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\DOI{10.5802/crbiol.124}
\datereceived{2023-06-06}
\daterevised{2023-08-29}
\dateaccepted{2023-08-29}
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\dateposted{2023-09-21}
\begin{document}

\begin{noXML}

\RubricEF{Review Article}{Article de revue}

\title{Cell migration in dense microenvironments}

\alttitle{La migration cellulaire dans des microenvironnements encombr\'es}

\author{\firstname{Guilherme Pedreira} \lastname{de Freitas Nader}\CDRorcid{0000-0002-1905-1903}\IsCorresp}
\address{Department of Pathology and Laboratory Medicine, Children's
Hospital of Philadelphia and Perelman School of Medicine, University of
Pennsylvania, 3615 Civic Center Boulevard, Abramson Research Center,
Philadelphia, PA 19104-4399, USA}
\email[G. P. de Freitas Nader]{naderg@chop.edu}

\author{\firstname{Juan Manuel} \lastname{Garc\'{i}a-Arcos}\CDRorcid{0000-0003-3420-6093}}
\address{Department of Biochemistry, University of Geneva, CH-1205
Geneva, Switzerland}
\email[J. M. Garc\'{i}a-Arcos]{Juan.Garcia@unige.ch}

\keywords{\kwd{Cell migration}\kwd{Nucleus}\kwd{Nuclear envelope}}

\altkeywords{\kwd{Migration cellulaire}\kwd{Noyau}\kwd{Enveloppe nucl\'eaire}}

% \shortrunauthors

\def\thanksname{Note}

\thanks{Guilherme Nader received the Grandes Avanc\'ees en Biologie 2022
award.}

\begin{abstract} 
The nucleus has been viewed as a passenger during cell migration that
functions merely to protect the genome. However, increasing evidence
shows that the nucleus is an active organelle, constantly sensing the
surrounding environment and translating extracellular mechanical inputs
into intracellular signaling. The nuclear envelope has a large membrane
reservoir which serves as a buffer for mechanical inputs as it unfolds
without increasing its tension. In contrast, when cells cope with
mechanical strain, such as migration through solid tumors or dense
interstitial spaces, the nuclear envelope folds stretch, increasing
nuclear envelope tension and sometimes causing rupture. Different
degrees of nuclear envelope tension regulate cellular behaviors and
functions, especially in cells that move and grow within dense
matrices. The crosstalk between extracellular mechanical inputs and the
cell nucleus is a critical component in the modulation of cell function
of cells that navigate within packed microenvironments. Moreover, there
is a link between regimes of nuclear envelope unfolding and different
cellular behaviors, from orchestrated signaling cascades to cellular
perturbations and damage.
\end{abstract}

\begin{altabstract}
Le noyau a longtemps \'et\'e consid\'er\'e comme un passager lors de la
migration cellulaire, servant simplement \`a prot\'eger le g\'enome.
Cependant, de plus en plus de preuves montrent que le noyau est un
organite actif, qui sonde le milieu environnant et traduit les
entr\'ees m\'ecaniques extracellulaires en signalisation
intracellulaire. L'enveloppe nucl\'eaire possède un grand r\'eservoir
membranaire qui sert de tampon face aux entr\'ees m\'ecaniques en se
d\'epliant sans augmenter sa tension. En revanche, lorsque les cellules
font face \`a des contraintes m\'ecaniques, telles que la migration au
travers de tumeurs solides ou despaces interstitiels denses, les plis
de l'enveloppe nucl\'eaire s'\'etirent, augmentant sa tension et
provoquant parfois sa rupture. Diff\'erents degr\'es de tension de
l'enveloppe nucl\'eaire r\'egulent les comportements et les fonctions
cellulaires, en particulier des cellules qui se d\'eplacent et se
d\'eveloppent dans des matrices denses. La signalisation crois\'ee
entre les entr\'ees m\'ecaniques extracellulaires et le noyau
cellulaire sont des composants essentiels dans la modulation de la
fonction des cellules qui naviguent dans des microenvironnements
encombr\'es. De plus, il existe un lien entre les r\'egimes de
d\'eploiement de l'enveloppe nucl\'eaire et les diff\'erents
comportements cellulaires, allant des cascades de signalisation
jusquaux perturbations et dommages cellulaires.
\end{altabstract}

