Cytoskeleton Dynamics in the Retina

Abstract

Background:
Cytoskeleton is one of the essential forms of protein, important in the existence of both eukaryotic as well as prokaryotic cells. Its transformation plays a vital role in cell division and intracellular transportation by facilitating intracellular vesicular traffic. Among the various tissue types in the body, the neural tissue exhibits the maximum heterogeneity, and hence the role of cytoskeleton at both developmental and functional levels becomes paramount. Cytoskeleton dynamics have been established in the neural physiology, but only at the level of axonal development and growth. Retina has not been adequately studied in the context of cytoskeletal proteins.

Methods:
We reviewed the last 10 years of literature with reference to the development, growth, degeneration, and regeneration of the retina and the role of cytoskeleton in each aspect. We have focused on various changes that the retina undergoes at the cytosolic and cytoskeletal levels in the course of degeneration as well as regeneration.

Findings:
For this review, we compiled research articles pertaining to the role of cytoskeletal and other associated proteins involved in development of retina, which used various animal models. The effect of SNPs in the cytoskeletal proteins and their impact in retinal degeneration is also discussed.

Conclusion:
Studies describing the role of cytoskeleton in the anatomy and physiology of retina and its layers, although they are few, collectively provide an opportunity to understand retinal development in the context of cytoskeleton dynamics.

Full text

Critical ReviewsTM in Eukaryotic Gene Expression, 24(3):255-268 (2014)
255
1045-4403/14/$35.00 © 2014 Begell House, Inc. www.begellhouse.com
Cytoskeleton Dynamics in the Retina
Akshay Anand1,*,** Sridhar Bammidi1,** & Parul Bali2
1Department of Neurology, PGIMER, Chandigarh 160012, India; 2Department of Biophysics, Panjab University, Chan-
digarh 160014, India
* Address all correspondence to: Akshay Anand, Associate Professor, Neuroscience Research Lab, Department of Neurology, PGIMER, Chan-
digarh- 16001; Tel.: +91-991429090; Email: akshay1anand@rediffmail.com.
**Both are rst authors with equal contribution.
ABSTRACT: Background: Cytoskeleton is one of the essential forms of protein, important in the existence of
both eukaryotic as well as prokaryotic cells. Its transformation plays a vital role in cell division and intracellular
transportation by facilitating intracellular vesicular trafc. Among the various tissue types in the body, the neural
tissue exhibits the maximum heterogeneity, and hence the role of cytoskeleton at both developmental and func-
tional levels becomes paramount. Cytoskeleton dynamics have been established in the neural physiology, but only
at the level of axonal development and growth. Retina has not been adequately studied in the context of cytoskel-
etal proteins. Methods: We reviewed the last 10 years of literature with reference to the development, growth, de-
generation, and regeneration of the retina and the role of cytoskeleton in each aspect. We have focused on vari-
ous changes that the retina undergoes at the cytosolic and cytoskeletal levels in the course of degeneration as well
as regeneration. Findings: For this review, we compiled research articles pertaining to the role of cytoskeletal
and other associated proteins involved in development of retina, which used various animal models. The effect
of SNPs in the cytoskeletal proteins and their impact in retinal degeneration is also discussed. Conclusion: Stud-
ies describing the role of cytoskeleton in the anatomy and physiology of retina and its layers, although they are
few, collectively provide an opportunity to understand retinal development in the context of cytoskeleton dynamics.
KEY WORDS: microtubule, intermediate lament, microlament, cytoskeleton
I. INTRODUCTION
The anatomical, physiological, and functional as-
pects of life start from a single cell and can be best
understood through study at the cellular level. The
zygote, from the time of fertilization, undergoes
a series of divisions, physiological and structural
changes, and a vast array of differentiation and
compartmentalization at both inter- and intracellular
levels to reach its fate. Furthermore, these cellular
mechanisms occur in a series of well-dened and
accurate mechanisms of transcription and transla-
tion of protein macromolecules that govern physi-
ological and phenotypic expression. The cytosol
stores various codes for its behavior and metabolic
activities such as migration, translocation, nutrition,
phagocytosis, division, differentiation, and signal-
ing. Codes in the form of proteins can express struc-
turally as well as functionally. One of the essential
forms of proteins that is of utmost importance in the
existence of cell (whether eukaryotic or prokary-
otic) is the cytoskeleton. The cytoskeleton forms the
scaffold or skeleton at the cellular level in the cyto-
plasm of the cell. The cytoskeleton participates in
a variety of activities of the cell. These involve the
structural matrix of the individual cell and also in
the cellular movement through cilia,1 agella,2 and
lamellipodia.3 The cytoskeleton and its transforma-
tion play vital roles in cell division and intracellular
transportation by facilitating intracellular vesicular
trafc. The fact that the shape of different cell types
is determined by the organization of a network of
tubules (eventually termed cytoskeleton) was estab-
lished by Nikolai Koltsov, in 1903. The term cyto-
skeleton (French: cytosquelette) was rst introduced
in 1931 by the French embryologist Paul Wintrebert.
The association of cytoskeletal proteins with the
plasma membrane and its rearrangement has a very
interesting and well-elaborated role in cell locomo-
tion,4 translocation, and defense mechanisms of the

