Clin Genet 2007: 72: 296304 # 2007 The Author

Printed in Singapore. All rights reserved Journal compilation # 2007 Blackwell Munksgaard
CLINICAL GENETICS
doi: 10.1111/j.1399-0004.2007.00888.x

Developmental Biology: Frontiers for Clinical Genetics

Section Editors:
Roderick R. McInnes, email: mcinnes@sickkids.ca
Jacques L. Michaud, email: jacques.michaud@recherche-ste-justine.qc.ca

Lissencephaly and LIS1: insights into the

molecular mechanisms of neuronal migration
and development
Wynshaw-Boris A. Lissencephaly and LIS1: insights into the molecular A Wynshaw-Boris
mechanisms of neuronal migration and development. Departments of Pediatrics and Medicine,
Clin Genet 2007: 72: 296304. # Blackwell Munksgaard, 2007 UCSD School of Medicine,
La Jolla, CA, USA
Lissencephaly is a severe human neuronal migration defect characterized
by a smooth cerebral surface, mental retardation and seizures. LIS1 was
first gene cloned in an organism important for neuronal migration, as it
was deleted or mutated in patients with lissencephaly in a heterozygous
fashion. Studies in model organisms, particularly Aspergillus nidulans, as
well as those in the mouse, have uncovered an evolutionarily conserved
pathway that involves LIS1 and cytoplasmic dynein. This pathway codes
for proteins in a complex with cytoplasmic dynein and positively Key words: 14-3-3e dynein
regulates its conserved function in nuclear migration. This complex interkinetic nuclear migration isolated
appears to be important for proliferation and neuronal survival as well as lissencephaly sequence LIS1
neuronal migration. One of the components of this complex, NDEL1, is lissencephaly MillerDieker
a phosphoprotein that is a substrate for CDK5 (or CDK2 in fibroblasts) syndrome NDEL1 neuronal
and Aurora-A, two mitotic kinases. CDK5-phosphorylated NDEL1 migration nuclear migration
binds to 14-3-3e, which protects it from phosphatase attack. Corresponding author: Professor Anthony
Interestingly, 14-3-3e is located 1 Mb from LIS1 and is heterozygously Wynshaw-Boris, MD, PhD,
deleted with LIS1 in patients with a severe form of lissencephaly, Miller Departments of Pediatrics and Medicine,
Dieker syndrome. Mouse models confirm that 14-3-3e plays an important UCSD School of Medicine,
role in neuronal migration, and mice that are double heterozygotes for La Jolla, CA 92-93-0627,
mutations in Lis1 and 14-3-3e, display more severe neuronal migration USA.
defects. The identification of LIS1 as the first lissencephaly gene, and the Tel.: 11 858 822 3400;
first gene required for neuronal migration has revealed the importance of fax: 11 858 822 3409;
the regulation of cytoplasmic dynein in the control of neuronal migration e-mail: awynshawboris@ucsd.edu
by modulating nuclear migration in a pathway conserved in virtually all Received 6 July 2007, revised and
eukaryotes. accepted for publication 10 July 2007

The human brain is the most elaborate structure
known and comprised of an integrated network of
more than 100 billion neuronal cells. The forma-
tion of this intricate neuronal network requires the
precise choreography of neurogenesis, neuronal
migration, pruning and synaptogenesis during
development. The mammalian cerebral cortex is
highly patterned and divided into anatomically
distinct, functionally specialized areas for cogni-
tive, executive, motor, somatosensory auditory
and visual functions. These different cortical areas
display a precise connectivity to provide specific
inputs and outputs to and from the cerebral
cortex.
Much has been learned about neuronal devel-
opment by studying the variations from the basic
pattern of forebrain organization in different
mammalian organisms, including the human.
One of the most striking abnormalities of human
neuronal migration is lissencephaly (or smooth
brain), a term originally used to describe the
smooth cerebral surface of lower mammals.