\maketitle

{\vspace*{12pt}}

\twocolumngrid

\end{noXML}

\section{Introduction}

During cell motility, cells must integrate multiple physical inputs
from the microenvironment as the extracellular matrix architecture,
fibers and cell packing limit the available space. Indeed, migratory
cells must constantly pass through narrow spaces as they migrate or
grow within densely packed microenvironments~\cite{1,2}.
{\vadjust{\pagebreak}}Therefore,
they frequently {experience} cellular and nuclear deformations. Since the
nucleus is the largest and stiffest cellular organelle, it~will often
constitute a limiting factor for confined cell migration. The nucleus
is a mechanosensory organelle that senses compressive forces and
rigidity of the extracellular environment and modulates migratory
strategies~\cite{3,4}. Importantly, in physiological contexts, when
growing in densely packed environments or migrating across landscapes
of varied porosities, geometries and densities, the nucleus is
subjected to a wide range of deformations that will in turn modulate
cell behavior, function and shape.

\section{Contexts of nuclear deformations: the example of cell
migration}
{\vspace*{-2pt}}
\subsection{Single cell migration}

Migration in complex 3D environments brings its own set of constraints
and presents many challenges to the cells, especially to their nuclei.
They often face the challenge of squeezing through narrow gaps and
constrictions in the surrounding tissue because of dense matrix
meshworks (interstitial migration). The crossing of narrow pores
results in a local compression of the cell and generates large
intracellular forces that are transmitted across the cytoskeleton to
the nucleus, leading to its deformation. Cells and cell collectives
possess different migratory modes and change their migratory behavior
in a tailored manner to adapt to complex 3D architectures of 
\textit{in~vivo} microenvironments. As a consequence of these highly
complex and dense scenarios, motile cells often exhibit features
associated with high nuclear deformability. For instance, leukocyte
transendothelial migration (TEM) involves the opening of micron-wide
gaps by the cells through paracellular junctions, followed by rapid
squeezing of their bulky nuclei through relatively tight endothelial
barriers (transmigration)~\cite{5,6,7,8} (Figure~\ref{fig1}).
Interestingly, a study has demonstrated an active role for the nucleus
in the process of TEM. Leukocyte nuclear lobes physically push and
generate gaps and pores in between the endothelial cells for successful
TEM~\cite{9}. Alternatively, several cell types change their protrusion
mode when migrating in constricted spaces, alternating between
lamellipodia and contractility-driven plasma membrane
blebs~\cite{10,11,12,13}. Some subtypes of aggressive invasive cancer
cells solve the problem by secreting extracellular proteases that
digest the matrix, therefore relieving the restrictive role of the
environment. However, many other types of motile cancer cells and a
large number of professionally migrating immune cell types do not
ubiquitously employ the matrix proteolysis strategy; rather, they adapt
their dimensions and migration choices to the available space around
them.{\vspace*{-6pt}} 

\begin{figure*}
{\vspace*{1pt}}
\includegraphics{fig01}
{\vspace*{1pt}}
\caption{\label{fig1}Schematic representation of the different contexts
of nuclear deformation and mechanosensitive signaling pathways
discussed in this review.}
{\vspace*{1pt}}
\end{figure*}