Critical ReviewsTM in Eukaryotic Gene Expression
Anand, Bammidi & Bali
256
cell, such as phagocytosis. The cytoskeleton has a
well-dened role in spindle formation in cell divi-
sion5 and also in pili formation during conjugation
of the prokaryotic cell.6
In the past two decades, researchers have in-
vestigated how the changes in the cell are governed
by the changes and modications in the cytoskel-
eton. The changes signify the overall reprogram-
ming of the cell while differentiating in the course
of tissue- or organ-level compartmentalization.7
The various subtypes of the cell and the cellular
organization, originating from the same germ lay-
er, are largely inuenced by their structural pattern,
which again is governed by cytoskeletal proteins.8
There has been increased emphasis on the role of
cytoskeleton in the development, growth, and dif-
ferentiation of the cell and its various types. The
cytoskeleton and its ultra-structural complexities
constitute not only the gross framework of a cell
but also the localization and functioning of its or-
ganelles.9
There has been a recent re-emergence of em-
phasis on the concepts of neural cytoskeleton. The
study of degeneration in the CNS and the neural
niche throughout different organisms has exposed
many myths about the nervous system. The dogma
that the CNS consists of immutable neurons whose
loss is permanent has been disproved by the ad-
vances in regeneration studies.10 The present evi-
dence clearly shows the presence and replacement
of neurons in the organism throughout life, at dif-
ferent levels. Among the various tissue types in the
body, the neural tissue exhibits the maximum het-
erogeneity,11 and hence the role of cytoskeleton at
both developmental and functional levels is very
important. The polarization of the neuron, con-
sisting of axonal and dendritic development and
arrangement has a series of mechanical and bio-
chemical cues in the rearrangement of the various
cytoskeletal proteins.12 The cytoskeleton plays a
major role in the formation and disruption of the
neural network, and in the formation, viability, and
degeneration of the neurons.
The retina, with its well-dened layers, has
been established as a good model with which to
study the CNS, due to its common ectodermal ori-
gin. The neural retina is extensively studied for its
nature, structure, function, and physiology and has
been manipulated in various ways in studying the
disease models.
This review focuses on the cellular arrange-
ment of neural retina and the pattern of different
cytoskeletal proteins participating in the develop-
ment, disease, and regeneration of the retina.
II. COMPOSITION OF THE CYTOSKELETON
Neurocytoskeleton is the protein skeleton distribut-
ed throughout the cytoplasm that serves such func-
tions such as providing mechanical strength, cell
shape, morphology, locomotion, and transportation
of organelle. Eukaryotic cells contain mainly three
types of cytoskeleton laments (microlament, in-
termediate lament, and microtubule), which show
dynamic behavior by their assembly and disassem-
bly. Microlaments are polymers of actin-forming
bundles and three-dimensional structures. The as-
sembly and disassembly of microlament is regu-
lated by the various actin-binding proteins. Inter-
mediate lament, mainly type-VI nestin, expresses
in dividing neuronal cells during development. Its
expression is higher in neuronal stem cells and di-
minishes in adult neurons. In adulthood, the injury
of CNS enhances expression of nestin.
Microtubule is the third component of cyto-
skeleton that is a polymer of tubulin protein. The
neuronal morphology shows a distinct arrange-
ment of microtubules. Axonal transportation re-
quires the assistance of the microtubule-associated
proteins kinesin and dynein. Kinesin is a coiled
structure with globular head that interacts with
microtubules to move toward the plus end while
dynein moves toward the minus end. Both kinesin
and dynein move opposite each other. Interruption
in axonal transport due to mutation in these mi-
crotubule-associated proteins is the key player in
mediating neuropathological states, as in the cases
of Alzheimer’s and Parkinson’s disease.
On close observation of the micrograph of a
cell, a dense disorganized network of laments
can be seen. There is a marked distinction in the
regions of cell protrusions or in the junctions of

