Biology subjects

Parkin, J.

Publications and source records attributed to Parkin, J..

4 recordsLinked to original sources

bioRxiv2018

Efa6 regulates axon growth, branching and maintenance by eliminating off-track microtubules at the cortex

Cortical collapse factors affect microtubule (MT) dynamics at the plasma membrane. They play important roles in neurons, as suggested by inhibition of axon growth and regeneration through the Arf activator Efa6 in C. elegans, and by neurodevelopmental disorders linked to the mammalian kinesin Kif21A. How cortical collapse factors influence axon growth is little understood. Here we studied them, focussing on the function of Drosophila Efa6 in experimentally and genetically amenable fly neurons. First, we show that Drosophila Efa6 can inhibit MTs directly without interacting molecules via an N-terminal 18 amino acid motif (MT elimination domain/MTED) that binds tubulin and inhibits microtubule growth in vitro and cells. If N-terminal MTED-containing fragments are in the cytoplasm they abolish entire microtubule networks of mouse fibroblasts and whole axons of fly neurons. Full-length Efa6 is membrane-attached, hence primarily blocks MTs in the periphery of fibroblasts, and explorative MTs that have left axonal bundles in neurons. Accordingly, loss of Efa6 causes an increase of explorative MTs: in growth cones, they enhance axon growth, in axon shafts, explorative MTs cause excessive branching, as well as atrophy through perturbations of MT bundles. Efa6 over-expression causes the opposite phenotypes. Taken together, our work conceptually links molecular and sub-cellular functions of cortical collapse factors to axon growth regulation and reveals new roles in axon branching and in the prevention of axonal atrophy. Furthermore, the MTED delivers a promising tool that can be used to inhibit MTs in a compartmentalised fashion when fusing it to specifically localising protein domains.\n\nSummary statementThe cortical collapse factor Efa6 inhibits microtubule polymerising outside axonal bundles. Thereby it limits axon growth and branching, but preserves microtubule bundle organisation crucial for axon maintenance.

cell biology↗

bioRxiv2017

Length and time scales of cell-cell signaling circuits in agar

A community of genetically heterogeneous cells embedded in an unmixed medium allows for sophisticated operations by retaining spatial differentiation and coordinating division-of-labor. To establish the principles of engineering reliable cell-cell communication in a heterogeneous environment, we examined how circuit parameters and spatial placement affect the range of length and time scales over which simple communication circuits interact. We constructed several \"sender\" and \"receiver\" strains with quorum-sensing signaling circuits. The sender cell colony produces acyl homoserine lactones (AHL), which diffuse across the semisolid medium. The receiver cell colony detects these signal molecules and reports by fluorescence. We have found that a single colony of one sender variant is sufficient to induce receiver response at more than 1.5cm separation. Furthermore, AHL degradase expression in receiver colonies produces a signal threshold effect and reduces the response level in subsequent receiver colonies. Finally, our investigation on the spatial placement of colonies gave rise to the design of a multicellular long-range communication array consisting of two alternating colony types. Its signal response successfully propagated colony-by-colony along a six-colony array spanning 4.8cm at a transmission velocity of 12.8 hours per colony or 0.075cm per hour. In addition, we have developed a reaction-diffusion model that recreates the observed behaviors of the many performed experiments using data-informed parameter estimates of signal diffusion, gene expression, and nutrient consumption. These results demonstrate that a mixed community of colonies can enable new patterning programs, and the corresponding model will facilitate the rational design of complex communication networks.

synthetic biology↗

bioRxiv2017

Enzyme-Free Nucleic Acid Dynamical Systems

Chemistries exhibiting complex dynamics--from inorganic oscillators to gene regulatory networks--have been long known but either cannot be reprogrammed at will, or rely on the sophisticated chemistry underlying the central dogma. Can simpler molecular mechanisms, designed from scratch, exhibit the same range of behaviors? Abstract coupled chemical reactions have been proposed as a programming language for complex dynamics, along with their systematic implementation using short synthetic DNA molecules. We developed this technology for dynamical systems, identifying critical design principles and codifying them into a compiler automating the design process. Using this approach, we built an oscillator containing only DNA components, establishing that Watson-Crick base pairing interactions alone suffice for arbitrarily complex dynamics. Our results argue that autonomous molecular systems that interact with and control their chemical environment can be designed via molecular programming languages.

bioengineering↗

bioRxiv2017

Engineering pulsatile communication in bacterial consortia

Lux-type quorum sensing systems enable communication in bacteria with only two protein components: a signal synthase and an inducible transcription activator. The simplicity of these systems makes them a popular choice for engineering collaborative behaviors in synthetic bacterial consortia, such as photographic edge detection and synchronized oscillation. To add to this body of work, we propose a pulsatile communication circuit that enables dynamic patterning and long-distance communication analogous to action potentials traveling through nerve tissue. We employed a model-driven design paradigm involving incremental characterization of in vivo design candidates with increasing circuit complexity. Beginning with a simple inducible reporter system, we screened a small number of circuits varying in their promoter and ribosomal binding site (RBS) strengths. From this candidate pool, we selected a candidate to be the seed network for the subsequent round of more complex circuit variants, likewise variable in promoter and RBS strengths. The selection criteria at each level of complexity is tailored to optimize a different desirable performance characteristic. By this approach we individually optimized reporter signal-to-background ratio, pulsatile response to induction, and quiescent basal transcription, avoiding large library screens while ensuring robust performance of the composite circuit.

synthetic biology↗