RA5a: Structure,environment and staffing policy
RA5a Research Structure, Environment and Staffing Policy:
The School of Biosciences was formed in 1999 by merging the Schools of Biological Sciences and Biochemistry. The new School’s research spans from biophysics to ecology and is grouped into five Themes: Structural Biology (SB); Molecular Microbiology (MM); Molecular Cell Biology (MCB); Plant Genetics & Cell Biology (PGCB); and Environmental Biology & Toxicology (EBT). Themes are communities of inter-related research groups, but individual research activities often combine interests from several Themes (see RA1). Research Groups principally affiliate to one Theme, and affiliations change as needed. Themes are foci for promoting specific research activities and collaborations, including seminars and training, for developing research strategy and for co-ordinating applications for funding. The key aims of our research strategy are:
• To promote and support research of international excellence in every Theme.
• To champion new research opportunities by catalysing scientific developments and interactions.
• To enhance our position as an internationally recognised centre for research training.
The Research Management Committee [RMC; including the Director of Research (Heath), Head of School (Trayer) and Theme Leaders] oversees the School’s research. It defines research strategy and co-ordinates research across Themes. To fulfil the above aims: we recruit, train and retain the best staff, whose talents match our scientific aims; we provide state-of-the-art infrastructure for research; and we promote a pervasive and vigorous research culture. The RMC’s roles include targeting major funding initiatives, recruiting excellent staff, coordinating research training and interacting with stakeholders and research users (especially research councils, charities and industry).
Recruitment and Staff Support: We currently have 50.7 FTE research-active staff. Retirements and new posts will allow ~20 new appointments from 2000 to ~2005, leading to ~65 HEFCE-funded staff (plus Fellows). By 2005, more than 40% of staff will have been in post for less than 10 years. The new School’s Theme-led research strategy defines clear foci for research development. During 1999-2000, Themes reviewed their scientific strengths, weaknesses and future priorities, and ~10 ‘target areas’ were identified in which to recruit staff to enhance the quality of existing research. Five appointments have been made since the merger: Blackburn (theoretical population ecology; from Imperial College); Filatov (population genetics; from Edinburgh University); Fütterer (protein crystallography; from Washington University, USA); Luo (quantitative genetics; from Fudan University, China) and Schrewe (from MPI Frieburg); O. Smart will take up a lecturership in Bioinformatics from June 2001. Recruitment is now under way in ‘toxicogenomics', ‘genetic analysis of eukaryotic signalling’, ‘microbial pathogenicity’, and ‘molecular ecology’. From 2002-4, we will a) recruit outstanding individuals with expertise in relevant new areas and/or technologies; and b) reinforce links between Themes and cement partnerships with other Schools (e.g. Medicine; Geography & Environmental Science; Chemistry; Mathematics & Statistics; Computer Science) – possibly with some joint appointments. We anticipate one or more professorial appointments but most appointees will be lecturers/senior lecturers.
Individuals holding independent personal research fellowships make major contributions to our research productivity and scientific vitality. We aim to recruit and support an increasing number of such Fellows in all Themes. At present there are 6 such Fellows (Dove, Insall, Kinnunen, Machesky, Michell, Turnbull): applications from others are being considered by relevant agencies. Since 1996, research achievement has been recognised by the promotion of several staff to Personal Chairs (Chipman, Franklin, Kearsey, Penn, Strain) and Readerships (Macaskie, Turnbull, Wharton).
New staff and fellows are housed in high quality laboratories with access to shared core technical resources and equipment, are given ‘start up’ equipment funding (typically £10-15k), and have light teaching and administrative loads for three years. They are encouraged to apply for local funding (e.g. Science and Medical Faculties) to ‘pump-prime’ their consumable costs, and Mentors advise on obtaining grants. Most of the recently appointed staff have quickly won peer-reviewed grants from research councils or charities. The University provides training in academic skills (interviewing/recruiting, safety, grant writing, funding opportunities with industry and the EU, etc.). We also emphasise career support for postdoctoral researchers. They are represented on key School Committees and participate in a Postdoctoral Forum to articulate training and support needs. In response, we have developed training in teaching, career management and grant writing.
Research infrastructure: About 60% of our existing laboratories have been refurbished since 1995, at a cost of ~£10m, mostly from external sources: the JIF scheme (Plant & Microbial Genomics); Wolfson Foundation and HEFCE (Molecular Microbiology & Environmental Biology); Glaxo-Wellcome Foundation (Bioinformatics). Priorities for the next 3 years are: a) to finish modernising our existing laboratories; b) following completion of an adjacent Teaching Resource Centre, to convert ex-teaching space to research; and c) to build new laboratories in which to co-locate research in molecular cell biology, genomics and structural biology. We have identified a site onto which to extend our building, and are bidding for much of the funding from the University’s £20.3m SRIF allocation. Recent investments in major equipment are detailed in RA6.
