardiovascular disease, including stroke and coronary artery disease. Given the consistency of this association and its proinflammatory properties, we examined the mechanisms whereby HDAC9 regulates vascular inflammation. HDAC9 bound and mediated deacetylation of NLRP3 in the NACHT and LRR domains leading to inflammasome activation and lytic cell death. Targeted deletion of the critical CRE in mice increased Hdac9 expression in myeloid cells to exacerbate inflammasome-dependent chronic inflammation. In human carotid endarterectomy samples, increased HDAC9 expression was associated with atheroprogression and clinical plaque instability. Incorporation of TMP195, a class IIa HDAC inhibitor, into lipoprotein-based nanoparticles to target HDAC9 at the site of myeloid-driven vascular inflammation stabilized atherosclerotic plaques, implying a lower risk of plaque rupture and cardiovascular events. Our findings link HDAC9 to atherogenic inflammation and provide a paradigm for anti-inflammatory therapeutics for atherosclerosis.

The glycocalyx, a highly heterogeneous glycoprotein layer of cilia regulates adhesion and force transduction and is involved in signaling. The high-resolution molecular architecture of this layer is currently not understood. The structure of the ciliary coat is described in the green alga Chlamydomonas reinhardtii by cryo-electron tomography and proteomic approaches and the high-resolution cryoEM structure of the main component, FMG1B is solved. FMG1B is described as a mucin orthologue which lacks the major O-glycosylation of mammalian mucins but is N-glycosylated. FMG1A, a previously undescribed isoform of FMG1B is expressed in C. reinhardtii. By microflow-based adhesion assays, increased surface adhesion in the glycocalyx deficient double-mutant fmg1b-fmg1a is observed. It is found this mutant is capable of surface-gliding, with neither isoform required for extracellular force transduction by intraflagellar transport. The results find FMG1 to form a protective layer with adhesion-regulative instead of adhesion-conferring properties and an example of an undescribed class of mucins.