The relation between the specific absorption rate and electromagnetic field intensity for heterogeneous exposure conditions at mobile communications frequencies

The relation between the incident electromagnetic field strength and both the whole‐body and the local specific absorption rate (SAR) was investigated for typical heterogeneous exposure scenarios for frequencies relevant for mobile communication. The results were compared to results from plane wave exposure. Heterogeneous exposure arises from multiple path propagation of the electromagnetic waves to the location of interest. It is shown that plane wave exposure does not represent worst‐case exposure conditions. When the electric field strength arising at plane wave exposure is compared to the electric field strength averaged over the volume of the human body occurring during multipath exposure, 12% of all heterogeneous cases examined represent worse exposure conditions than plane wave exposure for whole‐body exposure at 946 MHz, 15% at 1840 MHz, and 22% at 2140 MHz. The deviation between plane wave and heterogeneous whole‐body SAR ranges from ?54% to 54%. For partial‐body SAR averaged over 10 g of tissue, a range from ?93% to 209% was found when comparing multiple wave exposure to single incoming plane waves. The investigations performed using the Visible Human as phantom showed that the basic restrictions are met as long as the reference levels are not exceeded. However, this must not be necessarily the case when different phantoms are used to perform similar investigations because recent studies demonstrated that reference levels might not be conservative when phantoms of children are used. Therefore, the results of this work indicate the need to extend the investigations to numerical simulations with additional human phantoms representing parts of the human population having different anatomy and morphology compared to the phantom used within the frame of this project. This also applies to phantoms of children. Bioelectromagnetics 30:651–662, 2009. © 2009 Wiley‐Liss, Inc.     

A Genome-Scale Proteomic Screen Identifies a Role for DnaK in Chaperoning of Polar Autotransporters in Shigella

