Lee Niswander, Ph.D.
Genetic and Cellular Control of Vertebrate Embryonic Development
Our research focuses on the genetic and cellular mechanisms that control embryonic development. In particular we are interested in the processes required for closure of the neural tube and for development of the vertebrate limb and lung. Our approaches include a forward genetic screen in mice to identify mutations that affect specific processes of embryonic development and embryological and molecular manipulation of the chick embryo.
ENU-mutagenesis screen for genes required for mouse development. A forward
genetic approach is ideal for identifying key genes that regulate specific
developmental processes of interest. In an on-going screen in mice for
ENU (ethylnitrosourea)-generated mutations, we are identifying recessive
mutations that disrupt specific aspects of embryonic development. We have
focused on mutations that affect neural tube closure, limb and lung development.
To learn more about the screen, to view pictures of the various mutants,
and to follow our progress in identifying the genes responsible and their
mechanism of action, please go to our
Website - http://mouse.ski.mskcc.org/
Neural Development
Neural tube defects (NTD) are the second most common human birth defect,
affecting 1:1,000 births. However, the underlying genetic basis of NTD
in humans is very poorly understood. Failure to close the neural tube
in the brain leads to exencephaly (anencephaly in humans), failure to
close the spinal neural tube causes spina bifida. Neural tube closure
involves a variety of coordinated developmental processes including pattern
formation, proliferation, cell differentiation, and changes in cell shape
and cell movement. The mouse provides an excellent model to systematically
identify the genes that are required for closure of the neural tube. The
chick embryo provides an excellent experimental system to test the function
of the identified genes.
Our studies seek to identify the genes required and their mechanism of action.
Genetics of neural tube closure.
To advance an understanding of the causes of birth defects of the brain
and spinal cord in humans, we are using the mouse as a model to systematically
identify the genes involved in closure of the neural tube, followed by
a comprehensive cellular and molecular characterization of their mechanism
of action. From the ENU mutagenesis screen we have identified a large
number of mouse lines with NTDs. Our approach is to identify the gene
by genetic mapping and to characterize the phenotype to elucidate the
developmental process that is affected. Priority will be given to mutations
that are linked to potential human disease loci. Moreover, we will test
whether the NTD can be suppressed by dietary supplementation to understand
the genetics underlying the effects of, for instance, folic acid which
can suppress the incidence of NTDs by an unknown mechanism.
Experimental studies of neural development – BMPs and WNTs control
patterning and growth of the neural tube
The chick provides an experimentally amenable and rapid system to determine
the function of a gene. We are using the chick to functionally characterize
the genes arising from the mutagenesis screen, as well as other developmentally
important genes. In the latter category, we have explored the role of
the Bone Morphogenetic Protein (BMP) and WNT signaling in coordinating
patterning and growth in the embryonic spinal neural tube using gain and
loss of function approaches including in vivo knockdown of gene expression
by siRNAs. We found that BMPs are necessary to regulate pattern formation,
including the establishment of discrete expression domains of Wnt signaling
components along the dorsal-ventral axis of the neural tube and that WNTs
function as mitogenic signals regulating neural tube growth. Thus, BMPs,
acting through WNTs, couple patterning and growth to generate dorsal neuronal
progenitor populations in the appropriate proportions within the neural
tube.
Limb Development
Key signals that control limb growth and patterning have been identified
in the past 10 years, yet surprisingly little is known about how these
regulators function, what are their upstream activators and downstream
targets, or how patterning information is translated into skeletal elements
of appropriate size and shape.
Our interests in limb development continue to revolve around the formation of the skeletal primordia and the function of the apical ectodermal ridge (AER). The AER is the critical signaling center that controls limb outgrowth via the production of Fibroblast growth factors (FGFs). We are exploring the mechanisms that regulate AER formation and function through molecular manipulations of the chick embryonic limb, imaging techniques and mouse genetics. Our studies of the chick limb have determined that BMPs are upstream of two important developmental processes, that of AER formation and dorsal-ventral patterning. The BMP signal bifurcates at the level of two different transcription factors to mediate AER formation and dorsal-ventral patterning differentially.