Collaborative research - internal and external: The emphasis on promotion of research excellence within the School can only be fulfilled by maintaining strong external collaborations. Formal and informal research networks are a great strength of this University's Life Sciences community - particularly the links between Biosciences and Medicine. Our two MRC Co-operative Groups (see MCB Theme) exemplify the research links between these Schools, and some members of Biosciences (Dove, Kirk, Michell, Strain) have their laboratories in the Medical School. Our research will benefit directly from new developments in Medicine: the JIF-funded Institute of Biomedical Sciences (£18m) and 900 MHz Biomedical NMR facility (£5.7m). Geography has just moved into a new extension of the Biosciences building, consolidating the research links in environmental sciences and eco-toxicology (NERC-funded). There are growing research links with Computer Science in database mining and image processing (EPSRC-funded).
Among the collaborations with other UK research institutions are the ESRF synchrotron allocation (see RA6) and formal links with Horticulture Research International, John Innes Centre, Roslin Institute, Institute of Grassland & Environmental Research and British Antarctic Survey. The School participates enthusiastically in regional collaborations and Midlands research initiatives - e.g. with cognate Departments at Warwick, Leicester and Nottingham. There is peer-reviewed support for several international collaborations [e.g. from Human Frontiers Science Programme (Machesky, Turnbull); EU (Butler, Macaskie, Machesky, Taylor, Turnbull)], which yield numerous co-authored publications. A major aim is to enhance the School's research links with industry and other international agencies: Ford-Lloyd, Newbury & Kearsey maintain long-term funded collaborations with research centres of the Consult. Gp. of Intl. Agric. Res., e.g. IRRI (Manila); ICRISAT (Hyderabad). Many of our industrial research links involve funding for research collaborations and postgraduate students.
RESEARCH ACTIVITIES, 1996-2000: *denotes staff appointed since the 1996 RAE
STRUCTURAL BIOLOGY (SB): *Fütterer, Hyde, Jackson, Levine, Trayer (Theme Leader), Wharton, *White.
This Theme studies protein-protein and protein-ligand interactions on the molecular scale, yielding detailed understanding of fundamental processes: e.g. enzyme activity and control; gene regulation; energy conservation; signal transduction. A spectrum of biophysical techniques is employed: X-ray crystallography, spectroscopy (NMR, fluorescence, UV-VIS, IR, stopped-flow), microcalorimetry, SPR etc., with protein-chemical and molecular biology methods, computational modelling and bioinformatics. The first Birmingham-based solutions of high-resolution protein structures appeared recently. The new 900 MHz NMR facility, and the appointment of an outstanding NMR spectroscopist, will greatly enhance our NMR opportunities. The appointment of Smart to a HEFCE-funded position (from June, 2001) will enhance the Theme's computational capabilities, especially in the areas of ion channels and peptide design.
Transhydrogenase (TH) is an accessible model for the study of biological ion pumps. Bacterial TH provides biosynthetic NADPH, and mammalian muscle TH is involved in regulating Krebs cycle flux. Jackson’s laboratory initially determined three structures crystallographically and by NMR: the human and bacterial NADP(H)-binding subunits (c. *White and Trayer) and the bacterial NAD(H)-binding subunit (c. Rice, Sheffield). Jackson and *White recently published the first crystallographic structure of the complex. Asymmetries in the structure and novel modes of nucleotide binding, together with kinetic and thermodynamic studies (sometimes c. Rydstrom, Goteborg), have revealed that the proton-pumping step is linked to a change in NADP(H) binding energy. Protein engineering experiments (c. Thomas, MM Theme) are analysing the roles in H+ pumping of key amino acids, some of which were pinpointed through recently solved structures. Structure determinations on the complete, membrane-spanning TH are now underway.
*White has also determined the structure of a novel type of prokaryotic inorganic pyrophosphatase: it has a 2-domain structure with a buried active site at the interface. Future work will focus on mechanistic studies and inhibitor design. *White and Hyde solved the X-ray structure of nitroreductase and are constructing mutants to improve its potential for anticancer gene therapy (c. Searle, CRC Inst. Cancer Studies). *Fütterer obtained structures of an N-myristoyltransferase, the tandem SH2 domains of the Syk protein-tyrosine kinase and a bacterial pilus/chaperone complex during his post-doctoral work (c.Waksman, Washington U.). He is now focusing on signalling proteins such as the Arp2/3 complex with MCB Theme members.
Hyde primarily analyses DNA-binding proteins by NMR. Her group characterised complexes of Trp repressor with operator and co-repressor variants. With Dixon (John Innes, Norwich), they determined the secondary structure and DNA-binding of the C-terminal domain of NifA, an activator of transcription of nitrogen fixation genes. Future work will determine the tertiary structure of NifA and examine its other domains. With Busby (MM Theme), she is also examining transcription activation by MelR. She determined the structures of a potent inhibitor of platelet aggregation and is examining variants, to identify features essential for activity (c. Lu, Thrombosis Res. Inst., London).
