Helical swimming is a ubiquitous strategy for motile cells to generate self-gradients for environmental sensing. The model biflagellate Chlamydomonas reinhardtii rotates at a constant 1 – 2 Hz as it swims, but the mechanism is unclear. Here, we show unequivocally that the rolling motion derives from a persistent, non-planar flagellar beat pattern. This is revealed by high-speed imaging and micromanipulation of live cells. We construct a fully-3D model to relate flagellar beating directly to the free-swimming trajectories. For realistic geometries, the model reproduces both the sense and magnitude of the axial rotation of live cells. We show that helical swimming requires further symmetry-breaking between the two flagella. These functional differences underlie all tactic responses, particularly phototaxis. We propose a control strategy by which cells steer towards or away from light by modulating the sign of biflagellar dominance.

All living cells interact dynamically with a constantly changing world.
Eukaryotes in particular, evolved radically new ways to sense and react to
their environment. These advances enabled new and more complex forms of
cellular behavior in eukaryotes, including directional movement, active
feeding, mating, or responses to predation. But what are the key events and
innovations during eukaryogenesis that made all of this possible? Here we
describe the ancestral repertoire of eukaryotic excitability and discuss five
major cellular innovations that enabled its evolutionary origin. The
innovations include a vastly expanded repertoire of ion channels, endomembranes
as intracellular capacitors, a flexible plasma membrane, the emergence of cilia
and pseudopodia, and the relocation of chemiosmotic ATP synthesis to
mitochondria that liberated the plasma membrane for more complex electrical
signaling involved in sensing and reacting. We conjecture that together with an
increase in cell size, these new forms of excitability greatly amplified the
degrees of freedom associated with cellular responses, allowing eukaryotes to
vastly outperform prokaryotes in terms of both speed and accuracy. This
comprehensive new perspective on the evolution of excitability enriches our
view of eukaryogenesis and emphasizes behaviour and sensing as major
contributors to the success of eukaryotes.

Beating flagella exhibit a variety of synchronization modes. This synchrony
has long been attributed to hydrodynamic coupling between the flagella.
However, recent work with flagellated algae indicates that a mechanism internal
to the cell, through the contractile fibres connecting the flagella basal
bodies, must be at play to actively modulate flagellar synchrony. Exactly how
basal coupling mediates flagellar coordination remains unclear. Here, we
examine the role of basal coupling in the synchronization of the model
biflagellate \textit{Chlamydomonas reinhardtii} using a series of mathematical
models of decreasing level of complexity. We report that basal coupling is
sufficient to achieve inphase, antiphase, and bistable synchrony, even in the
absence of hydrodynamic coupling and flagellar compliance. These modes can be
reached by modulating the activity level of the individual flagella or the
strength of the basal coupling. We observe a slip mode when allowing for
differential flagellar activity, just as in experiments with live cells. We
introduce a dimensionless ratio of flagellar activity to basal coupling that is
predictive of the mode of synchrony. This ratio allows us to query biological
parameters which are not yet directly measurable experimentally. Our work shows
a concrete route for cells to actively control the synchronization of their
flagella.

Heterodimeric motor organization of kinesin-II is essential for its function in anterograde IFT in ciliogenesis. However, the underlying mechanism is not well understood. In addition, the anterograde IFT velocity varies significantly in different organisms, but how this velocity affects ciliary length is not clear. We show that in Chlamydomonas motors are only stable as heterodimers in vivo, which is likely the key factor for the requirement of a heterodimer for IFT. Second, chimeric CrKinesin-II with human kinesin-II motor domains functioned in vitro and in vivo, leading to a ~ 2.8 fold reduced anterograde IFT velocity and a similar fold reduction in IFT injection rate that supposedly correlates with ciliary assembly activity. However, the ciliary length was only mildly reduced (~15%). Modeling analysis suggests a nonlinear scaling relationship between IFT velocity and ciliary length that can be accounted for by limitation of the motors and/or its ciliary cargoes, e.g. tubulin.

Cilia are specialized cellular organelles that are united in structure and implicated in diverse key life processes across eukaryotes. In both unicellular and multicellular organisms, variations on the same ancestral form mediate sensing, locomotion and the production of physiological flows. As we usher in a new, more interdisciplinary era, the way we study cilia is changing. This special theme issue brings together biologists, biophysicists and mathematicians to highlight the remarkable range of systems in which motile cilia fulfil vital functions, and to inspire and define novel strategies for future research.
. This article is part of the Theo Murphy meeting issue ‘Unity and diversity of cilia in locomotion and transport’.