Volume 24, Number 3, 2014
Cytoskeleton Dynamics in the Retina 257
two adherent cells. These regions have laments
concentrated into bundles. These bundles further
spread out into the interiors of cells as a network
of laments. Bundles and networks form the most
common arrangements of cytoskeletal laments
in a cell. Bundles and networks structurally dif-
fer in the organization of laments. The laments
are packed closely into arrays in bundles, which
are arranged loosely in a crisscross in the case of
networks. Networks further show planar (two-di-
mensional) or net-like arrangements as in nuclear
and plasma membrane, and three-dimensional ar-
rangements, which, for example, gives cytosol its
gel-like properties. In each of these arrangements,
the laments are held together by various cross-
linking proteins.
Actin laments are responsible for the distinct
shape of any cell. The organization of the actin
laments and their associated proteins that con-
nect the microlaments to the membrane govern
the cell morphology. On attachment to a bundle of
laments, the membrane acquires the shape of a
microvillus.
III. RETINA AS A MODEL FOR STUDYING THE
CENTRAL NERVOUS SYSTEM
As an extension of the central nervous system, the
retina, with its well-studied layers, provides a rela-
tively simpler and convenient tool for investigating
the complex central nervous system. Relatively, it
can be easily manipulated from the outside (unlike
brain), making it feasible to test the homing and re-
generative potential of various types of stem cells,
pharmacological compounds, and neurotropic fac-
tors. The common ectodermal origin of retina and
brain during embryonic development makes retina
an excellent experimental tool for studying the
CNS. The similarity in cellular architecture of ret-
ina across various species of experimental organ-
isms has further led to a wide range of organisms
being used for retina-based investigations.
Mammalian retina is composed of seven dis-
tinct layers consisting of different neurons and
glia: a ganglion cell layer, amacrine cells, bipolar
cells and muller glia in inner nuclear layer that act
as inter neurons, rod and cone photoreceptors in
the photoreceptor layer (also considered the outer
nuclear layer), and the retinal pigment epithelial
layer. The axons of ganglion cells traverse through
the optic nerve and connect to the visual cortex of
the brain. These different and highly diverse cell
types arise from the population of retinal progeni-
tor cells, which are a population of quiescent and
actively dividing cells. Recent research has sought
to elucidate the mechanisms controlling the com-
plex processes of neuronal differentiation from the
same source.
IV. NEURONAL/AXONAL DEVELOPMENTAL
CASCADE (FIG. 1)
The cytoskeleton plays a very vital role in neuronal
development. The polarity of the neural cell is es-
tablished by the activity and expression of various
intermediate and associated proteins. Structurally,
neurons have a polarized morphology with two
types of processes, i.e., a long and slender axon and
several short and tapered dendrites. Neurons are not
in the polarized morphology in the course of their
development. They are a result of asymmetrical
cell division of the neural precursors, which is con-
trolled by cytoskeleton.13 The present understand-
ing of the role and contribution of cytoskeleton-
associated proteins in the cellular mechanisms is
not enough. One of the examples that can best sub-
stantiate this fact is the structural proteins like tau,
a microtubule-binding protein that has been exten-
sively studied in the eld of neurodegenerative dis-
eases. Its molecular mechanisms and functions are
well elaborated, but the function within the normal
or diseased neurons has not yet been elucidated.14,15
The study of cytoskeleton development in neurons
can be best reviewed by examining the dynamics
of axonal growth, although the available literature
is relatively scarce. Axons, among all cell types,
are the longest in mature and full-grown animals.
Axons propagate (electrical) messages toward the
synaptic connections in the target cells several
meters away through the neuronal cell body. The
unusual architecture of the axonal shafts is main-
tained by the bundles of parallel microtubules.