Wharton has extended his IR spectroscopic work on the reaction mechanism of serine proteases to that of b-lactam antibiotic resistance (c. Hoffmann-La Roche, Basel). Transpeptidases (the antibiotic targets) and the resistance-conferring b-lactamases both tolerate multiple mutations and so adapt to antibiotic challenge. Experiments to observe reaction intermediates and protein conformational changes are interpreted in terms of a bacterium’s need to minimise transpeptidase inhibition, whilst allowing flexible b-lactamases to hydrolyse multiple antibiotics. Other rapid reaction methods are being developed for studying highly reactive systems. QM/MM calculations with Smart will enhance interpretation of IR spectra.
Trayer and Levine investigate regulation of the contractile activity of vertebrate striated muscle, especially human heart. Trayer's mutational studies showed that the N-terminal region of A1-type myosin light chains bind actin through the 4 N-terminal residues and modulate myosin motor function via an extended (X-P)7 tandem repeat sequence next to the actin-binding motif. Protein engineering of troponin-I (TnI) and troponin-T defined the interactions with troponin-C (TnC) that lead to muscle activation. He also showed how TnI phosphorylation regulates TnC/TnI interactions. Future work will concentrate on how TnI phosphorylation and Ca2+ binding to TnC aid cardiac muscle relaxation (c. Miller, Glasgow). Levine's NMR studies defined the ordered dual-site phosphorylation of TnI and how this reduces TnC’s Ca2+ affinity. Similar events that underlie smooth muscle regulation by caldesmon are being investigated (c. Marston, Imperial).
MOLECULAR MICROBIOLOGY (MM): Brown (Theme Leader), Busby, Cole, Lund, Macaskie, Minchin, Penn, Sweet, Thomas.
This Theme studies fundamental processes in microbial systems and the roles and applications of these in pathogenesis and in the environment. Increasingly, the work exploits genome sequences. A new appointment will expand and further strengthen activities in these areas.
Busby has revealed detailed molecular mechanisms of transcription activation in bacteria. Multiple contacts between the activators, CRP and FNR, and RNA polymerase provide the major mechanism that couples bacterial promoters to different transcription factors: different activating regions make different contacts with RNA polymerase. Collaborations with Cole and Hyde (SB theme), respectively, revealed novel synergistic mechanisms whereby a second activator works with FNR, preventing repressors from binding to operator sites, and how two activators, MelR and CRP, interact directly. He defined (with Minchin) how promoters with an extended –10 sequence are recognised by sigma factors and the rules for sequence specificity and the rules for effective alpha-CTD/DNA and sigma/DNA interactions at different classes of promoter (c. Ishihama, Japan).
Cole is exploring the roles of multiple E. coli nitrate (NR) and nitrite reductases. Discoveries include: the selective advantage conferred by periplasmic NR to enteric pathogens during nitrate-limited growth; the differential roles of the two-component regulatory systems, NarXL and NarQP, in regulating gene expression; haem ligation to the active site of nitrite reductase by a new haem lyase, NrfEFG; and the identification of a secretion pathway for partially-folded proteins containing redox cofactors. These studies are being extended to pathogenic neisseria and mycobacteria and, with Penn, to Campylobacter.
Lund showed that ring-ring interactions in the essential protein GroEL can be reversibly disrupted. This supports models of ring-to-ring transmission of an allosteric change for GroES-dependent substrates. By varying chaperone levels in vivo, he showed (c. Fersht, Cambridge) that small GroEL-derived domains partially complement GroEL loss. E. coli is being developed for the study of eukaryotic protein-folding enzymes: the ER protein-disulphide isomerase (PDI) complements loss of the isomerase DsbC, and allows rapid characterisation of PDI mutants. Future work will include whole genome analyses of the heat-shock response and studies on chaperones in extremophiles.
By sequencing R751, a broad host range IncPß plasmid, Thomas and Lanka (MPI Mol. Genet., Berlin) defined conserved elements of this complex plasmid family revealing genomic motifs relevant to plasmid evolution and transposable element acquisition. Immunofluorescence microscopy showed how the IncP partitioning system directs plasmids from one replication zone to the next before cell division. With Jagura-Burdzy (Inst. Biochem. Biophys., Warsaw), he showed that the partitioning protein KorB represses transcription at a distance. It interacts with RNA polymerase and with plasmid-encoded proteins IncC (the driver of partitioning) and TrbA and KorA regulators through a conserved domain. Future work will focus on how mobile elements are maintained in bacterial populations and will monitor their activity in bioremediation and the spread of antibiotic resistance.