Autotransporters are outer membrane proteins that are widely distributed among gram-negative bacteria. Like other autotransporters, the Shigella autotransporter IcsA, which is required for actin assembly during infection, is secreted at the bacterial pole. In the bacterial cytoplasm, IcsA localizes to poles and potential cell division sites independent of the cell division protein FtsZ. To identify bacterial proteins involved in the targeting of IcsA to the pole in the bacterial cytoplasm, we screened a genome-scale library of Escherichia coli proteins tagged with green fluorescent protein (GFP) for those that displayed a localization pattern similar to that of IcsA-GFP in cells that lack functional FtsZ using a strain carrying a temperature-sensitive ftsZ allele. For each protein that mimicked the localization of IcsA-GFP, we tested whether IcsA localization was dependent on the presence of the protein. Although these approaches did not identify a polar receptor for IcsA, the cytoplasmic chaperone DnaK both mimicked IcsA localization at elevated temperatures as a GFP fusion and was required for the localization of IcsA to the pole in the cytoplasm of E. coli. DnaK was also required for IcsA secretion at the pole in Shigella flexneri. The localization of DnaK-GFP to poles and potential cell division sites was dependent on elevated growth temperature and independent of the presence of IcsA or functional FtsZ; native DnaK was found to be enhanced at midcell and the poles. A second Shigella autotransporter, SepA, also required DnaK for secretion, consistent with a role of DnaK more generally in the chaperoning of autotransporter proteins in the bacterial cytoplasm.The Shigella outer membrane protein IcsA is unusual in that it is secreted at the bacterial old pole (9, 13, 24). The secreted protein forms a cap at the old pole (Fig. ​(Fig.1A),1A), where during the infection of host cells, it interacts with cellular actin cytoskeletal proteins to induce the formation of propulsive actin tails (6, 43, 70). Actin tail formation is essential to the spread of Shigella spp. through cell monolayers and mammalian tissues (6, 43, 47) and is critical for Shigella virulence (15, 60). IcsA is a member of the autotransporter family of secreted proteins in gram-negative bacteria. Approximately 700 autotransporter proteins are predicted to be encoded within bacterial genomes that had been annotated as of 2003 (54). All autotransporter proteins for which the site of secretion has been determined are, like IcsA, secreted at the bacterial old pole (35).Open in a separate windowFIG. 1.Design of screen for proteins that, like IcsA, localize to potential division sites independent of FtsZ. (A) Localization of IcsA on the surface of S. flexneri. Immunofluorescence using antibody to IcsA. (B) Localization of IcsA_507-620-GFP (expressed from pBAD24-icsA_507-620::gfp) to poles of single cells of E. coli MC4100 leu::Tn10 ftsZ84(Ts) grown at the permissive temperature (30°C). (C) Localization of IcsA_507-620-GFP to potential cell division sites of E. coli MC4100 leu::Tn10 ftsZ84(Ts) grown at the nonpermissive temperature (42°C). (D) Diagram of the strategy used to identify proteins of E. coli that localize to potential cell division sites independent of FtsZ, displaying a localization pattern similar to that shown for IcsA in panel C. Image from DnaK-GFP localization (expressed from leaky promoter on pCA24N-dnaK, without induction) in screen well; incomplete overlay of GFP with phase-contrast microscopy is due to the movement of cells between capturing the two images, as cells were imaged live. Size bars = 2 μm (A and B) and 5 μm (C and D). Images are representative. O/N, overnight.Several other secreted bacterial proteins are also localized to one or both cell poles; these include the Listeria monocytogenes actin assembly protein ActA (39), components of the chemotaxis apparatus in Escherichia coli and Caulobacter crescentus (1, 46, 66), the Legionella pneumophila and Agrobacterium tumifaciens type IV secretion systems (14, 40), Pseudomonas aeruginosa type IV pili (8), protein components of the cell cycle regulatory pathways in C. crescentus (reviewed in reference 72), the DNA transfer apparatus in Bacillus subtilis and Streptomyces spp. (26, 28), and polar flagella in Vibrio cholerae, Campylobacter spp., Helicobacter spp., C. crescentus, and other gram-negative bacteria. In L. monocytogenes, the polarity of ActA is established after ActA secretion and likely depends on differential growth rates of the cell wall along the length of the bacterium (56). In C. crescentus, TipN serves as a polar developmental landmark (31, 42), and RcdA provides temporal and spatial specificity in the regulated proteolysis of key factors involved in polar asymmetry (50). Beyond these studies, relatively little is known about the molecular mechanisms that mediate the proper localization of polar bacterial proteins.Chemical or genetic blockade of cell division leads to the formation of filamentous cells without septa. In cells that have been filamented by either blocking FtsI or depleting functional FtsZ, a cytoplasmic derivative of IcsA localizes at or near potential cell division sites (36), which represent the sites of future cell poles. IcsA also localizes to potential division sites independent of nucleoid occlusion (36), together indicating that the positional information directing IcsA polarity is independent of these cell division proteins and chromosome positioning. The molecules that are required for the localization of IcsA to the cell pole have not been identified.One model of IcsA localization to the pole is that freely diffusing cytoplasmic IcsA recognizes and binds a protein receptor that is present at poles and future poles. Although icsA is present only in Shigella spp., upon heterologous expression, IcsA localizes to the poles of other Enterobacteriaceae (13, 58, 59), indicating that if targeting occurs via binding to a polar receptor, the receptor is likely conserved among members of this family. In addition, since IcsA localizes independently of FtsZ and FtsI, the localization of a putative polar receptor to the pole must also be independent of these cell division proteins. To find proteins that might serve as a polar receptor for IcsA, we first conducted a genome-wide screen designed to identify the subset of E. coli proteins that localize to poles and to potential cell division sites independently of functional FtsZ. For each conserved protein that displayed this localization pattern, we then tested whether it played a role in the polar localization of IcsA. We found that, under the conditions of our screen, a green fluorescent protein (GFP) fusion to the cytoplasmic chaperone DnaK localizes to cell poles and potential cell division sites. Although DnaK is not a polar receptor for IcsA, we demonstrated that it was required for the localization of IcsA to the pole in the bacterial cytoplasm in E. coli and for the secretion of both IcsA and a second Shigella autotransporter, SepA, in native Shigella flexneri, consistent with a critical role of DnaK in the chaperoning of IcsA and SepA, and perhaps autotransporter proteins more generally, in the bacterial cytoplasm.     