Critical ReviewsTM in Eukaryotic Gene Expression
Anand, Bammidi & Bali
258
This provides support for structural and long-dis-
tance transport.16 Microtubules in axons are not
continuous and, unlike most cells, are not attached
to the centrosome for their organization.17 The plus
ends of the microtubules mostly point distally.18–20
Short actin laments, organized by spectrin (scaf-
folding protein) into repetitive rings, line the cor-
tex of axon shafts. This linkage to the axonal cell
membrane is again brought about by a second set
of proteins called ankyrins.21 Ankyrins are the an-
chorage and nodal proteins which show robustness
in maintaining the axonal integrity as discussed in
the mouse model of cerebral hypoperfusion.22 The
specic size and shape (varying diameters) of the
different classes of axons are regulated further by a
number of intermediate laments.23 Axons have a
very long span of life, and the dying neurons are a
characteristic feature of the ageing brain.24 Severe
disorganization in the axonal microtubule bundles
are believed to trigger the axonal loss.25,26 The
structural microtubule-binding proteins such as tau
and MAP1B are believed to stabilize the axonal
bundles.27 Moreover, continuous polymerization of
microtubules has been reported in mature axons,
maintaining the steady turnover of microtubules.28
The structural plasticity of neurons demonstrated
by the shortening and lengthening,29–31 the branch-
ing, the re-growth, and the axonal pruning that oc-
curs for sizing the overgrowth,32 are all important
in the regeneration of the damaged neurons. This
regeneration and its associated processes are very
important in synapse formation related to learning
and memory.33,34 The formation of small neuritic
protrusions on the neuronal cell bodies mark the
onset of the formation of axons. The migration of
microtubules by pushing beyond the cell body and
the removal of barrier function is dynamically as-
sisted by the rearrangement of the cortical actin
cytoskeleton35–37 and the formation of lopodia for
extension of microtubules into the membrane.38,39
In a vertebrate neuron, an axon is formed from the
extension of a single neurite out of the many neu-
rites formed on the cell body simultaneously. This
selective process is termed neuron polarization,
and it involves a signaling cascade of cell polar-
ity factors for targeted and specic localization,40
which also requires specicity of stabilization of
microtubules of the selected neurite.41,42
FIG. 1: Hematoxylin & Eosin stain of mouse eye cryosection

Volume 24, Number 3, 2014
Cytoskeleton Dynamics in the Retina 259
After their establishment, axons follow a ste-
reotype in growth toward their target cell. This
process is generated and executed by growth cones
at the axon tips. The spatio-temporal pattern of
chemical signaling guides the growth cones to-
ward their specic target tissue.43 These guiding
signals have been identied and well elaborated
in their mechanism,44,45 controlling the behavior
of growth cone. The morphogenetic movements,
such as growth velocity, pausing, turning, retrac-
tion or collapse downstream of guidance signal-
ing, are achieved by the particular arrangement of
growth cone actin and microtubule cytoskeleton.46
The growth cones can be divided into three
different zones: the actin-rich peripheral zone; cen-
tral zone, rich in microtubules; and the transition
zone of microtubule and F-actin overlap. The mi-
crotubules spread into the peripheral zone starting
from the center of the growth cones comprising the
axonal tips. The peripheral zone consists of actin-
rich membrane protrusions, which include nger-
like lopodia and veil-like lamellipodia. Filopodia
(containing parallel F-actin bundles) and lamelli-
podia (containing F-actin lattices), after guidance
from the peripheral zone, present their receptors
on their surface. The continuous aggregation and
assembly of actin laments at the leading edge
constantly changes the shape. These laments ow
back in a reverse orientation, depolymerize, and
recycle further in the transition zone. Filopodia
then reach into the growth cone and behave as sig-
naling and adhesion sensors.47–50 The microtubule
behavior in guided axon extension is essentially in-
uenced by this series of events. Lamellipodia acts
as site of substrate adhesion and myosin-II–driven
contraction and force generation.
In another model of growth-cone guidance, the
stabilization of the single microtubule in the pe-
ripheral zone is triggered by external signals,51,52
which are further joined by a bulk of microtubules
from the center, giving rise to a new axonal seg-
ment. The nal step in elongation is the relocation
of the lamellipodia and lopodia in the axon tip.
Microtubules have a crucial role in the implemen-
tation of axon extension in this model, while the
directionality of the extension is achieved by F-
actin. Notably, therefore, the blocking of microtu-
bule dynamics results in the inhibition of axonal
growth.53–55 On the other hand, the inhibition of
actin polymerization either by pharmacological or
by genetic approaches had no suppression of the
axonal growth, in vitro or in vivo. Such interfer-
ence showed some aberration in the turning and
path-nding dynamics of the growth cone.56,57–61
In this regard, it is interesting to note that the
role of F-actin is not always essential in the di-
rectional movement of growth cones. Many sig-
naling mechanisms are targeted to microtubules.
Phosphorylation of some signaling molecules, like
MTBPs APC1, MAP1B, and/or CLASP, mediated
by GSK3 directly regulate the stability of microtu-
bules in growth cones.62–64
The generation of neurons and their differenti-
ation followed thereafter is regulated by a number
of intrinsic and extrinsic factors. The proteins be-
longing to the basic helix–loop–helix (bHLH) fam-
ily like Neurogenin 2 (Ngn2) or Mash (mammalian
achaete-scute complex)-1 or Math (mammalian
atonal homolog)-1 are some of the pro-neural genes
reported in neurogenesis. These mechanisms are
all regulated necessarily by an array of distinct
signaling molecules like Notch signaling, trans-
forming growth factor (TGF)-β/bone morphogenic
protein (BMP) and broblast growth factor (FGF).
The formation and generation of different types of
neuronal types from their progenitors is regulated
by regulatory proteins. The cell cycle is switched
off to regulate number of cells post neurogenesis.65
The regulation of cell cycle in this context is ex-
tremely important for the development of the ner-
vous system. The patterning of neural tissue is de-
termined by SRY (sex-determining region Y) box,
Sox, and the Pax (paired box) family of proteins.
Pax6 (a paired-box transcription factor), which is
highly conserved among vertebrate development,
is responsible for the overall patterning of the em-
bryonic nervous system.66 It directly regulates the
expression of Ngn2 and also upregulates the ex-
pression of cell adhesion molecules as well as the
cell-cycle inhibitors such as p27.67 This eventually
links together the differentiation and the withdraw-
al from the cell cycle. Again, the maintenance of

