Chief, Section on Molecular Development
Laboratory of Developmental Biology
Department of Cell and Molecular Biology
agroves@hei.org
Education:
Sidney Sussex College, Cambridge, UK, - MA, BA (1988)
Ludwig Institute for Cancer Research, University College/Middlesex Hospital Branch, London, UK - Ph.D. (1992)
Fellowships and Awards:
2005-2008: March of Dimes Research Grant
“The role of Notch signaling in inner ear development”
2003-2005: R21 Pilot Project Grant, DC06139 (Co-PI with Neil Segil)
"Marking hair cell progenitors with BAC transgenics"
2003-2007: RO1 Grant, DC06185
"Characterization of Inner Ear Stem Cells"
2001-2003: R21 Pilot Project Grant,DC04876
"A novel method for targeted gene disruption in the inner ear"
2001-2006: Hair Cell Regeneration Initiative Grant (Joint PI with Dr. Neil Segil, House Ear Institute)
National Organization for Hearing Research
"Sensory hair cell progenitors in the mammalian inner ear”
2001-2006: RO1 Grant, DC04675
"Molecular and cellular mechanisms of otic placode induction"
1999-2001: Basil O’Connor Starter Scholar Award, March of Dimes Birth Defects Foundation
"Investigation of the cellular signals in otic placode induction"
Professional Experience:
2001 -
Section Chief (Scientist II), Section on Molecular Development
Department of Cell and Molecular Biology, House Ear Institute
2000 -
Adjunct Assistant Professor, Department of Cell and Neurobiology and Clinical Assistant Professor
Department of Otolaryngology, University of Southern California.
1999-2001
Section Chief (Scientist I), Section on Molecular Development
Department of Cell and Molecular Biology, House Ear Institute
1997-1999
Senior Research Fellow in Biology, California Institute of Technology
Laboratory Head: Dr. Marianne Bronner-Fraser
1992-1996
Research Fellow in Biology, California Institute of Technology
Laboratory Head: Dr. David Anderson
Research:
My laboratory uses the inner ear as a model system to address fundamental questions in developmental biology and regeneration. The transformation of a simple piece of placodal epithelium into a sensory organ of extreme morphological complexity provides an opportunity to study competence, induction, pattern formation, cell-type differentiation and morphogenesis. In addition, the failure of sensory hair cells to regenerate in deafened mammals is a clinical problem that may prove tractable through an understanding of inner ear development and the role of stem cells and progenitor cells in hair cell formation. Our work exploits the complementary strengths of chick embryology and mouse genetics to address these problems.
Molecular mechanisms of inner ear induction: The nature of competence and patterning of developmental fields
The first focus of my lab’s research is on the earliest events in the formation of the inner ear. We have demonstrated that FGF signaling is both necessary and sufficient for the induction of many markers of the otic placode, the anlagen of the inner ear. More recently, we have shown that only ectoderm that lies adjacent to the anterior neural plate is competent to respond to FGF signaling. This “pre-placodal” ectoderm has been suggested to give rise to all craniofacial placodes, and is characterized by the expression of a number of transcription factors such as Dlx, Six, and Eya family members. We have shown that competence to respond to FGF correlates with the up-regulation of these transcription factors, but we do not yet know if any of these factors are either necessary or sufficient for FGF responsiveness. In the next phase of our work, we aim to test the roles of these transcription factors in the acquisition of competence to respond to FGF. To further understand the basis of this competence, we have also begun to analyze the intracellular signaling pathways by which FGFs induce otic placode genes, and whether these pathways are intact in competent versus non-competent ectoderm. We have also identified a novel Forkhead gene - Foxi3 - that is expressed in the pre-placodal region. We are currently knocking this gene out in mice to determine its role in the formation of the otic and other craniofacial placodes. This work forms the main part of my RO1 renewal (DC04675).
