Axial patterning in the vertebrate inner ear: the role of Hedgehog signalling
Lead Research Organisation:
University of Sheffield
Department Name: Biomedical Science
Abstract
The inner ear is enormously important for the senses of balance and hearing. It consists of an intricate fluid-filled labyrinth housing a variety of extraordinarily sensitive sensory structures that respond to sound, movement and gravity. For correct inner ear function it is essential that each of these components form in exactly the right place, as any disturbance can lead to deafness or balance disorders. Indeed, congenital deafness is an important clinical problem, affecting approximately one in every thousand children at birth. Our aim is to understand how the inner ear develops in the embryo, and the mechanisms that ensure that the different cell types in the ear arise in the correct positions so that they can function accurately. We use embryos of a small tropical fish, the zebrafish, in our research, as it is a superb model for the study of vertebrate inner ear development. Importantly, zebrafish possess an inner ear that is very similar in most respects to those of other vertebrates like ourselves, and the embryo is transparent, meaning that the inner ear - even though an internal organ - can be visualised in the live organism. Moreover, the zebrafish is a powerful genetic organism, meaning that we can disrupt individual genes specifically to discover their function. Many different lines of zebrafish exist in which the ear develops with specific defects, or in which different cell types are marked with fluorescent dyes, and these can be used to identify key genes that are required for correct formation of the ear. In addition, embryos are abundant, easily available and amenable to manipulation. In this project, we will focus on understanding the effects of Hedgehog (Hh) proteins on ear development. Hh proteins are signalling molecules that give instructions to cells in the embryo, telling them how and where to develop. It is known that Hh has a crucial and early role in distinguishing one region of the ear from another, but its exact function is not fully understood. In particular, it does not seem to play the same role in zebrafish and mammals: in fish, it primarily regulates development of the anterior-posterior (head to tail) axis in the ear, whereas in mammals, its primary role is in regulation of the dorsal-ventral (back to belly) axis, which is perpendicular to the anterior-posterior axis. This is surprising, as the orientation of the inner ear in the head of the adult animal, and the majority of inner ear structures, are very similar between the two groups. Are mechanisms of ear development really so different in mammals and fish? It is important to answer this question, as the zebrafish is widely used as a model for human hearing and deafness. Our preliminary investigations now suggest that the role of Hh is more similar between mouse and zebrafish than it first appears. In particular, our studies indicate that Hh signalling is also involved in distinguishing between dorsal and ventral regions of the zebrafish inner ear. We aim to confirm this using a novel and unique panel of zebrafish mutants. In these fish, the function of genes that code for inhibitors of Hh signalling is disrupted, meaning that all cells now experience high levels of this signalling molecule. We will use these mutants to explore the mechanisms involved in formation of dorsal-ventral patterning in the zebrafish ear. In particular, we aim to establish exactly when Hh is acting during ear development, and whether it acts together with a second group of signalling molecules, those of the Wnt family, in patterning the ear. We will also test whether Hh has additional AP patterning roles in the developing inner ear of the mouse embryo. This work will lead to a greater understanding of how the inner ear develops in the vertebrate embryo, providing important basic knowledge that will help to inform clinical research into the many genetic conditions that lead to deafness in humans.
Technical Summary
The inner ear of the zebrafish displays homologies at every level - from developmental mechanisms to cellular physiology - with that of the mammal, and is widely used as a model for hearing and deafness. Nevertheless, there are differences in ear anatomy between the two groups. The most notable of these is the lack of a cochlea (the specialised hearing organ in amniotes) in the zebrafish ear. The role of Hh signalling is also reported to have a different role in the two groups, predominantly affecting otic dorsoventral (DV) axis formation in the mouse, but otic anteroposterior (AP) patterning in the zebrafish. Despite these observations, there is evidence to suggest that both DV and AP axes may be affected by Hh signalling in both groups. In particular, we have found that ear phenotypes in a new series of zebrafish mutants, in which Hh signalling is constitutively active, suggest that Hh signalling must be kept repressed for DL structures to develop correctly. In this proposal, we aim to clarify how Hh signalling acts to pattern the ear in both zebrafish and mouse. In the zebrafish, we will exploit the new mutants in inhibitors of the Hh pathway, and define the timing, mechanism of action and effects of upregulation of Hh signalling in the ear. We will also re-examine the mouse Shh mutant phenotype using markers of AP pattern to see whether Hh signalling plays a role in otic AP patterning, in addition to its previously characterised DV patterning role. Finally, we will test the hypothesis that the dorsolateral otic defects we observe in zebrafish Hh inhibitor mutants reflect the inappropriate repression of Wnt signalling in the dorsal part of the ear, using a zebrafish transgenic reporter line in which Wnt activity can be visualised directly. The work will contribute to our understanding of the fundamental problems of axis formation during organogenesis and addresses many of the scientific remits of the BBSRC.
