postdoc/JRF, University of Cambridge
Absent a brain, how do microorganisms coordinate microscale appendages for diverse swimming gaits?
My research primarily concerns the motility of microorganisms and the biophysics of micro-swimming. The slender propulsion-generating appendages used by free-living eukaryotes for the purposes of swimming are structurally, morphologically, and behaviourally similar to the epithelial cilia that are at this very moment pumping physiological flows inside the human body. Consequently, the very same physical theory, namely one of “creeping” or “low-Reynolds number” flows in which viscous forces are dominant over inertial forces, can be used to elucidate the active dynamics.
One key mystery concerns the origins of ciliary or flagellar coordination: after all, there can be little benefit in having multiple flagella if you cannot coordinate them effectively (think three-legged races)… We sought to address this question using species of flagellate algae as our preferred model system. These span several orders of magnitude in size and flagella number, which allow us to reveal a fascinating balance between active biological control and passive physical interactions that changes or evolves with increasing system size or complexity.
For instance, the stereotypical biflagellate alga Chlamydomonas actuates two nearly- identical flagella in unison in a breaststroke gait, while the large spherical alga Volvox rotates through the fluid using thousands of surface attached flagella which beat metachronously. In the latter, neighbouring flagella interact directly through the fluid, so no centralised control is necessary for spontaneous emergence of waves, much like the rise and fall of hands in a stadium. Species that have precisely four flagella can even produce locomotion patterns that resemble the trotting, galloping, pronking gaits of horses and other quadrupeds.
Disruption or mutation of ciliary coordination can not only result in reduced swimming and navigational efficacy in algae, but moreover in complex syndromes or ciliopathies in mammals. Such ciliopathies can have further downstream consequences including blindness, obesity and kidney disease.
Abstract: The motility of cilia and flagella of eukaryotic cells is controlled by second messengers such as Ca(2+), cAMP, and cGMP. In this study, the cAMP-dependent control of flagellar bending of Chlamydomonas is investigated by applying cAMP through photolysis of 4,5-dimethoxy-2-nitrobenzyl adenosine 3',5'-cyclicmonophosphate (caged cAMP). When cAMP is applied to demembranated and reactivated cells, cells begin to swim with a larger helical path. This change is due to a larger turn about the axis normal to the anterior-posterior axis, indicating an increased imbalance in the propulsive forces generated by the cis-flagellum (flagellum nearer to the eyespot) and trans-flagellum (flagellum farther from the eyespot). Consistently, when cAMP is applied to isolated axonemes, some axonemes show attenuated motility whereas others do not. Axonemes from uni1 mutants, which have only trans-flagella, do not respond to cAMP. These observations indicate that cAMP controls the balance of the forces generated by cis- and trans-flagella in Chlamydomonas.
Pub.: 11 Aug '15, Pinned: 25 Jun '17
Abstract: In a multitude of life's processes, cilia and flagella are found indispensable. Recently, the biflagellated chlorophyte alga Chlamydomonas has become a model organism for the study of ciliary motility and synchronization. Here, we use high-speed, high-resolution imaging of single pipette-held cells to quantify the rich dynamics exhibited by their flagella. Underlying this variability in behaviour are biological dissimilarities between the two flagella-termed cis and trans, with respect to a unique eyespot. With emphasis on the wild-type, we derive limit cycles and phase parametrizations for self-sustained flagellar oscillations from digitally tracked flagellar waveforms. Characterizing interflagellar phase synchrony via a simple model of coupled oscillators with noise, we find that during the canonical swimming breaststroke the cis flagellum is consistently phase-lagged relative to, while remaining robustly phase-locked with, the trans flagellum. Transient loss of synchrony, or phase slippage, may be triggered stochastically, in which the trans flagellum transitions to a second mode of beating with attenuated beat envelope and increased frequency. Further, exploiting this alga's ability for flagellar regeneration, we mechanically induced removal of one or the other flagellum of the same cell to reveal a striking disparity between the beatings of the cis and trans flagella, in isolation. These results are evaluated in the context of the dynamic coordination of Chlamydomonas flagella.
Pub.: 28 Feb '14, Pinned: 25 Jun '17
Abstract: Cilia and flagella are model systems for studying how mechanical forces control morphology. The periodic bending motion of cilia and flagella is thought to arise from mechanical feedback: dynein motors generate sliding forces that bend the flagellum, and bending leads to deformations and stresses, which feed back and regulate the motors. Three alternative feedback mechanisms have been proposed: regulation by the sliding forces, regulation by the curvature of the flagellum, and regulation by the normal forces that deform the cross-section of the flagellum. In this work, we combined theoretical and experimental approaches to show that the curvature control mechanism is the one that accords best with the bending waveforms of Chlamydomonas flagella. We make the surprising prediction that the motors respond to the time derivative of curvature, rather than curvature itself, hinting at an adaptation mechanism controlling the flagellar beat.