Penn investigates bacterial pathogens. He identified a lipoprotein antigen of the gastric pathogen Helicobacter pylori as a flagellar sheath component, and is exploring the regulation of flagellar gene expression. Iron restriction of the food-borne pathogen Campylobacter jejuni led to identification of AhpC, an alkyl hydroperoxidase that protects against oxygen toxicity, and of PerR, a Fur homologue. He contributed to annotation of the Sanger Centre’s C. jejuni genome. He continues to exploit high throughput analyses of these pathogens (c. Wren, SHTM, London; Ketley, Leicester; & van Vliet, Amsterdam). Sweet established that the influenza virus neuraminidase (NA) acts at least twice in induction of apoptosis in epithelial cells: early in replication, and late in the infection cycle to remove sialic acid from virion haemagglutinin and enhance infection. Other involved virion components are are being identified. Anti-NA compounds (Gilead/Hoffman La Roche and Biocryst/Johnson Pharmaceuticals) were evaluated and are close to clinical use. Reverse genetics will be used to define the roles of 'flu proteins in fever and apoptosis. Murine cytomegalovirus (MCMV) ts mutants elicit an immune response and protect against virulent virus challenge, but produce no infectious virus: virulence of MCMV will be further explored through BAC-cloning.
Brown analyses the regulation of gene expression in bacterial metal resistance. He explained how expression of the metal resistance determinants pco and czc (c. Nies, Halle) and the major E. coli zinc exporter are regulated, and now focuses on regulators in the MerR family. A unitary model for the mechanism of Hg resistance transport proteins is being developed. With Mattiasson (Lund), a novel protein-based metal biosensor was patented that may delineate interactions between metal ions and proteins. Future work will focus on the control of metal ion homeostasis in E. coli - particularly for Cu at the whole genome level - and on how proteins discriminate between metals. He also helped local clinicians to identify Alliococcus otitis, a novel pathogen, as a possible cause of Otitis Media.
Macaskie investigates microbial interactions with heavy metals and radionuclides - to develop industrial and environmental applications such as bioremediation of pollutants and bioprocess modelling. Where traditional technologies had failed, biological treatments removed 237Np, 239/241Pu and 99Tc from solution. Desulfovibrio desulfuricans hydrogenase was used in a novel patented electrobioreactor to recover precious metals (e.g. Pd) from solution by bioreduction. Through biochemical events at the cell surface, Citrobacter causes metal phosphate crystal deposition. Such deposition can recover uranyl phosphate from acidic mine water (for use in nickel bioremediation by ion exchange) or make calcium phosphate crystals (of potential use in bone implants: patent application).
MOLECULAR CELL BIOLOGY (MCB): *Dove, Heath, Hotchin, *Insall, *Kinnunen, *Lalioti, *Machesky, Michelangeli, Michell (Theme Leader), Publicover, *Smith, Strain, *Turnbull, Wheatley.
This Theme addresses interactions between extracellular ligands and transmembrane receptors, and how the resulting signals control intracellular events - with a growing emphasis on genetic and structural biology approaches. It has expanded through the recruitment of independent research fellows, and includes two MRC Co-operative Groups: Integrated Functions of Transmembrane Adhesion and Growth Factor Receptors [Heath, Hotchin, Turnbull, Berdetchevski (Inst. Cancer Res.)]; and Cell Regulatory Roles of Phospholipids, Phospholipases and Inositol Phosphates [Michell, Wakelam (Inst. Cancer Res.), Michelangeli, Machesky, Insall].
Heath (c. *Lalioti) analyses interactions between polypeptide growth factors and their receptors. Crystallography (c. Jones, Oxford) has yielded receptor (e.g. gp130) and ligand (e.g. oncostatin M) structures and defined the specificities of recognition sites involved in assembly of gp130 signalling complexes: structures of multimeric receptor/ligand complexes are being determined. Heath & *Schrewe continue to analyse growth factor/receptor function and activin-inducible gene expression by genetic and biophysical methods.
*Turnbull (c. *Kinnunen) analyses the roles of heparan sulphate proteoglycans (HSPGs) in cell regulation and growth factor signalling. They are defining HS biosynthesis in vivo and the specificities of HS-protein interactions, notably in the nervous system: e.g. changes in FGF-binding HS and in HS sulphotransferase expression during neuroepithelial development (c. Ford, Melbourne). He showed that specific HS sulphation patterns are needed to activate FGF signalling, patented the first HS saccharide sequencing method (c. Gallagher, Manchester; Hopwood, Adelaide) and is developing MALDI-MS-based HS sequencing (c. Sasisekharan, MIT). He will continue to define how variant HS sequences act as dynamic regulators of proteins such as FGFs (c. Heath) and has started to analyse HS structure and biosynthesis in C. elegans.
*Smith aims to define the roles of stem cells and their growth factor regulation in skeletal muscle and to devise therapeutic strategies for human myopathies and ageing. Having identified IGF-II as an SMSC survival factor, she showed that SMSC apoptosis increases in dystrophic mdx mice: IGF-II reduces this and ameliorates the dystrophy (c. Ward, Bath).