296

Lissencephaly of the human brain is a severe
malformation in which the brain has a smooth
cerebral surface rather than the characteristic gyri
and sulci that make the human and primate brain
instantly recognizable. Lissencephaly can be
further specified as agyria (absence of gyri) or
pachygyria (reduced numbers of broadened gyri).
In this review, I will focus on LIS1, the first
neuronal migration gene cloned in any organism.
The discovery of LIS1 as the causative gene in
a subset of human lissencephaly led to the
elucidation of a complex conserved from yeast
to human that regulates the activity of cytoplas-
mic dynein. This complex is crucial for neuronal
migration, but more recent data indicates that it is
also broadly important during neurogenesis and
for neuronal survival during development. The
reader is referred to recent reviews for more com-
prehensive insights into the control of neuronal
migration (13) and LIS1 function (4).
Neuronal migration during cortical
development
Neuronal migration has been studied extensively
for over 30 years in diverse mammalian species
from the mouse to human. The basic sequence of
events that occurs during cortical development is
shared by all of these species, although the timing
of events is species dependent. For example, these
events occur between 5th and 22nd gestational
weeks of human development, while in the mouse
they occur between embryonic day 11 (E11) and
19 (E19).
Prior to E11 in the mouse, the neural tube
consists of a single layer of pseudostratified
neuroepithelial stem cells that proliferate rapidly.
Each of these stem cells stretches from the apical
to the basal surface of the neuroepithelium, and as
the cell progresses through the cell cycle, the nuclei
display interkinetic nuclear migration (Fig. 1).
Nuclei divide at the apical surface and synthesize
DNA at the basal surface of the neural epithelium.
Consequently, nuclei from cells in G1 migrate
from the apical to basal surface, while nuclei from
cells in G2 move from the basal to apical surface.
At about E11, shortly after closure of the rostral
neural tube, the neuroepithelial stem cells further
differentiate into a more restricted progenitor,
radial glial progenitor cells, and in a subpopula-
tion of these cells, the daughter cells of the radial
glial progenitors become post-mitotic, immature
neuroblasts while in and shortly after leaving the
ventricular zone (VZ). Young neurons begin to
move out from the VZ at about E12 radially along
the extensions of the radial glial progenitors that
act as both progenitor and scaffold for migration
Lissencephaly and LIS1
(a)
BASAL
APICAL
NESCs G1 S G2 M NESCs
(b)
CP
IZ
VZ
RGs G1 S G2 M RGs
Fig. 1. Interkinetic nuclear migration in neuroepithelial
stem cells and radial glial progenitor cells. (a) Neuro-
epithelial stem cells (NESC) exist as a pseudostratified
epithelium with each cell attached at both apical and basal
surfaces. Interkinetic nuclear migration occurs throughout
the length of the cell, with nuclei from cells in G1 moving
from the apical to the basal surface (arrow above G1),
where cells synthesize DNA (S). Nuclei from cells in G2
move from the basal to apical surface (arrow above G2),
where the cells divide (M), to produce two daughter NESCs.
(b) Interkinetic nuclear migration occurs in the more
restricted radial glial progenitor cells (RG), but this
migration is confined to the ventricular zone (VZ), and
excluded from the intermediate zone (IZ) and cortical plate
(CP). Note that RG cells divide at the apical surface.
(Fig. 2). The first migrating cells give rise to the
marginal zone or layer 1, which is made up largely
of the earliest differentiated neurons of the cortex,
the CajalRetzius cells. Between E13 and 19 in the
mouse, neurons generated in the VZ migrate
across the intermediate zone toward the cortical
plate at the pial surface and become the projection
neurons of the cortex. As a result, several
embryonic laminas are established over the next
few days. Interestingly, the proliferating radial
glial progenitors display interkinetic nuclear
migration, similar to the neuroepithelial stem
cells, but only in the VZ, a proliferative zone at the
apical surface (Fig. 1). The embryonic cortical
plate forms layers 2 through 6 in the post-natal
period. Classical studies have established that the
cerebral cortex is formed by an insideout
migration of cells from the VZ. The early arising
neuroblasts that migrate first to the cortical plate
occupy the deepest cortical layers in the adult
(layers 4, 5 and 6). The later born migrating young
neurons migrate past the earlier established
cortical plate cells to occupy more superficial
layers (layers 2 and 3) of the adult cortex.