Publications
Hammond KL
(2010)
Repression of Hedgehog signalling is required for the acquisition of dorsolateral cell fates in the zebrafish otic vesicle.
in Development (Cambridge, England)
Hammond KL
(2011)
Fgf and Hh signalling act on a symmetrical pre-pattern to specify anterior and posterior identity in the zebrafish otic placode and vesicle.
in Development (Cambridge, England)
Hammond KL
(2009)
Expression of zebrafish hip: response to Hedgehog signalling, comparison with ptc1 expression, and possible role in otic patterning.
in Gene expression patterns : GEP
Stooke-Vaughan GA
(2012)
The role of hair cells, cilia and ciliary motility in otolith formation in the zebrafish otic vesicle.
in Development (Cambridge, England)
Description | The inner ear is important for the senses of balance and hearing. It consists of an intricate labyrinth housing a variety of extraordinarily sensitive sensory structures that respond to sound, movement and gravity. It is essential that each of these components form in the embryo in exactly the right place, as any disturbance can lead to deafness or balance disorders. This project aimed to understand how the inner ear develops, and the mechanisms that ensure that the different cell types in the ear arise in the correct positions. The project used embryos of a small tropical fish, the zebrafish, as a model system. It focused on understanding the effects of Hedgehog (Hh) proteins on ear development. Hh proteins are signalling molecules that give instructions to cells in the embryo, telling them how and where to develop. We aimed to confirm a role for Hh signalling in ear patterning using a novel and unique panel of zebrafish mutants. This led to a greater understanding of how the inner ear develops in the vertebrate embryo, providing important basic knowledge that will help to inform clinical research into the many genetic conditions that lead to deafness in humans. |
Exploitation Route | Education of the public in developmental biology and basic research through events such as open days, public lectures, exhibitions, careers events and STEM clubs. Used by other researchers to gain a greater basic science understanding of ear development in the embryo, and of the role of signalling molecules in developing systems. An improved knowledge base contributes to our understanding of disease processes. This work has also been used to underpin further grant proposals. |
Sectors | Education,Healthcare,Other |
Description | Findings have been used to increase knowledge of mechanisms of otic development and to underpin further research in this area. |
First Year Of Impact | 2007 |
Sector | Education,Healthcare,Other |
Impact Types | Cultural |
Description | Modelling axial polarity in the developing zebrafish ear |
Amount | £75,000 (GBP) |
Funding ID | 1322544 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 10/2012 |
End | 09/2016 |
Description | University of Sheffield collaboration: Dr Freek van Eeden |
Organisation | University of Sheffield |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | Research collaboration, characterising the phenotype of new zebrafish mutant lines created in the van Eeden lab. Led to joint publication in Development (Hammond et al., 2010). |
Start Year | 2009 |
Description | Conference Talk (Madison, WI, USA) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2010: 9th International Zebrafish Development and Genetics Meeting, Madison, WI, USA no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |
Description | Conference Talk BSDB 2010 |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | 2010: BSDB Autumn Meeting: Development of the Senses, Oxford, UK no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |
Description | EMBO Practical Course: Animal models for physiology and disease |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | EMBO Practical Course no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |
Description | Invited seminar (Cincinnati, OH, USA) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2010: Division of Developmental Biology, Cincinnati Children's Hospital, Cincinnati, OH, USA no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |
Description | Invited talk (Uruguay) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2010: 9th International Congress on Vertebrate Morphology, Punta del Este, Uruguay (given by K. Hammond) no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |
Description | Invited talks and presentations |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2009: University of Konstanz, Germany no actual impacts realised to date |
Year(s) Of Engagement Activity | 2009 |
Description | Invited talks and presentations |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2009: 18th CDB Meeting: Common Themes and New Concepts in Sensory Formation, Kobe, Japan no actual impacts realised to date |
Year(s) Of Engagement Activity | 2009 |
Description | Invited talks and presentations |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | 2008: Department of Zoology, University of Cambridge, UK no actual impacts realised to date |
Year(s) Of Engagement Activity | 2008 |
Description | Invited talks and presentations |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Type Of Presentation | Paper Presentation |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2010: BSDB Autumn Meeting: Development of the Senses, Oxford, UK 2010: 9th International Zebrafish Development and Genetics Meeting, Madison, WI, USA 2010: 9th International Congress on Vertebrate Morphology, Punta del Este, Uruguay (given by K. Hammond) 2010: Workshop: Development of pattern in the nervous system, Minerve, France 2010: Division of Developmental Biology, Cincinnati Children's Hospital, Cincinnati, OH, USA 2009: Universitat Pompeu Fabra, Barcelona, Spain 2009: University of Konstanz, Germany 2009: 18th CDB Meeting: Common Themes and New Concepts in Sensory Formation, Kobe, Japan 2008: Department of Zoology, University of Cambridge, UK no actual impacts realised to date |
Year(s) Of Engagement Activity | 2007 |
Description | Invited talks and presentations |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2009: Universitat Pompeu Fabra, Barcelona, Spain no actual impacts realised to date |
Year(s) Of Engagement Activity | 2009 |
Description | Public exhibition |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | Yes |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | 2009: Royal Society Summer Science Exhibition, London: Fishing for Clues: Why Medical Researchers are Glowing with Excitement (MRC CDBG entry). MRC CDBG team exhibit at the Royal Society Summer Science Exhibition 2009 Exhibition delivered to public, including live demonstrations, videos, quizzes, leaflets. Demonstration and resources have been used subsequently at many events, such as open days. no actual impacts realised to date |
Year(s) Of Engagement Activity | 2009 |
Description | Undergraduate teaching - Sheffield |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | Yes |
Geographic Reach | National |
Primary Audience | Public/other audiences |
Results and Impact | Research-led teaching to undergraduate students via lectures, tutorials and laboratory projects Encouragement of students to pursue a career in research science. |
Year(s) Of Engagement Activity | 2006,2007,2008,2009,2010 |
Description | Workshop participation (Minerve, France) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | 2010: Workshop: Development of pattern in the nervous system, Minerve, France no actual impacts realised to date |
Year(s) Of Engagement Activity | 2010 |