Pub.: 12 May '16, Pinned: 25 Jun '17
Abstract: The mechanisms underlying the coordinated beating of cilia and flagella remain incompletely understood despite the fundamental importance of these organelles. The axoneme (the cytoskeletal structure of cilia and flagella) consists of microtubule doublets connected by passive and active elements. The motor protein dynein is known to drive active bending, but dynein activity must be regulated to generate oscillatory, propulsive waveforms. Mathematical models of flagellar motion generate quantitative predictions that can be analysed to test hypotheses concerning dynein regulation. One approach has been to seek periodic solutions to the linearized equations of motion. However, models may simultaneously exhibit both periodic and unstable modes. Here, we investigate the emergence and coexistence of unstable and periodic modes in three mathematical models of flagellar motion, each based on a different dynein regulation hypothesis: (i) sliding control; (ii) curvature control and (iii) control by interdoublet separation (the 'geometric clutch' (GC)). The unstable modes predicted by each model are used to critically evaluate the underlying hypothesis. In particular, models of flagella with 'sliding-controlled' dynein activity admit unstable modes with non-propulsive, retrograde (tip-to-base) propagation, sometimes at the same parameter values that lead to periodic, propulsive modes. In the presence of these retrograde unstable modes, stable or periodic modes have little influence. In contrast, unstable modes of the GC model exhibit switching at the base and propulsive base-to-tip propagation.
Pub.: 04 Apr '15, Pinned: 25 Jun '17
Abstract: Cross-linked filament bundles, such as in cilia and flagella, are ubiquitous in biology. They are considered in textbooks as simple filaments with larger stiffness. Recent observations of flagellar counterbend, however, show that induction of curvature in one section of a passive flagellum instigates a compensatory counter-curvature elsewhere, exposing the intricate role of the diminutive cross-linking proteins at large scales. We show that this effect, a material property of the cross-linking mechanics, modifies the bundle dynamics and induces a bimodal L(2) - L(3) length-dependent material response that departs from the Euler-Bernoulli theory. Hence, the use of simpler theories to analyse experiments can result in paradoxical interpretations. Remarkably, the counterbend dynamics instigates counter-waves in opposition to driven oscillations in distant parts of the bundle, with potential impact on the regulation of flagellar bending waves. These results have a range of physical and biological applications, including the empirical disentanglement of material quantities via counterbend dynamics.
Pub.: 02 Jun '17, Pinned: 25 Jun '17
Abstract: A fundamental issue in the biology of eukaryotic flagella is the origin of synchronized beating observed in tissues and organisms containing multiple flagella. Recent studies of the biflagellate unicellular alga Chlamydomonas reinhardtii provided the first evidence that the interflagellar coupling responsible for synchronization is of hydrodynamic origin. To investigate this mechanism in detail, we study here synchronization in Chlamydomonas as its flagella slowly regrow after mechanically induced self-scission. The duration of synchronized intervals is found to be strongly dependent on flagellar length. Analysis within a stochastic model of coupled phase oscillators is used to extract the length dependence of the interflagellar coupling and the intrinsic beat frequencies of the two flagella. Physical and biological considerations that may explain these results are proposed.
Pub.: 24 Nov '11, Pinned: 24 Jun '17
Abstract: Cilia and flagella often exhibit synchronized behavior; this includes phase locking, as seen in Chlamydomonas, and metachronal wave formation in the respiratory cilia of higher organisms. Since the observations by Gray and Rothschild of phase synchrony of nearby swimming spermatozoa, it has been a working hypothesis that synchrony arises from hydrodynamic interactions between beating filaments. Recent work on the dynamics of physically separated pairs of flagella isolated from the multicellular alga Volvox has shown that hydrodynamic coupling alone is sufficient to produce synchrony. However, the situation is more complex in unicellular organisms bearing few flagella. We show that flagella of Chlamydomonas mutants deficient in filamentary connections between basal bodies display markedly different synchronization from the wild type. We perform micromanipulation on configurations of flagella and conclude that a mechanism, internal to the cell, must provide an additional flagellar coupling. In naturally occurring species with 4, 8, or even 16 flagella, we find diverse symmetries of basal body positioning and of the flagellar apparatus that are coincident with specific gaits of flagellar actuation, suggesting that it is a competition between intracellular coupling and hydrodynamic interactions that ultimately determines the precise form of flagellar coordination in unicellular algae.
Pub.: 02 May '16, Pinned: 24 Jun '17
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