Strain uses human liver epithelial cells to examine important clinical questions in liver pathophysiology. He showed that human liver contains bipotent stem cells that can differentiate to hepatic epithelium. Human hepatocytes (for transplantation, gene therapy, toxicology, bioartificial liver systems, etc.) are scarce, and such studies, complemented by studies with embryonal cell-lines, may allow the development of alternative sources. He is developing models of human hepatic epithelial morphogenesis (c. Auth, Essen; Fabris, Padua),
Wheatley analyses the structure and function of G protein-coupled receptors (GPCR) for vasopressin (AVP) and oxytocin. He identified the first extracellular loop as part of the AVP binding site and also an N-terminal sub-domain that is critical for agonist, but not antagonist, binding. He developed linear V1a antagonists and chimeric ligands that co-incorporate vasopressin with other peptides (c. Langel & Östenson, Stockholm). The latter are functionally distinct from both progenitors and offer a novel strategy for ligand design. He improved V1R sub-type selectivity and stability by including unusual amino acids and retro-inverso analogues. Future work will explore the binding site architecture and the complex effects of post-translational modification on GPCR function.
Hotchin explores how small GTPases of the Rho/Rac/Cdc42 family regulate epithelial cell responses (migration, differentiation, etc.) following ligation of cadherins or integrins. He showed that Rho- and Rac-mediated responses require specific integrin-ECM interactions (c. Mardon, Oxford). Assembly and maintenance of cadherin-based adherens junctions require activated Rho and Rac1 (c. Braga, UCL), and he showed that clathrin-independent endocytic mechanisms are needed for Rac1 regulation. To define the role of Rac1 in cell migration in vivo, GFP-Rac1 was expressed in chick myoblast precursors and cell movement followed by high resolution imaging (c. Horwitz, Virginia). Having shown that a major H. pylori virulence factor needs Rac1 for activity, Cover (Vanderbilt) & Hotchin are defining how H. pylori attachment activates epithelial Rho signalling.
Michell & *Dove discovered the ‘novel’ phosphoinositide, phosphatidylinositol 3,5-biphosphate (PtdIns(3,5)P2) (c. Parker, ICRF). Though found in yeast, where its synthesis contributes to intra- cellular vesicle trafficking and osmotic stress responses, PtdIns(3,5)P2 is ubiquitous in eukaryotes. They defined Fab1p and its animal and plant orthologues as the Type III PtdInsP kinases that make PtdIns(3,5)P2. They now aim to identify protein targets for PtdIns(3,5)P2 and to define its role in membrane trafficking. Dove & Pical (Lund) also observed dramatic effects of saline stress on phosphoinositide turnover in Arabidopsis, and Michell identified a novel PtdIns(1,4,5)P3 kinase that makes InsP6 - and may be implicated in nuclear mRNA export. With Brown (Immunology), Michell defined a proliferative burst that occurs immediately after promyeloid cells commit to differentiation.
*Machesky and *Insall explore how control of the actin cytoskeleton - which provides structural support and generates force - dictates eukaryotic cell motility. *Machesky discovered a major signalling pathway to actin polymerisation (c. Pollard, La Jolla). Downstream of activated receptors, WASP family proteins signal to the actin-organising Arp2/3 protein complex, inducing polymerisation. Small GTPases, tyrosine kinases and heterotrimeric G-proteins target this mechanism. She implicated these events in the assembly of lamellipodia and stress fibres, in phagocytosis and in the intracellular motility of Listeria – and she now aims to define the molecular links between receptors, signal transducers, actin assembly and motility in multiple cell contexts (c. *Futterer, SB Theme). Multicentre EU and Human Frontiers grants (co-ordinator *Machesky: 7 participants world-wide) will facilitate this work. *Insall is exploiting genetics to understand Dictyostelium motility, especially roles of the Arp2/3 complex (c. Gerisch, Martinsried) and Ras-related proteins. Ras gene disruptions gave diverse defects: in polarity and cytokinesis (RasG-); in phototaxis (RasD-); and in phagocytosis and cell movement (RasS-). He identified and disrupted multiple RasGEFs and is now defining functions of the Ras proteins and characterising Ras targets (c. Weeks, Vancouver; Firtel, UCSF). He is also examining lipid-based signalling in Dictyostelium (c. Wakelam, Cancer Inst.; Cockroft, UCL).
Michelangeli investigates the diversifying families of Ca2+ pumps and channels. He showed that mastoparan inhibits ER ATPase activity and abolishes co-operative Ca2+ binding (c. Langel, Stockholm). The Ca2+-binding geometry of ER Ca2+ pumps has been uncertain, and this result suggested a side-by-side arrangement - as was recently shown crystallographically. He showed that some popular Ins(1,4,5)P3 receptor agonists are partial agonists, suggesting that channels permit ‘unproductive’ ligated state(s). With Galione (Oxford), he showed that cADP-ribose-activated and NAADP-activated intracellular Ca2+ channels are kinetically and pharmacologically distinct. 2-hydroxycarbazole was identified as a more potent ryanodine receptor inhibitor than caffeine, and others adopted it. Future work will analyse the mechanisms that regulate these transporters.