The otic placode develops from a field of cells that express the Pax2 gene. We have used Cre-Lox genetic fate mapping to show that only a medial subset of Pax2 cells will form the otic placode, with the rest differentiating as epidermis. We have shown that the canonical Wnt signaling pathway is active in this medial subset of Pax2 cells but not the more lateral population. We hypothesize that Wnt signaling may mediate a placode-epidermis fate decision, such that Pax2 cells receiving high levels of Wnt signaling differentiate as otic placode cells, and those receiving little or no Wnt signaling differentiating as epidermis. Since b-catenin is a key mediator of Wnt signaling, we have used Cre-Lox technology to conditionally delete b-catenin in Pax2 cells, which drastically reduces the size of the otic placode. However, constitutive activation of b-catenin in Pax2 cells greatly enlarges the otic placode at the expense of epidermis. This confirms our hypothesis that Wnt signaling acts instructively to direct Pax2 cells to an otic placode fate. In the future, we aim to understand how Wnt signaling instructs Pax2 progenitors to an otic placode cell fate, and whether there is synergy between the Wnt and FGF signaling pathways in this process. We have identified a second novel Forkhead gene - Foxi2 - that marks the epidermis immediately adjacent to the otic placode, but is excluded from the placode itself. We are knocking this gene out in mice to determine if it is involved in actively restricting the size of the otic placode, and we will also test whether this gene – and, by extension, epidermal differentiation - is negatively regulated by Wnt signaling.Testing a compartment boundary model of ear development: The application of Cre-Lox technology to the developing inner ear
One of the challenges of developmental biology is to understand how morphologically complex structures arise from simple derivatives. In the case of the inner ear, it has been proposed that different sensory and non-sensory structures are derived from distinct developmental compartments established when the ear is still a simple spherical otocyst. A number of transcription factors have been shown to be spatially restricted in the otocyst, with their expression frequently defined by the intersection of the dorso-ventral, antero-posterior or medio-lateral otocyst axes. We have evidence to suggest that the Iroquois (Irx) family of transcription factors play a key role in the formation of developmental compartments in the otocyst, and that as a result, different members of this gene family may variously specify the differentiation of the VIIIth ganglion, the cochlea, the utricle and saccule and the formation of the semicircular canals. We will use a combination of genetic lineage tracing and gain- and loss-of-function approaches in mouse embryos to test the role of the Iroquois family in the formation of these developmental compartments and their derivatives.
In order to achieve the precise spatial and temporal control of gene deletion or over-expression necessary for these studies, we have established the use of Cre-Lox technology in our lab. Our first BAC transgenic Cre mouse line, Pax2-Cre, can be used to delete or over-express genes in the entire inner ear, and we are now constructing further BAC mouse lines that drive expression of Cre recombinase, or the tamoxifen-inducible Cre-ER2 in more restricted regions of the inner ear. These lines will be used to activate or inactivate Iroquois gene expression in different regions of the inner ear to test their role in compartment formation and inner ear differentiation.The role of Notch signaling in inner ear neurogenesis, morphogenesis and hair cell formation: A combination of canonical and non-canonical signaling pathways?
Notch signaling plays a central role in the formation of boundaries and the segregation of different cell types during development. Notch receptors and their Delta and Jagged ligands are expressed at every point in inner ear development – in the otic placode, during delamination of the VIIIth ganglion, during morphogenesis of the inner ear, and during the formation of sensory hair cells. We have begun to test the function of Notch signaling in ear development in collaboration with Pamela Stanley of Albert Einstein College of Medicine by conditionally targeting O-fucosyltransferase (an enzyme absolutely required for Notch signaling) in the inner ear. Surprisingly, disruption of this gene has no effect on ear development until the time of hair cell formation, where it leads to a massive over-production of hair cells. Moreover, we find that a variety of Hes and Hey gene family members (which have been proposed to mediate canonical Notch signaling) are expressed in the inner ear, but are only Notch-responsive during hair cell differentiation, not at earlier times. This suggests that both canonical and non-canonical modes of Notch signaling are deployed during inner ear development, with non-canonical signaling occurring early, and canonical signaling occurring later during hair cell differentiation. We will confirm these results with a conditional allele of RBPJ/CSL, a key mediator of canonical Notch signaling, in collaboration with Tasuku Honjo, and by examining non-canonical Notch signaling pathways in collaboration with Gerry Weinmaster at UCLA. This work is currently supported by a research grant from the March of Dimes, and by an RO1 (DC06185), on which Dr. Neil Segil of the House Ear Institute is a co-investigator.
Sensory hair cells of the mammalian organ of Corti do not regenerate when lost as a consequence of injury, disease, or age-related deafness. Deafness in mammals is thus permanent and irreversible. This lack of regenerative ability contrasts with other vertebrates such as birds, where the death of sensory hair cells causes the surrounding supporting cells to re-enter the cell cycle and give rise to both new hair cells and supporting cells. It is not clear whether the lack of mammalian hair cell regeneration is due to an intrinsic inability of supporting cells to divide and differentiate or to an absence or blockade of regenerative signals. In an effort to evaluate the capacity of the mammalian inner ear for regeneration, we first established a cell culture system in which progenitor cells isolated from the embryonic mouse auditory or vestibular system were able to proliferate and generate hair cells over extended periods in culture. We then developed methods to purify supporting cells from the postnatal mouse cochlea, and surprisingly found that these cells also retained the capacity for division and hair cell formation in the first postnatal week of life. However, this capacity is severely diminished in animals two weeks of age or older. We have shown that this inability to re-enter the cell cycle in older animals is partially dependent on the cyclin-dependent kinase inhibitor p27Kip1, as supporting cells isolated from juvenile p27Kip1 mutants retain the capacity for proliferation and differentiation.