\subsection{Collective cell migration}

Besides the physical challenges experienced by single cells, mechanical
constraints are also present during migration of cell collectives,
potentially on a greater scale. Collective invasion of multiple cells
occurs when moving cells maintain functional cell--cell adhesion,
leading to a ``supracellular'' organization of the actin cytoskeleton
as well as multicellular polarity and traction force
generation~\cite{14,15}. In tissues, cells do not exist in isolation,
and the ability of cells to migrate collectively is crucial in shaping
organisms during the complex morphogenetic events of development, and
for several physiological and pathological events such as wound healing
and cancer metastasis. For instance, leader cells of collective
melanoma invasion developed adaptive invasion patterns along preformed
tracks of complex topography, combining single-cell and collective
migration modes, without immediate anatomic tissue remodeling or
destruction~\cite{16}. These events led to both cell body and nuclear
deformation. Likewise, the complex mechanical behavior of biological
tissues and their active nature constantly generate laminar flows,
swirling motions of cell clusters and vortices~\cite{15,17,18}. In
particular, cell extrusion events were shown to be preceded by a
coordinated, long-range flow of cells towards the eventual location of
the extrusion. This causes considerable bending of cells, leading to
high compressive stresses that are sufficient to trigger the extrusion
of a nearby cell~\cite{17}. Thus, such dynamic regime of tissues is
likely to impose intermittent physical strain to the cells and their
nuclei, ultimately affecting cell behavior, fate and tissue
homeostasis. Likewise, the movement of the collective strand of cells,
including invasion through tissue barriers, can inflict severe physical
challenges to the nucleus. When tissue gaps are smaller than the cell
width, the proteolytic degradation of boundaries or non-proteolytic
track widening (by passive outward pushing and space expansion) impose
strong deformations to the cells and their nuclei. This is illustrated
when breast cancer cells digest and breach the myoepithelial layer of
the confining mammary duct to invade into the stroma. During this
metastatic process, the collective motion of the strand of cells
breaching the barrier encounters a ``collective constriction'' (the
surrounding myoepithelial layer) that imposes high nuclear deformation
and DNA damage to the invasive strand~\cite{19,20} (Figure~\ref{fig1}).

In summary, cells can squeeze themselves not only when individually
navigating or growing in complex and dense microenvironments. They can
also squeeze each other in motile invasive strands or \mbox{during} aberrant
tissue overgrowth, such as in solid tumors~\cite{19}
(Figure~\ref{fig1}). Therefore, cells are constantly receiving
mechanical input from extracellular environments of different
topographical and physical features, and the nucleus has been a central
player in modulating the cellular responses. Critically, nuclear
plasticity and integrity will affect cell behavior and function,
ultimately impinging on physiological and pathological processes. This
demands the existence of specific mechanosensing mechanisms through
which cells can ``sense'' the nuclear deformation and the tension state
of the nuclear envelope (NE) to appropriately respond to them, thus
maintaining proper cellular and tissue functions.