Critical ReviewsTM in Eukaryotic Gene Expression
Anand, Bammidi & Bali
260
neural cells in the undifferentiated state is brought
about by Sox1, Sox2, and Sox3 proteins via sup-
pression of neurogenesis.68
Coordination of transcriptional program in the
neurogenesis occurs also at the epigenetic level.
The overexpression of HDAC4 (histone deacety-
lase) in granule neurons, shields them from apop-
tosis without undergoing the PI3K or ERK/MAPK
pathways. The phenomenon of neuroprotection is
brought about by blocking the abnormal progres-
sion of cell cycle by the inhibition of CDK1 (cell
cycle kinase).69
The overall transcriptional programs and epi-
genetic regulation is mediated by signal transduc-
tion pathways. The signal transduction pathways
are further regulated by the events in cell cycle.
The essential role of Notch signaling in neurogen-
esis has been demonstrated both in vitro70 and in
vivo.71 It inhibits the neural gene expression later-
ally, and hence limits the number of differentiating
neurons. In vertebrate neurogenesis, the notch and
delta signaling are restricted only to some stages in
the events of cell cycle.72
V. THE GROWTH AND DIFFERENTIATION
DYNAMICS
After exit from the cell, the generation of intri-
cate networks of the microtubule in neurons and
the extensions has to be brought about by a well-
orchestrated differentiation program. This is also
brought about by the timed entry and exit of the
cell cycle. While studying the mutations in dynac-
tin in a zebrash model, a premature cell cycle exit
and acceleration of neurogenesis was exhibited by
the retinal progenitors. The cell signaling also gov-
erns cell fate. The defects in trafcking of Notch
to the apical region of retinal progenitors has been
reported in affecting the early cell fates at the ex-
pense of late ones.73
Neurons migrate to their functional location
from the site of their origin via polarized move-
ment of the cytoskeleton.74 In the case of neuro-
genesis in the retina, a specic pattern is followed.
During the development of the retina, retinal gan-
glion cells are the rst to differentiate, then cones,
followed by horizontal cells. After these, the rod
cells and the amacrine cells are formed. The last
cells to differentiate are the bipolar cells. Retinal
neurons in mammals differentiate before birth.75–77
Neurogenesis of retinal ganglion cells is deter-
mined intrinsically. A number of genes have been
identied in this process of neurogenesis.78 The
deletion of Pax6, Barhl2, or Math5 reduces and/
or blocks the differentiating RGCs in mice.79, 81–83
VI. CYTOSKELETAL PROTEINS IN THE RETINA
AT DIFFERENT LEVELS
From the beginning, understanding the cell-fate
specications and mechanisms has remained a
challenge in the study of retinal development. The
Rho family plays a key role in cytoskeletal arrange-
ments in the RPE in the TGF-β1–induced pathway.
It has been demonstrated experimentally that treat-
ment with TGF-β1 induces cytoskeleton reorga-
nization, expression of α-SMA, an increase in the
phosphorylation of ERK, Smad, AKT, and activa-
tion of Rac1 and RhoA. These mechanisms are in-
hibited by treatment with Rho inhibitors. TGF-β1
also increases LIMK and colin phosphorylation.
Based on these observations, it has been well pro-
posed and established that the cytoskeletal actin re-
arrangement in human RPE is induced by TGF-β1
via the Rho GTPase-dependent pathways modulat-
ing LIM kinase and colin activity.84
The role of cytoskeleton proteins can also be
well elaborated in the phagocytosis mechanism of
the RPE. The shed photoreceptor outer segments
are phagocytosed by the cells of the RPE (retinal
pigment epithelium) through a mechanism that
involves αv integrins upstream of Rho GTPases
and tyrosine kinases. Retinal pigment epithelial
cells have the tendency to activate and redistribute
Rac1, but they do not do the same with RhoA or
Cdc42 during phagocytosis. The particle engulf-
ment, in this case by the RPE, is prevented when
dominant-negative Rac1 is overexpressed as well
as when Rac1 expression is decreased. It has been
further experimentally demonstrated that αvβ5
integrin and its ligand MFG-E8 (milk fat globule
EGF factor-8), are required for the activation of