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Wynshaw-Boris
3V
Cortex
HIP HIP LV NCX
LGE
Caudate, MGE
putamen
Globus
pallidus
Fig. 2. Direction of migration of neurons during mamma-
lian cortical development. The schematic represents a coro-
nal section through an E12-13 mouse cortex. Arrows
represent radial migration from the hippocampus (HIP,
green), cortex (aqua), lateral ganglionic eminence (LGE,
red) to the caudate, putamen and medial ganglionic
eminence (MGE, black) to form the projection neurons.
Additionally, cells migrate tangentially from the LGE (blue)
to the caudate, putamen and MGE (pink) to form the
inhibitory interneurons in the cortex. These tangentially
migrating cells interact with radially migrating cells born at
the same time in the intermediate zone of the cortex, and
then migrate together to the proper layer in the cortex. LV,
lateral ventricle; NCX, neocortex; 3V, third ventricle.
In addition to radial migration of the projection
neurons of the cortex, the inhibitory interneurons
migrate tangentially from the medial ganglionic
eminences (MGE) and lateral (LGE) ganglionic
eminences (Fig. 2). As one example of the tight
choreography of neuronal migration, cells born
in the cortical VZ meet cells born at the same time
in the MGE and LGE in the IZ, where they in-
teract and migrate together to the appropriate
cortical layer.
Human lissencephaly
Lissencephaly is one of the best characterized
and studied human genetic malformations of the
cerebral cortex. The primary defect underlying the
smoothening of the brain of lissencephalic pa-
tients is defective neuronal migration. The inabil-
ity of post-mitotic neurons to reach their final
destination and correctly populate the cortical
plate of the cerebral cortex consequently leads to
abnormal cortical thickness and reduced or absent
gyri and sulci of its surface.
Classical lissencephaly defines a subgroup of
human neuronal migration disorders character-
ized by generalized agyria/pachygyria, four
abnormal cortical layers, enlarged ventricles,
298
and generalized neuronal heterotopias (5). There
are two major types of classical lissencephaly:
isolated lissencephaly sequence (ILS) and Miller
Dieker syndrome (MDS). ILS is a heterogeneous
disorder consisting of variably severe lissence-
phaly with no other major malformations such as
craniofacial dysmorphism. Children with ILS are
also severely retarded and typically have epilepsy
(6). MDS consists of more severe lissencephaly
than ILS patients, characteristic facial anomalies,
and occasionally other malformations. Children
with MDS are severely retarded and suffer from
epilepsy. Craniofacial dysmorphism includes mi-
crocephaly with bitemporal narrowing, a high
forehead, a small nose with anteverted nares, thin
vermilion border, and micrognathia (7). The
disorder is invariably fatal in early childhood.
The major pathology in both disorders is usually
considered to be in the cerebral cortex, but
cerebellar and olivary heterotopias have been
described in MDS patients as well.
MDS (100%) and some cases of ILS (40%) are
the result of haploinsufficiency at human chromo-
some 17p13.3, with visible or submicroscopic
deletions detectable by fluorescence in situ hybrid-
ization (8, 9). The LIS1 gene was cloned from this
region (10). LIS1 was disrupted in an ILS patient
with a translocation, and several other key MDS
patients (9). Point mutations and an intragenic
deletion in LIS1 were identified in ILS patients who
showed no gross structural chromosomal rear-
rangements, confirming that mutations or dele-
tions of the LIS1 gene are responsible for classical
lissencephaly in ILS and MDS patients (11, 12).