A disruption in AER formation and function appears to be the primary defect in a mouse mutant we found in the mutagenesis screen that leads to missing digits, dorsal-ventral digit duplications, and aberrant ossification. We have identified other limb mutants that cause polydactyly (extra digits), soft tissue syndactyly (the webbing between the digits does not undergo regression), and other skeletal defects including shortening of some limb elements. Moreover, we are further exploring genetic interactions between various mutant lines to determine whether they regulate similar functions or act within a genetic pathway. These studies provide an unbiased means to identify key developmental regulators and will contribute significantly to a greater understanding of the genetic and cellular control of vertebrate limb development. (Grants from the National Institutes of Health provided support for the limb work.)
Lung Development and Disease
The lung and other highly branched organs such as the kidney and lacrimal
gland develop from a simple epithelial bud into a complex three dimensionally
patterned functional organ. This happens through a process called branching
morphogenesis. We wish to elucidate the fundamental processes underlying
the development of the lung, as well as the pathogenesis of lung disease.
To do so we use a combination of forward and reverse genetics in mice
and molecular manipulations in organ cultures to identify key regulators
of branching morphogenesis. We have identified novel regulators of lung
development through the mouse ENU mutagenesis screen. Moreover we have
manipulated key signaling pathways such as BMP and WNT to elucidate the
mechanisms by which the position and shape of the bud are determined.
The future goal is to study if and how developmentally important genes
are involved in tissue injury and repair, in particular in diseased lungs.
(This work is supported by the National Institutes of Health and by the
Sandler Program for Asthma Research)
Publications
Niswander, L. and Martin, G.R. (1992). Fgf-4 expression during gastrulation,
myogenesis, limb and tooth development in the mouse. Development 114:
755-768.
Niswander, L. and Martin, G.R. (1993). FGF-4 and BMP-2 have opposite effects
on limb growth. Nature 361, 68-71.
Niswander, L., Tickle, C., Vogel, A., Booth, I., and Martin, G.R. (1993).
FGF-4 replaces the apical ectodermal ridge and directs outgrowth and patterning
of the limb. Cell 75, 579-587.
Niswander, L., Jeffrey, S., Martin, G.R., and Tickle, C. (1994). Positive
feedback loop coordinates growth and patterning in the vertebrate limb.
Nature, 371, 609-612.
Yang, Y. and Niswander, L. (1995). Interaction between the signaling molecules
WNT and SHH during vertebrate limb development: dorsal signals regulate
anteroposterior patterning. Cell 80, 939-947.
Zou, H. and Niswander, L. (1996). Requirement for BMP signaling interdigital
apoptosis and scale formation. Science 272, 738-741.
Kuhlman, J. and Niswander, L. (1997). Limb deformity protein: Role in
mesodermal induction of the apical ectodermal ridge. Development 124,
133-139.
Zou, H., Wieser, R., Massagué, J. and Niswander, L. (1997). Distinct
roles of type I bone morphogenetic protein receptors in the formation
and differentiation of cartilage. Genes and Development 11, 2191-2203.
Zou, H., Choe, K-M., Lu, Y., Massagué, J., and Niswander, L. (1997).
BMP signaling and vertebrate limb development. Cold Spring Harbor Symposis
on Quantitative Biology, Vol. LXII, 269-272.
Yang, Y., Drossopoulou, G., Chuang, P.T., Duprez, D., Marti, E., Bumcrot,
D., Vargersson, N., Clarke, J., Niswander, L., McMahon, A. and Tickle,
C. (1997). Relationship between dose, distance and time in Sonic Hedgehog-mediated
regulation in the chick limb development. Development 124, 4393-4404.
Niswander, L. (1997). Limb mutants: what can they tell us about normal
limb development? Current Opinion in Genetics and Development 7, 530-536.
Crowe, R., Henrique, D., Ish-Horowicz, D., and Niswander, L. (1998). A
new role for Delta in cell fate decisions: patterning the feather array.
Development 125, 767-775.
Crowe, R. and Niswander, L. (1998). Disruption of scale development by
Delta-1 misexpression. Dev Bio 195, 70-74.
Bushdid, P. B., Brantley, D. M., Yull, F. E., Blaeuer, G. L., Hoffman,
L. H., Niswander, L., and Kerr, L. D. (1998). Inhibition of NF-kB activity
results in disruption of the apical ectodermal ridge and aberrant limb
morphogenesis. Nature 392, 615-618.
Varley, J. E., McPherson, C. E., Zou, H., Niswander, L. and Maxwell, G.