Volume 24, Number 3, 2014
Cytoskeleton Dynamics in the Retina 261
Rac1 during phagocytosis, but they do not involve
the receptor tyrosine kinase MerTK. FAK inhibi-
tion hinders the tyrosine kinase signaling down-
stream of αvβ5 toward MerTK. This specic hin-
drance does not prevent Rac1 activation nor does
it prevent the recruitment of F-actin during phago-
cytosis. Inhibition of Rac1 does not effect FAK or
MerTK activation. Therefore, activation/recruit-
ment of MerTK and F-actin via FAK and Rac1,
respectively, require MFG-E8–ligated αvβ5 integ-
rin. These pathways have independent activation
and are essential for clearance phagocytosis. The
major source of endothelin-1 (ET-1) at the blood
retinal barrier is the retinal pigment epithelium
(RPE). Thrombin can act on the RPE and can prob-
ably injure the blood retinal barrier. The activation
of protein kinase C and the transient intracellular
calcium mobilization brought about by thrombin
probably play a minor role in endothelin-1 synthe-
sis in the retinal pigment epithelium.85
The outgrowth of the neurites, and eventually
the growth cone repulsion, are mediated by a mem-
ber of the subfamily of cell adhesion molecules,
i.e., protein tyrosine phosphatase (RPTP). PTPμ
acts as a permissive substrate for the growth of
retinal ganglion cell (RGC) neurites while it inhib-
its the growth of temporal retinal ganglion cells.
In temporal neurons, Rac 1 acts as the regulating
switch between the permissive and repulsive re-
sponses to PTPμ. For the repulsion of temporal
neurons, Rac 1 needs to be inhibited. Being an
inhibitor for nasal growth, the activity of Cdc42
is essential for both permissive and repulsive re-
sponses to PTPμ. However, inhibition of Rac1 ac-
tivity is the key to PTPμ-dependent repulsion in
temporal RGC neurons.86
Studies on the activity of actin/colin proteins
have shown an active role in axonal extension87–91
and have an implications in the regulation of neu-
rotrophins on lopodial responses.92,93 A recent
study showed that the direct role of actin/colin
proteins in response to bone morphogenic proteins
displays growth-cone guidance in the spinal neu-
rons of Xenopus laevis.94 Another very important
member of this group of molecules is the Rho fam-
ily of GTPases. Of the various molecules amongst
the small GTPase related to Rho, Rac1 is involved
in the regulation of cytoskeletal dynamics and is
critically involved in the development of neuron,
axonal growth, and cell survival. Rac1 in its con-
stitutively active form has been shown to promote
the growth of axons in inhibitory environments of
in vivo95 as well as in vitro96 models. It has been
reported to promote survival in vivo as well.97, 98 In
studies involving the developing chick retina, NF-
M, i.e., mid-size neurolament99–101 and a chick-
specic MAP (microtubule associated protein),
have been extensively studied to identify ganglion
cells and ganglion precursor cells.102 It has been
demonstrated that the expression of MAP(RA4)
and NF-M proteins in the chick retina is not re-
stricted to ganglion cells alone as otherwise thought
and accepted. Rather, it is established that these
proteins are expressed transiently in a develop-
mentally regulated manner by all neuronal retinal
progenitors.103 The Lim-kinase (LIMK) proteins
essentially regulate the actin cytoskeleton. It is
therefore believed that they control many process-
es, including cell cycling, cell integrity, migration
and axonal guidance, during development.104 The
loss of LIMK1 resulted in defects in the neuronal
growth cone, dendritic spines, and the organiza-
tion of actin. These results were demonstrated by
the different experiments in-vitro and in-vivo and
resulted in abnormal long-term potentiation of the
hippocampus and synaptic changes.105 Glycogen
synthase kinase (GSK3) regulates a large number
of transcription factors, including β-catenin, c-Jun,
and CREB (cAMP response element-binding pro-
tein), to name a few.106 A study involving the regu-
lation of transcription factors by GSK3β shows the
involvement of kinase essentially in the pruning of
developmental axon both in vitro and in vivo.107 A
wide range of cytoskeleton dynamics and several
types of purinergic receptors are controlled by ex-
tracellular ATP. An intracellular Ca(2+) increase,
induced by the ATP, follows colin rod formation.
Cells that express another variant of colin (un-
phosphorylatable) do not exhibit ATP-induced co-
lin rod formation. The formation of colin rods is
induced by the calcineurin-dependent dephosphor-
ylation of colin induced by the inux of Ca(2+)