\section{Signaling through nuclear mechanosensing}

As cells sense the complexity of the extracellular microenvironments,
mechanical signals are transmitted to the nucleus via the cytoskeleton
and converted into signaling pathways to regulate cell behavior and
function. The first study to show mechanical continuity between cell
surface receptors and the nucleus was performed by the group of Donald
Ingber. Upon micromanipulation of ligand-coated microbeads, they
observed that cytoskeletal filaments reoriente, nuclei distorte, and
nucleoli redistributs along the axis of the applied tension
field~\cite{21}. Moreover, forces applied to integrins also induced
displacements of nuclei up to 20~$\upmu$m away from the site of force
application, whereas generalized deformation of the surface bilayer
only produced local effects at the cell surface~\cite{22}. Now, we know
that such phenomenon of mechanosensation and force transmission through
the cytoplasm to the nucleus occurs via the LINC
complex~\cite{23} (Figure~\ref{fig1}). Additionally, nucleoskeleton and
transmembrane NE proteins bind chromatin, generating stress at the
level of the NE, leading to nuclear deformation and changes in
chromatin organization~\cite{24,25,26} (Figure~\ref{fig1}). Therefore,
these mechanical signals ultimately influence gene expression and
control multiple aspects of cell behavior, including growth and
differentiation, and even cancer progression. Additionally, during
nuclear mechanosensing, nuclear pores physically expand and contract,
thus altering their transport rates. Indeed, more recently, the direct
effect of nuclear deformation on the import of transcription factors
has brought back into the picture the deformation of the nucleus, and
more precisely the NE as a sensor of mechanical constraints.
Importantly, this study contributed with a potential general
applicability for mechanosensing pathways in transcriptional
regulation~\cite{27}. Altogether, these findings demonstrate that
mechanical forces that are applied at the cell surface do more than
activate membranesignaling events. The subsequent mechanosensing
pathways promote structural rearrangements deep in the nucleus. The
magnitude of the signal will shape both short- and long-term \mbox{responses,}
differently impacting gene \mbox{regulation,} cell behavior and cell fate.
Broadly, modulation of the NE state can affect more than an individual
cell at a given time. The consequences can be far-reaching, be it
spatially or temporally, all the way to impacting tissue homeostasis
and developmental fate. Critically, extreme perturbations on cells and
their nuclei might also have detrimental effects. Therefore, how cells
adapt their responses or avoid harmful perturbations when growing and
migrating within confined 3D microenvironments becomes critical.
Indeed, in more extreme cases, nuclear blebs have been shown to
systematically form and rupture at sites of high local nuclear
curvature~\cite{28,29,30,31}. Critically, these events might bring
harmful consequences such as repeated loss of nucleo-cytoplasmic
compartmentalization and chronic DNA damage~\cite{1,2,19,32}. In
contrast, severe nuclear deformations might also trigger mechanisms
that help cells navigate the complexity of dense matrices. In an
elegant work, Lomakin and colleagues showed that cells measure the
degree of spatial confinement using their nucleus. When deformed below
a specific height, the NE unfolds to its maximum and it becomes tensed.
The augmented NE tension leads to recruitment of cytosolic
phospholipase A2 (cPLA2) to the inner NE surface, causing an increase
in cortical actomyosin contractility, producing pushing forces to
resist physical compression~\cite{33}. NE stretching has also been
shown to play a critical role in modulating inflammatory response via
cPLA2~\cite{34} (Figure~\ref{fig1}).

Another major perturbation experienced by cells under mechanical strain
lies at the level of the chromatin. Chromosome dynamics and motion are
locally transmitted to discrete sites of the NE via lamina-associated
domains (LADs). LADs and heterochromatinization can be responsible for
forming large NE folds that will pull on the NE and generate
topological tension~\cite{35}. Moreover, there is evidence of
cross-talk between the NE and chromatin state during cell migration.
For instance, epigenetic reprogramming was shown to start shortly after
initiation of migration. Likewise, treatment of cells with drugs to
abolish epigenetic modifications led to decreased
motility~\cite{36,37}. It is also plausible that the fluctuations on NE
tension experienced by cells migrating on stiff matrices or in confined
3D microenvironments might change their transcription
programs~\cite{26,38}.

\section{Conclusions}

The crosstalk between microenvironmental physical strains and the cell
nucleus triggers nuclear mechanosensing pathways that regulate cellular
homeostasis and cell behavior. Critically, in order to maintain
cellular homeostasis, a fine balance must exist between what cells
sense as a perturbation (potentially threatening their integrity), and
the signaling pathways that orchestrate their behavior, function, and
at a greater level, tissue homeostasis. In this context, the nucleus
has emerged as a key mechanosensory organelle engaged in tuning these
responses. In the light of evolution, frequent and unavoidable sources
of damages, such as extreme nuclear deformations and NE ruptures, are
likely to be integrated into signaling pathways to modulate cellular
behaviors, fate and tissue physiopathology.

\section*{Declaration of interests}

The authors do not work for, advise, own shares in, or receive funds
from any organization that could benefit from this article, and have
declared no affiliations other than their research organizations.

\section*{Acknowledgements}
We thank Dr. Matthieu Piel (Institut Curie, Paris, France) for all the
scientific input, invaluable mentoring and for providing the funding
for the postdoctoral research of GPFN, which made this article
possible. 

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