The more severe neuronal migration phenotype
and facial dysmorphisms displayed by MDS
patients suggest that genes other than LIS1, such
as 14-3-3e are responsible for these phenotypes.
Mouse models for Lis1 functional studies
Mouse models for these disorders have aided in
the understanding of the function of LIS1 and the
pathways associated with it during brain devel-
opment. Although human studies demonstrated
that LIS1 mutations are responsible for lissence-
phaly, it is not possible to study the neuronal
migration defects that occur embryologically
during human development. To examine the
consequences of dosage reduction and complete
deficiency of Lis1 in vivo, we produced two
knockout and one conditional knockout Lis1
mutant alleles by gene targeting in the mouse (13).
The two knockout alleles (one from germline
Cre-mediated deletion of the conditional knock-
out allele) are nulls, while the conditional allele
is hypomorphic, because of the disruption of

transcription or splicing by the insertion of PGKneo
in intron 3. By mating mice with various Lis1
alleles, mice were produced with graded reduction
in LIS1 dosage. Mice with decreased levels of Lis1
exhibited dose-dependent disorganized cortical
layers, hippocampus, cerebellum and olfactory
bulb because of cell autonomous neuronal migra-
tion defects (13) and are a good model for the
human disorder. Complete loss of LIS1 results in
peri-implantation lethality, a result confirmed in
another Lis1 knockout (14), demonstrating that
Lis1 is an essential gene.
Further studies demonstrated impairments of
motor coordination and cognition in Lis1-null
heterozygotes (15), the types of deficits expected
based on the human phenotype. The disruption of
hippocampal cellular and synaptic physiology
(16) seen in the Lis1 mutant mice were the result
of neuronal migration defects. Golgi impregna-
tion studies demonstrated that the dendritic arbor
of neurons from both wild type and Lis11/KO
cells located within stratum pyramidale were not
significantly different, but the dendritic arbor
of heterotopic pyramidal cells was significantly
smaller compared with both cell types, with
stunted dendrites that make fewer dendritic
branches. The enhanced excitability displayed by
slices from Lis11/KO mice provides a potential
explanation for the intractable seizures seen in ILS
patients.
Several lines of evidence support the conclusion
that there are in vivo migrational defects in mice
with reduction of LIS1 dosage (13). These include
histological analysis during development, BrdU
birthdating experiments, and in vitro migration of
granule cells from cerebellar cell re-aggregates.
Other studies from our laboratory have extended
and confirmed these findings (17, 18). The
development of the pre-plate, CajalRetzius cells
and the radial glial scaffold appeared unaffected
by LIS1 levels. LIS1 also was required for the
normal generation and survival of cortical VZ
neuroblasts. This study suggests that LIS1 is
important not only for neuronal migration, but
also for cell proliferation during neurogenesis as
well as for neuronal survival. Consistent with
these studies, Lis1 is co-localized with cytoplasmic
dynein at the mitotic kinetochores, indicating
a role of Lis1 in chromosomal behavior (19). For
example, microinjection of anti-Lis1 antibody
resulted in a delay of mitotic progression accom-
panied with chromosomal defects at the meta-
phase plate. Lis1 is also expressed at the cortical
(marginal) area of proliferating epithelial cells,
and its overexpression results in disruption of
the mitotic spindle organization as a result of
reductions in cortical distribution of dynein.
Lissencephaly and LIS1
Thus, the Dynein/Lis1 complex plays an impor-
tant role in the regulation of spindle orientation.
More recently, it has been possible to directly
examine neuronal migration in slice cultures from
embryonic cortex. Lipophilic dyes or in utero tran-
sfection with green fluorescent protein are used to
label the migrating cells, and migration is recorded
by confocal time-lapse video microscopy. The use
of RNAi knockdown of Lis1 definitively demon-
strated that LIS1 is required for neuronal migration
(20, 21). Additionally, LIS1 knockdown was
associated with neural stem cell division (21),
consistent with an important role for LIS1 in
proliferation.