D. (1998). Expression of a constitutively active type I BMP receptor using
a retroviral vector promotes the development of adrenergic cells in neural
crest cultures. Developmental Biology 196, 107-118.
Gibson-Brown, J., Agulnik, S.I., Silver, L.M., Niswander, L., and Papaioannou,
V.E. (1998) Involvement of T-box genes Tbx2-Tbx5 in vertebrate limb specification
and development. Development 125, 2499-2509.
Crowe, R., Zikherman, J. A. and Niswander, L. (1999). Delta-1 negatively
regulates the transition from prehypertrophic to hypertrophic chondrocytes
during cartilage formation. Development 126, 987-998.
Pizette, S. and Niswander, L. (1999). BMPs negatively regulate structure
and function of the limb apical ectodermal ridge. Development 126, 883-894.
Obara-Ishihara, Kuhlman, J., Niswander, L. and Herzlinger, D. (1999).
The surface ectoderm is essential for nephric duct formation in intermediate
mesoderm. Development 126, 1103-1108.
Pizette, S. and Niswander, L. (2000). BMPs are required at two steps of
limb chondrogenesis: formation of prechondrogenic condensations and their
differentiation into chondrocytes. Dev. Biology 219, 237-249.
Barna, M., Hawe, N., Niswander, L., Pandolfi, P. P. (2000). PLZF regulates
limb and axial patterning. Nature Genetics 25, 166-172.
Timmer, J., Johnson, J. and Niswander, L. (2001). The use of in ovo electroporation
for the rapid analysis of neural-specific murine enhancers. genesis 29,
123-132.
Holmes, G. and Niswander, L. (2001). Expression of slit-2 and slit-3 during
chick development. Dev Dynamics 222, 301-307.
Pizette, S., Abate-Shen, C. and Niswander, L. (2001). BMP controls proximodistal
outgrowth, via induction of the apical ectodermal ridge, and dorsoventral
patterning in the vertebrate limb. Development 128, 4463-4474.
Niswander, L. (2002). Interplay between the molecular signals that control
vertebrate limb development. International Journal of Developmental Biology
46, 877-881.
Timmer, J., Wang, C. and Niswander, L. (2002). BMP signaling patterns
the dorsal and intermediate neural tube via regulation of Homeobox and
Helix-Loop-Helix transcription factors. Development 129, 2459-2468.
Niswander, L. and Anderson, K. V. (2002). Hopeful monsters and morphogens
at the beach. Nature Cell Biology 11, E259-262.
Atit, R. and Niswander, L. (2003). EGF signaling patterns the feather
array by promoting the interbud fate. Developmental Cell 4, 231-40.
Niswander, L. (2003). Pattern Formation: old models out on a limb. Nature
Reviews Genetics 4, 133-143.
Liu, A.,, Li, J.Y.H, Bromleigh, C., Lao, Z., Niswander, L. and Joyner,
A. (2003). FGF17b and FGF18 have different midbrain regulatory properties
from FGF8b and activated FGF receptors. Development 130, 6175-6185.
Huangfu, D., Liu, A., Rakeman, A. S., Murcia, N. S., Niswander, L. and
Anderson, K. V. (2003). Hedgehog Signaling in the Mouse Requires Intraflagellar
Transport Proteins. Nature 426, 83-87.
Holmes, G. Crooijmare, R., Groenen, M., Niswander, L. (2003). ALC (ADJACENT
TO LMX1 IN CHICK) is a novel dorsal limb mesenchyme marker. Gene Expression
Patterns 3, 735-741.
Rouzankina, I., Abate-Shen, C. and Niswander, L. (2004). Dlx genes integrate
positive and negative signals during feather bud development. Developmental
Biology 265, 219-233.
Niemann, S., Zhao, C., Pascu, F., Stahl,, Aulepp, U., Niswander, L., Weber,
J., Ulrich Müller, U. (2004). Homozygous WNT3 mutation causes tetra-amelia
in a large consanguineous family. Amer J Human Genetics 74, 558-63.
Chesnutt C & Niswander (2004) Plasmid-based short-hairpin RNA interference
in chicken embryo. Genesis 39 73-8
Chesnutt, C., Burrus, L. Brown, A.M.C., and Niswander, L. (2004). Coordinate
regulation of neural tube patterning and proliferation by TGF? and WNT
signaling. Developmental Biology, in press