Characterization of cellular uptake and distribution of coenzyme Q10 and vitamin E in PC12 cells

Coenzyme Q (CoQ) is a well-known electron transporter in the mitochondrial respiratory chain. Furthermore, ubiquinol (UQH(2))--a reduced form of ubiquinone (UQ)--has been shown to act as a radical-scavenging antioxidant. Some studies have reported the beneficial effect of CoQ addition to cultured cells; however, the cellular uptake and distribution of CoQ have not been elucidated. In the present study, we used rat pheochromocytoma PC12 cells to investigate and compare the cellular uptake and distribution of CoQ(10) and alpha-tocopherol (alphaT). UQ(10) or UQ(10)H(2) treatment resulted in an increase in the cellular content of both CoQ(10) in a time- and concentration-dependent manner. A subcellular fractionation study revealed that the added UQ(10) as well as UQ(10)H(2) mainly localized in the mitochondrial fraction, which is similar to the localization of endogenous CoQ but different from that of alphaT. The cellular distribution of alphaT directly corresponded to the lipid distribution, while the CoQ distribution did not show any relationship with the lipid distribution, particularly in the mitochondrial and microsomal fractions. These results indicate that the cellular distribution of CoQ is completely different from that of alphaT; moreover, a certain system which accumulates CoQ preferentially in mitochondria may be suggested.     

Replication Initiator DnaA of Escherichia coli Changes Its Assembly Form on the Replication Origin during the Cell Cycle

DnaA is a replication initiator protein that is conserved among bacteria. It plays a central role in the initiation of DNA replication. In order to monitor its behavior in living Escherichia coli cells, a nonessential portion of the protein was replaced by a fluorescent protein. Such a strain grew normally, and flow cytometry data suggested that the chimeric protein has no substantial loss of the initiator activity. The initiator was distributed all over the nucleoid. Furthermore, a majority of the cells exhibited certain distinct foci that emitted bright fluorescence. These foci colocalized with the replication origin (oriC) region and were brightest during the period spanning the initiation event. In cells that had undergone the initiation, the foci were enriched in less intense ones. In addition, a significant portion of the oriC regions at this cell cycle stage had no colocalized DnaA-enhanced yellow fluorescent protein (EYFP) focus point. It was difficult to distinguish the initiator titration locus (datA) from the oriC region. However, involvement of datA in the initiation control was suggested from the observation that, in ΔdatA cells, DnaA-EYFP maximally colocalized with the oriC region earlier in the cell cycle than it did in wild-type cells and oriC concentration was increased.Initiation of DNA replication is highly regulated to coordinate with cell proliferation. It begins with a series of events in which the replication machinery is assembled at the replication origin of the chromosomal DNA (15, 26, 28, 38). Central to this process are the initiator proteins that bind to the origin of replication and eventually lead to the unwinding of the origin and to helicase loading on the unwound region. Previous biochemical studies and recent structural studies of the bacterial initiator protein DnaA have proposed the molecular mechanism of the action of ATP-DnaA in forming a large oligomeric complex to remodel the unique origin, oriC, and trigger duplex melting (12, 26). However, it is still not clear how the timing of initiation is controlled so that it takes place at a fixed time in the cell cycle. It has been reported that a basal level of DnaA molecules is bound by high-affinity DnaA binding sites (DnaA boxes R1, R2, and R4) at oriC throughout the cell cycle (9, 37). It is also suggested that noncanonical ATP-DnaA binding sites within oriC are occupied at elevated levels of the initiator molecules prior to the initiation event (18, 25). Thus, regulation of the activity and availability of DnaA is an important factor for the initiation control.At least three schemes are known to prevent untimely initiations in Escherichia coli. First, oriC is subject to sequestration, a process that prevents reinitiation, possibly by blocking ATP-DnaA from binding to newly replicated oriC (8, 24). E. coli oriC contains 11 GATC sites that are normally methylated on both strands by Dam methyltransferase. Immediately after passage of the replication fork, GATC sites are in a hemimethylated state, with the newly synthesized strands remaining unmethylated. SeqA binds specifically to such sites and, at oriC, protects these regions from reinitiation for about one-third of the cell cycle (6, 39). Second, in a process termed regulatory inactivation of DNA (RIDA), ATP-DnaA molecules are converted to an inactive ADP-bound form after initiation by the combined action of a β subunit of DNA polymerase III holoenzyme and Hda (16, 17). Newly synthesized DnaA molecules are able to bind ATP for the next initiation event, since its cellular concentration is much higher than that of ADP. ATP-DnaA is also regenerated from the inactive ADP-DnaA later in the cell cycle (21). Finally, the chromosomal segment datA serves to reduce the level of free DnaA protein by titrating a large number of DnaA molecules after replication of the site close to oriC (20).Cytological studies would be very useful for developing our understanding of the regulation mechanisms associated with the initiation step. In the present study, we tagged E. coli DnaA with a fluorescent protein in order to monitor its behavior in live cells. Microscopic observation revealed that DnaA is distributed all over the nucleoid. Remarkably, the majority of cells bore distinct foci that emitted brighter fluorescence against a weak fluorescent background on the nucleoid. We analyzed the behavior of these foci during the cell cycle with respect to oriC and datA.     