Publicover focusses on signalling in bone cells and sperm. He established that mechanotransduction in bone involves particular voltage-operated Ca2+ channels (c. El Haj, Keele; Zamponi, Calgary) and demonstrated glutamatergic signalling through multiple receptors in osteoblasts. With Barrett (Assisted Conception Unit), he showed biphasic Ca2+ influx into human sperm during the progesterone-induced acrosome reaction. They found novel a1G and a1H Ca2+ channel subunits in male germ cells and are now investigating activation of these channels and other pathways by zona pellucida, in the hope of obtaining a lead for male contraceptive development.
PLANT GENETICS AND CELL BIOLOGY (PGCB): Caten, *Filatov, Ford-Lloyd, Franklin, Franklin-Tong, Green, Jones, Kearsey (Theme Leader), *Luo, Newbury.
This Theme integrates basic and applied research into the genetics and natural variation of higher plants and fungi, and characterises cell signalling. Major foci are Arabidopsis and Brassica relatives and international (e.g. with IRRI, ICRISAT, ICARDA) and national (e.g. JIC, HRI, IACR, IGER) collaborative projects on rice, grain legumes and other crops. The new JIF-funded Genomics facility allows high throughput genotyping, sequence, transcriptome and proteome analysis. SNPs will be used for fine genetic mapping of Arabidopsis, brassicas and rice. Transcriptome analysis, in conjunction with substitution lines, will be used to locate and identify quantitative trait loci (QTL) and with proteomics, further define plant meiosis. A future aim is to develop new multi-site programs to explore natural variation in Arabidopsis and brassicas and to improve knowledge of rice genetics.
The Reproductive Biology group focuses on self-incompatibility (SI) and meiosis. Studies using Papaver rhoeas have established SI and the control of pollen tube growth as models for elucidating plant signalling cascades. Franklin has used site–directed mutagenesis of the stigmatic S-protein ligand to identify residues involved in pollen recognition and confirmed the key role of the pollen S-protein binding protein in mediating the SI reaction. Franklin-Tong and Franklin are investigating the signals that arrest tip growth of incompatible pollen - protein kinase activation and programmed cell death are involved. Imaging revealed that Ca2+ is a second messenger in the pollen tube SI response and implicated Ins(1,4,5)P3-regulated Ca2+ pools. Franklin-Tong is now analysing (c. Staiger, Purdue) how these signals regulate the polymerisation status of the actin cytoskeleton. Franklin (c. Ride) found a family of genes homologous to poppy S protein (SPHs) in Arabidopsis. Antisense experiments suggest that SPHs may regulate aspects of plant development. Jones & Franklin (c. Santos, Madrid) have developed cytogenetic techniques in Arabidopsis that have been used to identify and characterise 32 meiotic mutants. They have cloned several meiotic genes, including ASY1 a gene involved in homologous pairing. Antibodies raised against recombinant meiotic proteins are being used in conjunction with the mutants to investigate recombination and synapsis.
The Comparative Genomics and Genetic Variation group study marker gene mapping and QTL analysis, and applications in biodiversity, conservation and breeding. Jones has used FISH to develop an Arabidopsis karyotype that permits chromosome identification, and defined a cytogenetic map of the short arm of chromosome 4. With Kearsey, he is attempting to integrate genetic and physical maps of B. oleracea. Multiple probes, many linked to mapped RFLPs have been placed on all nine chromosomes, allowing orientation with genetic linkage maps. Jones & Kearsey (c. JIC and HRI) are attempting to contig three Brassica BAC libraries and to compare the maps with Arabidopsis. Kearsey (c. JIC and HRI) has developed resources to exploit the synteny of Arabidopsis and Brassica to facilitate the detailed genetical analysis of QTL. A B. oleracea genetic map (comprising 547 markers) was used to locate QTL controlling traits such as flowering time, size (Kearsey) and transformation ability (Newbury). Some map to regions syntenous to candidate Arabidopsis genes. To locate B. oleracea QTL more precisely, Kearsey used mapped markers and selective back-crossing to construct >70 chromosome substitution lines that contain short donor tracts in a constant background. These were used to refine QTL locations and revealed hidden QTL linked in repulsion. They have developed stepped recombinant lines in Arabidopsis as an international resource to enable precise QTL localisation (<1 cM). Kearsey continues to develop Web-based JAVA software for QTL (c. Roslin Inst. and Edinburgh U.). *Luo has developed statistical methods and advanced designs to study QTL with improved precision in complex populations (e.g. humans, farm animals), as well as software to construct genetic maps in autotetraploids (tetrasomic inheritance) to aid potato genetics. He leads a major project to locate human thyroid carcinoma susceptibility genes: there are associations with DNA polymorphisms in thyroid peroxidase. Using a novel statistical method for identifying marker/QTL associations, Ford-Lloyd and Newbury (c. IRRI) identified and mapped QTL for several rice traits, including agronomic performance. They found that diversity estimates can be marker-dependent and the common use of genetically mapped markers can bias diversity estimates. They found genetic diversity in a widespread species (e.g. Oryza longistaminata) correlates with rainfall (with predictive implications for climate change); and molecular markers, particularly SNPs, are invaluable in discriminating between problematic species and subspecies. *Filatov uses molecular and computational tools to study evolutionary and population genetics. He is analysing evolution of the Y chromosome and speciation, using as models the plant genus Silene and the cichlid fishes of ancient lakes. He has developed software that couples sequence editing and DNA polymorphism analysis.