LIS1, MTs and dynein
Neuronal migration is driven by complex mech-
anisms that are orchestrated by many proteins
that interact mainly, but not exclusively, to
promote the recruitment, organization, stability
and movement/function of microtubules and
actin cytoskeleton. LIS1 is an atypical microtu-
bule associated protein. LIS1 consists of seven
spaced WD-40 repeats, and probably assumes the
structure of a propeller wheel similar to other
WD-40 repeat proteins. Bovine and murine LIS1
are almost identical in amino acid sequence. In
mammals, LIS1 has at least two identified func-
tions. LIS1 is a non-catalytic subunit of platelet-
activating factor acetylhydrolase (PAFAH)
isoform Ib, an inactivating enzyme for platelet-
activating factor (PAF) (22), so the formal gene
name for LIS1 is PAFAH1B1. The catalytic alpha
subunits (a1 and a2) inactivate PAF, although the
role of LIS1 in regulating enzymatic function is
unknown. In addition, LIS1 has a non-enzymatic
function; it is part of a highly conserved pathway
that regulates nuclear migration in fungi (23). The
LIS1 homologue in the slime mold Aspergillus
nidulans, NudF is part of a signaling pathway that
regulates nuclear migration along microtubules
via the regulation of dynein motor function. In
this organism, nuclei migrate toward the growing
tip of hyphae to establish regular spacing, a pro-
cess termed nucleokinesis (24, 25). Several nuclear
distribution mutants (nud mutants) have been
generated in this organism (25, 26). One of these,
nudF is the fungal homologue of LIS1 (27), while
other nud mutants directly implicated cytoplasmic
dynein and dynactin in this process (28, 29).
The nud pathway is remarkably conserved in
eukaryotes, and the regulation of dynein motor
function by LIS1 and microtubule organization is
conserved in mammalian cells (Fig. 3). LIS1
associates with the microtubule fraction of cell
supernatants (30) and is present in regions of high
299
Wynshaw-Boris
microtubule density in neurons and fibroblasts,
especially at the centrosome and microtubule-
organizing center (MTOC) (19, 31). A large body
of data provide strong evidence that LIS1 dosage
regulates cytoplasmic dynein motor function, but
how does LIS1 regulate neuronal migration?
It appears that LIS1 is required for nuclear
movement during neuronal migration by coupling
the nucleus to the centrosome (18). Cerebellar
granule cells from Lis11/2 mice displayed slowed
migration and a lengthening of the distance
between the centrosome and the nucleus. LIS1 is
localized to the centrosome with some extension
toward the nucleus. Consistent with an important
role in dynein function, inhibition of dynein by
overexpression of dynamitin resulted in a similar
phenotype. Arrest of nuclear movement have been
observed using RNAi knockdown of LIS1 in
embryonic cortical slice cultures (20, 21). These
studies strongly suggest that nuclear movement
during neuronal migration is mediated by a LIS1
complex regulating cytoplasmic dynein function,
a function that is conserved through evolution
from Aspergillus to mammals.
Central role of NudE homologs in LIS1 function
The nud pathway is conserved in all eukaryotes,
including mammals. Several groups, including our
own, reported the cloning of mammalian NudE
homologues from yeast two-hybrid screens using
LIS1 as bait (3235), and some of these studies
PP2A
PR65
LIS1
NDE1 LIS1 NDEL1 P
P Ser198
LIS1
Centrosomal CDLC CDHC
Proteins
CDK5
Centrosome
function
Microtubule
dynamics
Aurora
Nuclear movement
neuronal migration
300
provided a direct link between LIS1 and dynein
motors. There are at least two mammalian NudE
homologues: mNudE (now termed NDE1) and
NUDEL (now termed NDEL1). Both are coiled
coil proteins of 344-345 amino acids and contain
10% serines. NDEL1 and NDE1 proteins co-
localize with LIS1 at the centrosome or MTOC in
mitotic embryonic neuroblasts or fibroblasts. Both
NDEL1 and LIS1 directly interact with the CDHC,
providing a direct link between LIS1 and cytoplas-
mic dynein motors. mNudE (NDE1) binds to the
dynein light chain, and its mechanism of dynein
regulation has not been as extensively studied at this
time as NDEL1. Overall, these results are consistent
with the hypothesis that LIS1 and NDEL1 are
responsible for the localization of CDHC (with
associated proteins) and regulate dynein motor
function and organelle transport in the cell.