. The phenomenon of ciliary coordination has garnered increasing attention in recent decades and multiple theories have been proposed to explain its occurrence in different biological systems. While hydrodynamic interactions are thought to dictate the large-scale coordinated activity of epithelial cilia for fluid transport, it is rather basal coupling that accounts for synchronous swimming gaits in model microeukaryotes such as
. Chlamydomonas.
. Unicellular ciliates present a fascinating yet understudied context in which coordination is found to persist in ciliary arrays positioned across millimetre scales on the same cell. Here, we focus on the ciliate
. Stentor coeruleus
. chosen for its large size, complex ciliary organization, and capacity for cellular regeneration. These large protists exhibit ciliary differentiation between cortical rows of short body cilia used for swimming, and an anterior ring of longer, fused cilia called the membranellar band (MB). The oral cilia in the MB beat metachronously to produce strong feeding currents. Remarkably, upon injury, the MB can be shed and regenerated de novo. Here, we follow and track this developmental sequence in its entirety to elucidate the emergence of coordinated ciliary beating: from band formation, elongation, curling and final migration towards the cell anterior. We reveal a complex interplay between hydrodynamics and ciliary restructuring in
. Stentor
. and highlight for the first time the importance of a ring-like topology for achieving long-range metachronism in ciliated structures.
.
. This article is part of the Theo Murphy meeting issue ‘Unity and diversity of cilia in locomotion and transport’.

. Living creatures exhibit a remarkable diversity of locomotion mechanisms, evolving structures specialized for interacting with their environment. In the vast majority of cases, locomotor behaviours such as flying, crawling and running are orchestrated by nervous systems. Surprisingly, microorganisms can enact analogous movement gaits for swimming using multiple, fast-moving cellular protrusions called cilia and flagella. Here, I demonstrate intermittency, reversible rhythmogenesis and gait mechanosensitivity in algal flagella, to reveal the active nature of locomotor patterning. In addition to maintaining free-swimming gaits, I show that the algal flagellar apparatus functions as a central pattern generator that encodes the beating of each flagellum in a network in a
. distinguishable
. manner. The latter provides a novel symmetry-breaking mechanism for cell reorientation. These findings imply that the capacity to generate and coordinate complex locomotor patterns does not require neural circuitry but rather the minimal ingredients are present in simple unicellular organisms.
.
. This article is part of the Theo Murphy meeting issue ‘Unity and diversity of cilia in locomotion and transport’.

© 2018 the Author(s). Propulsion by slender cellular appendages called cilia and flagella is an ancient means of locomotion. Unicellular organisms evolved myriad strategies to propel themselves in fluid environments, often involving significant differences in flagella number, localisation and modes of actuation. Remarkably, these appendages are highly conserved, occurring in many complex organisms such as humans, where they may be found generating physiological flows when attached to surfaces (e.g. airway epithelial cilia), or else conferring motility to male gametes (e.g. undulations of sperm flagella). Where multiple cilia arise, their movements are often observed to be highly coordinated. Here I review the two main mechanisms for motile cilia coordination, namely, intracellular and hydrodynamic, and discuss their relative importance in different ciliary systems.

© 2018 authors. Published by the American Physical Society. Published by the American Physical Society under the terms of the »https://creativecommons.org/licenses/by/4.0/» Creative Commons Attribution 4.0 International license. Further distribution of this work must maintain attribution to the author(s) and the published article's title, journal citation, and DOI. Active living organisms exhibit behavioral variability, partitioning between fast and slow dynamics. Such variability may be key to generating rapid responses in a heterogeneous, unpredictable environment wherein cellular activity effects continual exchanges of energy fluxes. We demonstrate a novel, noninvasive strategy for revealing nonequilibrium control of swimming - specifically, in an octoflagellate microalga. These organisms exhibit surprising features of flagellar excitability and mechanosensitivity, which characterize a novel, time-irreversible "run-stop-shock" motility comprising forward runs, knee-jerk shocks with dramatic beat reversal, and long stops during which cells are quiescent yet continue to exhibit submicron flagellar vibrations. Entropy production, associated with flux cycles arising in a reaction graph representation of the gait-switching dynamics, provides a direct measure of detailed balance violation in this primitive alga.

Cilia and flagella often exhibit synchronized behavior; this includes phase locking, as seen inChlamydomonas, 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 algaVolvoxhas 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 ofChlamydomonasmutants 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.

Flows generated by ensembles of flagella are crucial to development, motility and sensing, but the mechanisms behind this striking coordination remain unclear. We present novel experiments in which two micropipette-held somatic cells of Volvox carteri, with distinct intrinsic beating frequencies, are studied by high-speed imaging as a function of their separation and orientation. Analysis of time series shows that the interflagellar coupling, constrained by lack of connections between cells to be hydrodynamical, exhibits a spatial dependence consistent with theory. At close spacings it produces robust synchrony for thousands of beats, while at increasing separations synchrony is degraded by stochastic processes. Manipulation of the relative flagellar orientation reveals in-phase and antiphase states, consistent with dynamical theories. Flagellar tracking with exquisite precision reveals waveform changes that result from hydrodynamic coupling. This study proves unequivocally that flagella coupled solely through a fluid can achieve robust synchrony despite differences in their intrinsic properties.

. 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.
.