The Molecular Basis of Fungal Pathogenesis group focuses on pathogen genetics and the cell biology of infection (partly c. HRI and IACR). Caten has developed methods for molecular analysis of the filamentous fungus Stagonospora nodorum (wheat glume blotch). He cloned its nitrate reductase gene (NIA1) and used it as a selectable marker for transformation: transforming DNA is targeted to the resident NIA1 gene. Unlike many other phytopathogenic fungi, targeted gene disruption and replacement is readily achieved. Gene silencing occurred when multiple copies of a truncated nitrite reductase gene were inserted at an ectopic site. This offers ways of attenuating S. nodorum gene expression that the agrochemical industry can use for validating fungicide targets. Green has used monoclonal antibodies, GFP-fusions and expression cloning to explore the molecular and cellular basis of infection structure development by biotrophic fungal plant pathogens (e.g. Colletotrichum lindemuthianum). He cloned the CIH1 gene. This encodes a glycoprotein, expressed during the biotrophic phase of bean infection, that is found in a matrix at the fungal-plant interface, is proline-rich, can be oxidatively cross-linked and binds carbohydrates. It may mimic plant cell wall proteins and fool the plant into accepting the fungal invader. He has identified a major glycine-rich glycoprotein in the C. lindemuthianum spore coat that is involved in spore adhesion to hydrophobic surfaces (e.g. plant leaf cuticle).
ENVIRONMENTAL BIOLOGY AND TOXICOLOGY (EBT): Bale (Theme Leader), *Blackburn, Butler, Callow, Chipman, Coleman, Leadbeater, Pritchard, Pullin,*Schrewe, Taylor, *Wang, Waring, Woakes.
The Theme investigates many species at many levels - from the molecular and cellular to whole organisms and macroscale ecology. It addresses fundamental (organism physiology, carcinogenesis), strategic (alien invasions) and applied (marine biofouling, pollution monitoring) problems. Core strengths in animal and plant ecophysiology and toxicology have been enhanced by appointments in Macroecology (*Blackburn) and Mouse Genetics (*Schrewe). Future appointments in Molecular Ecology and Molecular Toxicology will strengthen environmental genomics and toxicogenomics.
The Animal Physiology group investigates function at the organ and whole animal levels – through mechanistic laboratory studies and investigations of the energetics, physiology and behaviour of free-living animals. Woakes develops advanced implantable heart-rate and temperature data-loggers, which he and Butler use to estimate energy expenditure in aquatic top predators (fur seals and penguins). Body temperature decreases during penguin foraging, reducing metabolic expenditure and maybe prolonging foraging. New loggers will store behavioural, positional (GPS) and physiological data for at least a year. They will permit studies of year-round energy expenditure, hence total food requirement, and will help to quantify the effects of predators on fish stocks and of fishing on predators. New insights have been gained into the behaviour and energetics of post-breeding barnacle geese migrating from Spitsbergen to Scotland. Leg muscles develop faster than pectoral muscles in waterfowl. Butler found that these muscles develop similarly in wild and captive geese - but domestic ducks show faster pectoral muscle growth and slower leg muscle growth than mallards (their wild ancestors). Reasons for this are being sought.
Taylor is characterising central control of the vertebrate cardiorespiratory system. He showed that particular dietary lipids dramatically affect vertebrate physiology including heart function. Taylor & Butler showed that seasonal temperature variations and low environmental pH, with or without copper, adversely affect the swimming performance of salmonid fish: elevated plasma NH4+ may reduce white muscle activity. A new project will attempt to define what factors (e.g. temperature, O2 and NH4+ levels, toxic metal ions, organic xenobiotics) influence recolonisation of urban rivers by fish.*Wang works on the 'primitive', incompletely divided, circulation of amphibians and reptiles and has generated new ideas on metabolic rate regulation during periods of inactivity.
The Molecular Toxicology group analyses the molecular mechanisms of cell and genetic toxicities of pharmaceutical and environmental chemicals, investigates the genetic and dietary bases of human toxin susceptibility and develops pollutant biomarkers. Chipman showed that heterologously expressed human cytochrome P4501A2 can activate a heterocyclic amine carcinogen to DNA-damaging products, and that protectants (e.g. isoflavonoids, sulforaphane) modulate P450 and inhibit DNA oxidation. Understanding of basic mechanisms leads to development of sensitive biomarkers of pollutant impact: for example, by using a reporter gene assay based on flounder cytochrome P4501A (Chipman). cDNA arrays are being developed for investigating pollutant responses (e.g. expression of flounder 'stress' genes in pollutant-exposed fish: Chipman & Minchin). Waring identified inter-individual differences (genetic and dietary) in sulphate metabolism as likely contributors to carcinogen and neurological disease susceptibility, and identified flavonoids, which may be cancer-chemopreventive, as potent sulfotransferase inhibitors. A population study is asking whether dietary flavonoids might increase free oestrogen levels and slow the progression of hormone-dependent breast cancers.