NDEL1 is a phosphoprotein. It is a bona fide
substrate for Cdk5/p35 in vitro and in vivo at three
sites: S198, T219 and S231 (34, 35). Cdk5 phos-
phorylation of NDEL1 controls its cellular locali-
zation and probably influences dynein motor
function. These same serine residues are conserved
in mNudE (NDE1), suggesting that NDE1 may be
a target of Cdk5 as well. However, direct in vitro
and in vivo experiments can address whether or not
NDE1 is a substrate for the Cdk5 complex.
In addition to CDK5 (or CDK1 in fibroblasts),
NDEL1 is phosphorylated by the polo-like
mitotic kinase Aurora-A (36). Polo-like kinases
are conserved in all eukaryotes (37). At the G2/M
Fig. 3. The LIS1/NDE/dynein com-
plex. LIS1 interacts directly with
NDE1, which interacts with the cyto-
plasmic dynein light chain (CDLC) to
regulate centrosomal protein localiza-
tion and function. LIS1 also directly
interacts with NDEL1, and LIS1 and
14-3-3
NDEL1 directly interact with the cyto-
plasmic dynein heavy chain (CDHC) to
Thr219
Ser231
regulate centrosomal protein localiza-
tion and function as well as microtubule
dynamics. These interactions are critical
for nuclear movements and neuronal
p35
migration. NDEL1 appears to be the
focus of regulation, as it is phosphory-
lated by two different mitotic kinases:
CDK5 with its required co-activator p35
Ser251 phosphorylates NDEL1 on three sites
(S198, T219 and S231); and Aurora-A
kinase phosphorylates NDEL1 on S251.
CDK5-phosphorylated NDEL1 binds
to 14-3-3e, which protects NDEL1 from
A de-phosphorylation, perhaps by phos-
phatase 2A (PP2A), as NDEL1 binds to
the PP2A regulatory subunit PR65a.

transition and during early stages of mitosis, Plks
have been implicated in the maturation of
centrosomes, the activation of Cdk1/cyclin B,
disassembly of the Golgi complex and removal of
cohesin subunits from chromatin. Aurora-A kinase
efficiently phosphorylates NDEL1 at serine 251 at
the beginning of mitotic entry coincidently with
Aurora-A activation at the G2-prophase transi-
tion, while S251 phosphorylation suddenly disap-
peared after prophase (36). Comparing Aurora-A
mediated S251 phosphorylation with CDK1
mediated T219 NDEL1 phosphorylation, it ap-
pears that phosphorylation by CDK1 inhibited
further phosphorylation by Aurora-A, suggesting
that Aurora-A mediated phosphorylation of
NDEL1 is transient and may trigger the entrance
into prophase, followed by degradation and
CDK1 mediated phosphorylation. Persistent and
simultaneous phosphorylation of NDEL1 by both
Aurora-A and CDK1 appeared to be inhibitory to
the generation of normal spindle and chromo-
some segregation. Phosphorylation of NDEL1 by
CDK1 facilitates katanin p60 recruitment to the
centrosome to trigger microtubule remodeling
and shortening (38). NDEL1 phosphorylation by
Aurora-A is essential for mitotic entry and
centrosome separation by recruitment of TACC3
and XMAP215 at the centrosome (36), critical
events for centrosome-mediated MT assembly in
mitosis. This coordinate regulation of NDEL1
phosphorylation by two mitotic kinases suggests
that NDEL1 is a central component of the LIS1
complex to regulate proliferation, and the activity
of NDEL1 and the LIS1/dynein complex appears
to be tightly regulated by a typical signal trans-
duction pathway (Fig. 3). The role of these
phosphorylation events during neuronal migra-
tion require further investigation.