Critical ReviewsTM in Eukaryotic Gene Expression
Anand, Bammidi & Bali
262
through P2X receptors. This pathway may essen-
tially elucidate how ATP effects development, de-
generation, and injury of a neuron.108
VII. ROLE OF CYTOSKELETON IN NEURAL/
NEURONAL DEVELOPMENT IN ANIMAL
MODELS
The study of neuronal polarity and synaptogenesis
and associated proteins has been studied in Dro-
sophila, Caenorhabditis elegans, Xenopus, and
zebrash models. A study of Drosophila demon-
strated that when Cyp, a cytoplasmic FMRP-
interacting protein, is mutated, it inhibits the l-
amentous actin (F-actin) to assemble, which is a
key regulating aspect of synaptogenesis. The study
also proposed that Cyp inhibits actin assembly
as well, regulating the development of synapse as
well as endocytosis.109 Intraagellar transport (IFT)
is essential in developing the ciliated sensory outer
segments (OS), which sequester a number of pro-
teins in the vertebrate phototransduction machin-
ery. Rho GTPases are essential regulators of cyto-
skeletal reorganization in the case of Drosophila as
well. Colin is a protein responsible for the depo-
lymerization of actin, very essential in the process
of neurogenesis. It is regulated by the inhibition
of LIM kinase and activation of Slingshot phos-
phatase. During Axon growth, the genetic analysis
of the different pathways have revealed conver-
gent and divergent pathways from Rho GTPases
to the cytoskeleton achieved by biases in pathway
selection.110 In one of the experiments involving
Drosophila, neuronal cells in vitro demonstrated
that the regulation of cellular actin dynamics, the
deciency of pantothenatekinase (PANK), a cata-
lyst that converts pantothenic acid to co-enzyme
A, leads to abnormal organization in the F-actin. A
reduction in in the activity of PANK up-regulates
the phosphorylated form of colin, which is a con-
served protein responsible for the severing of actin
lament.111
However, the mechanism of the cytoskeletal
proteins and the associated proteins/molecules have
been studied in the C. elegans model. In a study
using C. elegans, it was demonstrated that during
axonal outgrowth, the Arp2/3 complex regulates
the initiation of growth cone lopodia.112 The axon
guidance in C. elegans has been shown to be af-
fected by UNC-115/abLIM and UNC-34/Enabled,
which are actin modulatory proteins.113–115 Enabled
essentially regulates the formation of lopodia in
neurons. It probably acts by blocking the actin-
capping activity, resulting in long lament growth
in lopodia.116,117 Some other studies show that En-
abled might express the anti-capping independent
roles118 involved in lopodia formation.11 9 OSM-3,
belonging to the family of homodimeric kinesin 2
motor, has an essential role in a few sensory cil-
ia.120,121 Kif17, a homologue for the OSM-3 gene in
vertebrates, is known to be responsible for the traf-
cking of dendrites in neurons. Its role in the pro-
cess of ciliogenesis has not yet been dened. Kif17
is widely expressed in the zebrash nervous sys-
tem and retina.122 Sufcient evidence has proven
the role of ADF (actin depolymerizing factor) and
colin in the regulation of axonal growth and actin
dynamics. The expression of ADF/colins is high
in neuronal growth cones, especially in the areas
where there is a high rate of actin disassembly.123
Furthermore, the overexpression of these proteins
in a Xenopus model showed a nearly 50% increase
in the length of neurites.124
VIII. SNPS IN CYTOSKELETON IMPAIRMENT
Various molecules have been studied to acquire
deeper mechanistic understanding of retinal de-
generation. The mutations and/or deletions or
single nucleotide polymorphisms seen in the mol-
ecules during disease or degeneration lack proper
co-relation with the cytoskeleton. The dynamics of
the cytoskeleton plays a critical role in neural-crest
cell migration. One important molecules discussed
from the beginning of this review is colin. The
dynamics of cytoskeleton components, especially
in neural tube development, is regulated by colin.
Colin is encoded by the gene C1, and its poly-
morphisms have been investigated and established
as a risk factor of spina bida. In a population-based
control study in California of non-Hispanic white
infants with spina bida, ve SNPs in the gene