Gap-junctional cell-cell communication restrains uncontrolled proliferation and is tumour-suppressive. A non-genotoxic rodent hepatocarcinogen causes connexin-32 phosphorylation and inhibits hepatocyte communication, affecting the mitogenesis/apoptosis balance (Chipman). With Minchin, aberrant control is being investigated in a connexin-32 reporter gene assay. Using polarised hepatocyte couplets, Coleman identified compounds that protect against cholestatic liver diseases, at least partly by effects on signalling mechanisms that regulate cytoskeletal function and transporters to the bile canaliculus. The recent appointment of *Schrewe will allow mechanistic links to be made between molecular signalling and toxicological responses in development and carcinogenesis.
In the Bioadhesion & Biofilm group, Callow’s work (c. M Callow) on the motile spores of the alga Enteromorpha established a comprehensive model for analysing bioadhesion during marine biofouling. Antibodies have enabled partial characterization of the glycoprotein adhesive and its secretion: genes encoding the adhesive are being cloned. Adhesive visco-elastic properties and ‘curing’ have been measured by Atomic Force Microscopy. Surface micro-topography and physical chemistry strongly influence spore settlement and development - as do surface/diffusible cues from biofilms and previously settled spores. Studies of signalling mechanisms and gene expression will yield insights into bacterial-algal communication. Leadbeater (c. M Callow) has shown how algal biofilms control nutrient fluxes between sediments and overlying water: phosphate/calcite co-precipitation in hard waters reduces the phosphate load. Applied research is assessing how biofilms contribute to ecologically sensitive water treatments (reed beds, straw bales) and influence pollutant (Cu) and nutrient (P) release from river sediments.
The Ecology, Conservation and Biodiversity group aims to understand fundamental processes in the ecology and environmental physiology of animals and plants and applies this knowledge to strategic problems. Bale investigates how the thermal ecology of insects influences species distributions and abundance. Elucidation of the unique life cycle of an Arctic aphid and its response to field-manipulated temperatures allowed a comparative analysis of climate warming effects on polar and temperate species. Similar UK studies emphasised how photoperiodically-induced diapause can override the development-promoting effects of warming. Asexual overwintering of specialised cold-tolerant morphs explains the local persistence of lettuce root aphid. He is now investigating the value of a thermal 'discriminating index’ to predict the establishment potential in the UK of alien insects (pests or biocontrol agents). Bale & Pullin are exploring the ecophysiological adaptation and phylogeography of Antarctic terrestrial arthropods along a latitudinal gradient into Antarctica. They recently detected differences in allele frequencies in several loci at different spatial scales (<3 and >500 km). Pullin‘s research combines landscape ecology and molecular genetics, providing evidence-based support for conservation actions. He established a phylogeography for the Aricia species complex and the endangered Euphydryas aurinia in northern Europe. He demonstrated substantial gene flow at the landscape scale between UK populations of Papilio machaon, suggesting that habitat quality is as important as connectivity for species persistence - as is observed with grassland butterflies in urban areas. He produced a framework to promote evidence-based practice for conservation actions.Pritchard's work combines plant physiology, biochemistry and molecular biology to understand plant function in variable environments. He has demonstrated highly heterogeneous responses to stress (drought, pollution, herbivores) at the single cell level, and - by exploiting sap-feeding insects to sample long distance transport pathways - has revealed some of the regulatory mechanisms of these compartments (e.g. role of xylem composition in salt tolerance, homeostatic nature of the sieve element). Analysis of phloem-specific gene expression is uncovering molecular mechanisms that underlie the physiological observations. With Bale, he is relating phloem diet quality (accessed by stylectomy) to insect performance (development, reproduction). Future work will build bridges between cell physiology and emerging genomic resources (e.g. in Arabidopisis, aphids and their symbiotic bacteria).
*Blackburn investigates large-scale ecological patterns and processes that influence the structures of the smaller-scale communities studied by most ecologists. Large-scale effects on local community structure have been demonstrated in communities of microarthropods living in patches of moss on rocks - and also in simulations of the structure of a woodland bird community. A study of New Zealand birds explored factors that have driven species extinctions following sequential waves of human colonisation and that have influenced the geographic ranges attained by introduced species. Future work will focus on patterns of avian introduction success, in New Zealand and elsewhere.
Copyright 2002 - HEFCE, SHEFC, ELWa, DEL
Last updated 17 October 2003