We have also shown that another factor in the failure of regeneration in the mammalian inner ear is the Notch signaling pathway. As described above, disruption of all Notch signaling in the cochlea by conditional deletion of O-fucosyltransferase leads to a massive over-production of hair cells. Moreover, treatment of supporting cell cultures or neonatal cochlear organ cultures with gamma secretase inhibitors that block Notch signaling give rise to increased numbers of hair cells. This is the first demonstration that treatment with soluble small molecule pharmaceuticals can lead to hair cell regeneration
Our work suggests that two distinct molecular pathways may be central to understanding hair cell regeneration in mammals. On one hand, manipulation of p27Kip1 may stimulate re-entry of post-mitotic supporting cells in the organ of Corti, whilst manipulation of the Notch signaling pathway may promote hair cell differentiation from supporting cells. Our future work seeks to more fully understand the biology of these two pathways, whilst at the same time examining their potential in therapeutic approaches to hair cell regeneration.
This work is part of a collaboration between my lab and Dr. Neil Segil of the House Ear Institute, and is currently supported in part by an RO1-sized grant from the National Organization for Hearing Research on which Dr. Segil and I are listed as joint investigators, and by an RO1 (DC06185), on which Dr. Segil is a co-investigator.
Selected Publications:
White, P.M., Doetzlhofer, A., Lee, Y.-S., Groves, A.K. and Segil, N. (2006). Cochlear supporting cells retain the ability to divide and transdifferentiate into sensory hair cells. Nature, in press.
Arnold, J.S., Braunstein, E.M., Ohyama, T., Groves, A.K., Adams, J.C., Brown, M.C. and Morrow, B.E. (2006). Tissue specific roles of Tbx1 in the development of the outer, middle and inner ear, defective in 22q11DS patients. Human Molecular Genetics, in press.
Ohyama, T., Mohamed, O.A., Taketo, M.M., Dufort, D. and Groves, A.K. (2006). Wnt signals mediate a fate decision between otic placode and epidermis. Development, 133, 865-875.
Martin, K. and Groves, A.K. (2006). Competence of cranial ectoderm to respond to FGF signaling suggests a two step model of otic placode induction. Development, 133, 877-887.
Kil, S.-H., Streit, A., Brown, S.T., Agrawal, N., Collazo, A., Zile, M.H. and Groves, A.K. (2005). Distinct roles for hindbrain and paraxial mesoderm in the induction and patterning of the inner ear revealed by a study of vitamin A-deficient quail. Dev. Biol. 285, 252-271.
Brown, S.P., Wang, J. and Groves, A.K. (2005). Dlx gene expression during chick inner ear development. Journal of Comparative Neurology, 483, 48-65.
Doetzlhofer, A., White, P.M., Johnson, J.E., Segil, N. and Groves, A.K. (2004). In vitro growth and differentiation of mammalian sensory hair cell progenitors: A requirement for EGF and periotic mesenchyme. Dev. Biol. 272, 432-447.
Ohyama, T., and Groves, A.K. (2004a). Generation of Pax2-Cre mice by modification of a Pax2 bacterial artificial chromosome. Genesis 38, 195-199.
Ohyama, T. and Groves, A.K. (2004b). Expression of mouse Foxi class genes in early craniofacial development. Developmental Dynamics, 231, 640–646.
Groves, A.K. and Bronner-Fraser, M. (2000). Competence, specification and commitment in otic placode induction. Development 127, 3489-3499.
Morrison, S.J., Csete, M., Groves, A.K., Melega, W., Wold, B., and Anderson, D.J. (2000). Culture in reduced levels of oxygen promotes clonogenic sympathoadrenal differentiation by isolated neural crest stem cells. J. Neurosci. 20, 7370–7376.
Mujtaba, T., Piper D., Kalyani A., Groves A.K. , Lucero M. T., and Rao M. S. (1999). Lineage-restricted neural precursors can be isolated from both the mouse neural tube and cultured ES cells. Dev. Biol. 214, 113-127.
Shah, N.M., Groves, A.K., and Anderson, D.J. (1996). Alternative neural crest cell fates are instructively promoted by TGFb superfamily members. Cell 85, 331-343.
Verdi, J.M., Groves, A.K., Farinas, I., Jones, K., Marchionni, M., Reichardt, L.F., and Anderson, D.J. (1996). A reciprocal cell-cell interaction mediated by neurotrophin-3 and neuregulins on the early survival and development of sympathetic neurons. Neuron 16, 515-527.
Groves, A.K., George, K.M., Tissier-Seta, J.-P., Engel, J.D., Brunet, J.-F., and Anderson, D.J. (1995). Differential regulation of transcription factor gene expression and phenotypic markers in developing sympathetic neurons. Development 121, 887-901.
Groves, A.K., Entwistle, A., Jat, P.S. and Noble, M. (1993). The characterisation of astrocyte cell lines that display properties of glial scar tissue. Dev. Biol. 159, 87-104.
Groves, A.K., Barnett, S.C., Franklin, R.J.M., Crang, A.J., Mayer, M., Blakemore, W.F., and Noble, M. (1993). Repair of demyelinated lesions by transplantation of purified O-2A progenitor cells. Nature 362, 453-455.
