Dr HELEN DOOLEY
Current: Lecturer, University of Aberdeen. Aberdeen, UK
2008-2011: Principal research scientist, Pfizer Inc. Aberdeen, UK (legacy Wyeth)
2007-2008: Research associate, University of Maryland School of Medicine. Baltimore, USA
2002-2007: Post-doctoral fellow, University of Maryland School of Medicine. Baltimore, USA
2001: Scientist, Auvation Ltd. Aberdeen, UK
2001: Ph.D Antibody engineering. University of Aberdeen, UK
1998: M.Sc Antibody engineering. University of Aberdeen, UK
1996: B.Sc (hons) Genetics. University of Aberdeen, UK
For the past 10 years or so my research has focused upon trying to understand how the adaptive immune system (AIS) and its component molecules evolved. To do this I use a comparative approach, examining a specific molecule or immune mechanism in different species across phylogeny to look for shared features and/or general rules governing their function. Key to this are our studies on the immune system of the cartilaginous fishes (sharks, skates and rays); this group, comprising over 700 living species, diverged from a common ancestor with other jawed vertebrates approximately 500 million years ago. They are also the most ancient species to have a ‘mammalian-like’ adaptive immune system, using somatic recombination to generate diverse repertoires of antibodies and T-cell receptors.
Phylogeny of the four shark species used in my work. Nurse shark and bamboo shark are the most closely related, both being members of the order Orectolobiformes. In contrast nurse shark and spiny dogfish are estimated to have diverged from a common ancestor over 200 million years ago.
My work on the nurse shark (Ginglymostoma cirratum) showed for the first time that cartilaginous fish mount a highly complex, multi-layered serum antibody response complete with affinity maturation and immunological memory following antigen challenge. We have since expanded our studies to include three other, phylogentically divergent, shark species (the brown-banded bamboo shark Chiloscyllium punctatum; the small-spotted catshark Scyliorhinus canicula; and the spiny dogfish Squalus acanthias) in order to compare their immune response and look for features that are conserved amongst the different species. In my future work I want to look in more depth at how the different antibody isotypes combine to protect sharks from infection; of immediate interest are the antibodies that protect the mucosal surfaces of the gill and gut and thus constitute the sharks ‘first line of protection’ from pathogen invasion.
Another aspect that is of particular interest to me is IgNAR, a novel antibody isotype found only in the cartilaginous fishes. Unlike conventional antibodies found in mammals and other species, IgNAR is a heavy-chain homodimer which does not associate with light chains; instead it binds to its antigen via a pair of highly soluble, single-domain binding regions. My work showed that phage-displayed libraries of these binding regions (so-called VNARs) can be constructed from suitably immunised animals and selected for clones that bind with high affinity and specificity to the chosen target. Additionally, that via their unusually long CDR3 loop (the main determinant for antigen binding), VNARs can bind to epitopes on antigens (such as enzymatic active sites) that are inaccessible to conventional antibodies.
Comparison of (a) a mouse Fv antibody fragment and (b) a shark VNAR bound to the enzyme hen egg-white lysozyme (HEL). The VNAR actually inserts its CDR3 loop into the active site of HEL, blocking the interaction with its natural oligosaccharide ligand (c; bound oligosaccharide shown in red), and inhibiting its enzymatic activity.
Due to their small size and the presence of addition disulphide bonds VNAR domains are much more stable than conventional antibodies, retaining high levels of functionality after extended periods at temperatures in excess of 70°C. I plan to exploit these unusual innate properties of VNARs to generate new and/or improved diagnostic and therapeutic reagents.
Contributions to Journals
- Crouch, K., Smith, LE., Williams, R., Cao, W., Lee, M., Jensen, A. & Dooley, H. (2013). 'Humoral immune response of the small-spotted catshark, Scyliorhinus canicula'. Fish & Shellfish Immunology, vol 34, no. 5, pp. 1158-1169.
[Online] DOI: 10.1016/j.fsi.2013.01.025
- Li, R., Wang, T., Bird, S., Zou, J., Dooley, H. & Secombes, CJ. (2013). 'B cell receptor accessory molecule CD79a: characterisation and expression analysis in a cartilaginous fish, the spiny dogfish (Squalus acanthias)'. Fish & Shellfish Immunology, vol 34, no. 6, pp. 1404-1415.