Recent structural studies have suggested that
subunits of PAF-AH compete with NDEL1 for
LIS1 binding (39). They solved the structure of
LIS1 in complex with the a2/ a2 PAF-AH
homodimer. One LIS1 homodimer binds sym-
metrically to one a2/ a2 homodimer via the highly
conserved top faces of the LIS1 b propellers. The
same surface of LIS1 contains sites of mutations
causing lissencephaly and overlaps with a putative
dynein binding surface. NDEL1 competes with
the a2/ a2 homodimer for LIS1 via a complex
interaction that requires both the N- and C-
terminal domains of LIS1.
Functional studies of NDEL1 and NDE1
The studies in the previous section suggest that
NDEL1 has a critical and central role in mediating
the effects of LIS1. Consistent with this notion, we
Lissencephaly and LIS1
made a knockout and conditional knockout of
Ndel1 (40). Complete loss of Ndel1 results in early
embryonic lethality, demonstrating that Ndel1 is
an essential gene. RNAi knockdown studies of
Ndel1, Lis1 or dynein were performed by in utero
electroporation into the developing neocortex
(20). RNAi knockdown of any of these three
genes impaired neuronal positioning and caused
the uncoupling of the centrosome and nucleus,
similar to our studies with Lis1 mutants (18).
Overexpression of LIS1 partially rescued the
positioning defect caused by Ndel1 RNAi but
not dynein RNAi, whereas overexpression of
NDEL1 did not rescue the phenotype induced
by Lis1 RNAi. These findings with NDEL1 are in
contrast to genetic ablation of the LIS1-interacting
protein NDE1 in mouse (41). Complete loss of
Nde1 results in viable mice with microcephaly,
especially in the cerebral cortex. Cortical lamina-
tion was mostly preserved, but the mutant cortex
had fewer neurons and very thin superficial
cortical layers (layers 24), with dramatic defects
in mitotic progression, mitotic orientation, and
mitotic chromosome localization in cortical pro-
genitors. The small cerebral cortex seems to reflect
both reduced progenitor cell division and altered
neuronal cell fates. Taken together, these results
provide strong evidence that NDEL1 and NDE1
interaction with LIS1 are particularly important
for microtubule organization, nuclear transloca-
tion, neuronal positioning and development.
NDEL1 and 14-3-3e: a molecular explanation for
MillerDieker syndrome
To further investigate the potential differences
between ILS and MDS, we examined the function
of genes deleted in MDS, but not ILS. We found
that 14-3-3e binds to P-NDEL1 (42), and that
14-3-3e is always deleted in MDS patients (43).
The 14-3-3 family of genes is found in all
eukaryotic cells, and there are seven separate
genes or isoforms present in mammalian cells
(denoted by Greek letters). These highly con-
served regulatory molecules bind to sequence-
specific phosphoserine and phosphothreonine
motifs. More than 100 binding partners for 14-3-3
proteins have been discovered, resulting in mod-
ulation of signal transduction, cell cycle control,
apoptosis, stress responses, vesicular transport
and malignant transformation (44). We produced
mice deficient for 14-3-3e (42). These mice display
developmental brain and neuronal migration
defects similar to Lis1 heterozygous mutant mice.
14-3-3e1/2 mice showed mild disorganization
in the hippocampus and thinning of the cortex
that was more severe in the 14-3-3e2/2 mice.