Volume 24, Number 3, 2014
Cytoskeleton Dynamics in the Retina 263
enhanced disease risk by greater than two-fold.125
The regulation of the cytoskeleton is essentially
brought about by protein–protein interactions.
The interaction of integrin and actin is regulated
by the FLNA gene. Mutations in the FLNA gene
result in congenital malformations in craniofa-
cial structures.126 In one genetic study of two sib-
lings having an unusual set of disease conditions
(i.e. optic atrophy, early-onset spastic paraplegia,
and neuropathy), one homozygous c.316C>T(p.
R106C) variant in the Trk-fused gene (TFG) was
the only plausible mutation upon linkage analysis,
genome-wide SNP-typing, and exome sequencing.
The characterization of this protein did not yield
any self-assembly into an oligomeric complex,
which is essential for the normal functioning of
TFG. The inhibition of TFG alters endoplasmic
reticulum (ER) morphology while slowing down
its protein secretion. This results in the disruption
of peripheral tubules and eventually the ER col-
lapses over the cytoskeleton beneath it. This study
highlights the link between the involvement of
cytoskeleton through altered ER architecture and
neurodegeneration.127 TUBA8 gene, encodes a
variant of α-tubulin with unknown function and
it regulates the function of microtubules during
the migration of cortical neurons. One study has
reported that the mutation of this gene results in
generalized polymicrogyria along with optic nerve
hypoplasia (PMGOH). This study suggests the role
of TUBA8 in the regulation of brain development
in mammals.128 Mutations in PFN1, a gene encod-
ing the actin monomer-binding protein prolin 1,
were recently reported in 1–2% of familial amyo-
trophic lateral sclerosis (ALS) patients. Two rare
non-synonymous variants (E117D and E117G)
have been identied in sporadic ALS patients.129
Cerebral PGV (proliferative glomeruloid vascu-
lopathy) is a severe autosomal recessive disorder
of brain angiogenesis that results in abnormal
thickening and unusually perforated vessels which
lead to hydranencephaly. Mutation and deletion in
the FLVCR2 gene, which encodes a 12-transmem-
berane domain constituting a putative transporter,
results in the absence of alpha-smooth muscle ac-
tin, suggesting a decit in pericytes.130
IX. CONCLUSION
The eld of retinal neural cytoskeletal dynamics
in the growth and development of the vertebrate
retina needs denition. This review highlights the
role of retinal architecture and the involvement of
cytoskeletal proteins in the growth and differentia-
tion of the neuronal cells, thus indicating the need
for translational research. Therefore, the study of
retinal development, growth, and differentiation is
likely to contribute signicantly in the context of
functional retinal repair. The mechanisms underly-
ing differentiation and compartmentalization pro-
vide new insights into retinal development. In this
regard, regenerative approaches in stem cell trans-
plantation need to integrate such methodologies to
account for poor results and irreproducible data.
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