[Online] DOI: 10.1016/j.fsi.2013.02.015
- Castro, CD., Ohta, Y., Dooley, H. & Flajnik, MF. (2013). 'Noncoordinate expression of J-chain and Blimp-1 define nurse shark plasma cell populations during ontogeny'. European Journal of Immunology.
[Online] DOI: 10.1002/eji.201343416
- Li, R., Dooley, HM., Wang, T., Secombes, CJ. & Bird, S. (2012). 'Characterisation and expression analysis of B-cell activating factor (BAFF) in spiny dogfish (Squalus acanthias): cartilaginous fish BAFF has a unique extra exon that may impact receptor binding'. Developmental and Comparative Immunology, vol 36, no. 4, pp. 707-717.
[Online] DOI: 10.1016/j.dci.2011.11.010
- Smith, LE., Crouch, K., Cao, W., Müller, MR., Wu, L., Steven, J., Lee, M., Liang, M., Flajnik, MF., Shih, HH., Barelle, CJ., Paulsen, J., Gill, DS. & Dooley, H. (2012). 'Characterization of the immunoglobulin repertoire of the spiny dogfish (Squalus acanthias)'. Developmental and Comparative Immunology, vol 36, no. 4, pp. 665-679.
[Online] DOI: 10.1016/j.dci.2011.10.007
- Goodchild, SA., Dooley, H., Schoepp, RJ., Flajnik, M. & Lonsdale, SG. (2011). 'Isolation and characterisation of Ebolavirus-specific recombinant antibody fragments from murine and shark immune libraries'. Molecular Immunology, vol 48, no. 15-16, pp. 2027-2037.
[Online] DOI: 10.1016/j.molimm.2011.06.437
- Dooley, HM., Buckingham, EB., Criscitiello, MF. & Flajnik, MF. (2010). 'Emergence of the acute-phase protein hemopexin in jawed vertebrates'. Molecular Immunology, vol 48, no. 1-3, pp. 147-152.
[Online] DOI: 10.1016/j.molimm.2010.08.015
- Flajnik, MF. & Dooley, H. (2009). 'The generation and selection of single-domain, v region libraries from nurse sharks'. Methods in Molecular Biology, vol 562, pp. 71-82.
[Online] DOI: 10.1007/978-1-60327-302-2_6
- Stanfield, RL., Dooley, H., Verdino, P., Flajnik, MF. & Wilson, IA. (2007). 'Maturation of shark single-domain (IgNAR) antibodies: evidence for induced-fit binding'. Journal of Molecular Biology, vol 367, no. 2, pp. 358-372.
[Online] DOI: 10.1016/j.jmb.2006.12.045
- Dooley, H., Stanfield, RL., Brady, RA. & Flajnik, MF. (2006). 'First molecular and biochemical analysis of in vivo affinity maturation in an ectothermic vertebrate'. PNAS, vol 103, no. 6, pp. 1846-51.
[Online] DOI: 10.1073/pnas.0508341103
- Dooley, H. & Flajnik, MF. (2006). 'Antibody repertoire development in cartilaginous fish'. Developmental and Comparative Immunology, vol 30, no. 1-2, pp. 43-56.
[Online] DOI: 10.1016/j.dci.2005.06.022
- Dooley, H. & Flajnik, MF. (2005). 'Shark immunity bites back: affinity maturation and memory response in the nurse shark, Ginglymostoma cirratum'. European Journal of Immunology, vol 35, no. 3, pp. 936-45.
[Online] DOI: 10.1002/eji.200425760
- Stanfield, RL., Dooley, H., Flajnik, MF. & Wilson, IA. (2004). 'Crystal structure of a shark single-domain antibody V region in complex with lysozyme'. Science, vol 305, no. 5691, pp. 1770-3.
[Online] DOI: 10.1126/science.1101148
- Rumfelt, LL., Lohr, RL., Dooley, H. & Flajnik, MF. (2004). 'Diversity and repertoire of IgW and IgM VH families in the newborn nurse shark'. BMC Immunology, vol 5, pp. 8.
[Online] DOI: 10.1186/1471-2172-5-8
Chapters in Books, Reports and Conference Proceedings
- Müller, MR., O'Dwyer, R., Kovaleva, M., Rudkin, F., Dooley, H. & Barelle, CJ. (2012). 'Generation and isolation of target-specific single-domain antibodies from shark immune repertoires'. in P Chames (ed.), Methods in Molecular Biology: Antibody Engineering. vol. 907, Methods and Protocols, Humana Press, pp. 177-194.
[Online] DOI: 10.1007/978-1-61779-974-7_9
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