301
Wynshaw-Boris
CP
IZ
VZ
Interkinetic Neuronal
nuclear migration
migration
Neuronal migration was also quantitated by
BrdU analysis and revealed that, similar to Lis1
mutant mice, as the dose of 14-3-3e is reduced, the
rate of neuronal migration decreases. Double
heterozygotes of 14-3-3e and Lis1 display more
severe migration defects in both the cortex and
hippocampus than single heterozygotes, provid-
ing genetic evidence that 14-3-3e may be respon-
sible for most severe form of LIS, complete agyria,
seen in MDS. 14-3-3e binds directly to CDK5/
p35-phosphorylated NDEL1, and this binding
maintains NDEL1 phosphorylation (Fig. 3). As
NDEL1 also binds to a regulatory subunit of the
PP2A phosphatase (Fig. 3), it is likely that
binding of 14-3-3e to P-NDEL1 protects it from
phosphatase attack. Similar to LIS1, reduction of
14-3-3e results in mislocalization of NDEL1 and
LIS1, consistent with reduction of cytoplasmic
dynein function. These results establish a crucial
role for 14-3-3e in neuronal development by
sustaining the effects of CDK5 phosphorylation.
It also provides a molecular explanation for the
differences in severity of human neuronal migra-
tion defects with 17p13.3 deletions, namely ILS
and MDS. In addition, it places 14-3-3e in the
LIS1 pathway.
Conclusions and future directions
The identification of LIS1 as the first lissence-
phaly gene and the first gene required for neuronal
migration has illustrated the importance of
cytoplasmic dynein in the regulation of neuronal
migration by modulating nuclear migration in
a pathway conserved in virtually all eukaryotes.
By examining the effect of reduction of LIS1 either
genetically or by RNAi knockdown, it is clear that
LIS1 has a broader effect on proliferation and
survival. Does LIS1 and its complex regulate
distinct aspects of proliferation, migration and
survival? Or is there a similarity between these
cellular processes that LIS1 and its complex
regulates? Although direct experiments have not
been performed to address these questions, it is
apparent that there is similarity between inter-
kinetic nuclear migration and neuronal migration
302
Fig. 4. Similarity between interkinetic
nuclear migration and neuronal migra-
tion. Labeling is the same as Fig. 1,
except the centrosome is represented as
a red circle. A cell undergoing interki-
netic nuclear migration is enlarged on
the left, while a migrating neuron is
enlarged on the right. Microtubules that
stretch from the centrosome to the
nucleus are red.
(Fig. 4). During interkinetic nuclear migration,
the centrosome is at the apical surface and the
nucleus is more basal. The nuclei move in a basal
direction during G1, and during G2 toward the
apical surface, where they divide. The nucleus is
surrounded by microtubules anchored at the
centrosome. During neuronal migration, the
migration of the nucleus takes place in a similar
fashion, but reversed by 180. The centrosome is
ahead of the nucleus and is attached to the nucleus
by microtubules. The centrosome moves forward
into the leading process, followed by the nucleus
that is tethered to the centrosome by micro-
tubules. Interkinetic nuclear migration and neu-
ronal (nuclear) migration both involved an
oscillation in which the distance between the
nucleus and centrosome lengthens and then
contracts. As LIS1 and the dynein complex are
essential for interkinetic nuclear migration and
neuronal migration by coupling the centrosome to
the nucleus, it is reasonable to hypothesize that
both processes act in similar cell biological
fashions. As the major difference between these
processes is the position of the centrosome and
nucleus relative to the apical surface, it is likely
that apical-basal polarity is an important factor in
controlling the position of the centrosome and the
action of the LIS1 complex on nuclear-centrosomal
coupling.
Acknowledgements
The author is grateful for all those who have worked in his
laboratory on neuronal migration, and for his colleagues in the
neuronal migration field. Because of restraints on the number
of references, it was impossible to cite all relevant papers. This
work was supported by grants from the National Institutes of
Health (NS41030 and HD047380).
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