BMP and Hh signaling affects primordial germ cell division in Drosophila

The germline is segregated from the remainder of the soma during early embryonic development in metazoan species. In Drosophila, female primordial germ cells (PGCs) continue to proliferate during larval development, and become germline stem cells at the early pupal stage. To elucidate the roles of growth factors in larval PGC division, we examined expression patterns of a bone morphogenetic protein (BMP) growth factor, Decapentaplegic (Dpp), and Hedgehog (Hh), along with factors downstream of each, in the ovary during larval development. Dpp signaling appeared in the ovarian soma from early larval development, and was prominent in the terminal filament cells at late larval stage, whereas Hh appeared in the ovarian soma and PGCs from the third instar larval stage. The number of PGCs decreased when components of these signal transduction pathways were abrogated by RNAi in the PGCs, indicating that both Dpp and Hh signals directly regulate PGC proliferation. Experiments on the up- and down-regulation of Dpp and Hh with a tissue-specific Gal4 driver indicated that Dpp and Hh act as extrinsic and autocrine growth factors. Furthermore, heat-pulse experiments with hs-Gal4 showed that Dpp is active in PGC proliferation throughout larval development, whereas Hh has effects only during late larval development. In addition to Dpp, the reduction of Glass bottom boat (Gbb), another BMP molecule, caused a decrease in the number of PGCs and initiation of larval PGCs differentiation into cystocytes, indicating that Gbb functions to promote PGC division and repress differentiation.     

The DNA Damage Checkpoint Regulates a Transition between Yeast and Hyphal Growth in Schizosaccharomyces japonicus

Dimorphic yeasts change between unicellular growth and filamentous growth. Many dimorphic yeasts species are pathogenic for humans and plants, being infectious as invasive hypha. We have studied the determinants of the dimorphic switch in the nonpathogenic fission yeast Schizosaccharomyces japonicus, which is evolutionarily close to the well-characterized fission yeast S. pombe. We report that camptothecin, an inhibitor of topoisomerase I, reversibly induced the unicellular to hyphal transition in S. japonicus at low concentrations of camptothecin that did not induce checkpoint arrest and the transition required the DNA checkpoint kinase Chk1. Furthermore, a mutation of chk1 induced hyphal transition without camptothecin. Thus, we identify a second function for Chk1 distinct from its role in checkpoint arrest. Activation of the switch from single cell bipolar growth to monopolar filamentous growth may assist cells to evade the source of DNA damage.Yeasts and molds are major members of the kingdom Fungi. Molds grow as multicellular filamentous hyphae. On the other hand, yeasts propagate in a unicellular fashion by budding or by binary fission. However, many types of yeast can switch their growth modes, changing from unicellular growth to filamentous branching multicellular hyphae. This hyphal transition can be induced by a wide variety of environmental changes ranging from pH to the nature of the carbon source, and many species of dimorphic yeasts that are pathogenic for humans and plants are infectious in the hyphal form (15, 20).Hyphal transition is a simple mode of cellular differentiation program that is turned on upon environmental changes. The fungi may differentiate to adapt to the environmental challenges. Especially in the case of Candida albicans strains that infect humans, the hyphal transition may function as an action to resist against attack from macrophages or neutrophils. Hyphae are more difficult to phagocytose (16). It can also eventually kill macrophages if hyphal transition is triggered after ingestion by macrophage (14). Indeed, C. albicans cells that cannot form hyphae are avirulent. However, inducing hyphal growth in pathogenic yeasts is not always readily achievable in the laboratory, and genetic analysis of the hyphal growth phase and transition to this phase is often limited by the lack of appropriate tools. Thus, genetically tractable nonpathogenic dimorphic yeasts are attractive models for investigating invasive hypha.The nonpathogenic fission yeast Schizosaccharomyces japonicus is evolutionarily close to the well-characterized fission yeast Schizosaccharomyces pombe (5, 24). S. japonicus is dimorphic, transiting between unicellular and hyphal growth, and thus offers itself as an appropriate model to study this differentiation mechanism and the requirements of hyphal growth (25). In S. japonicus, hyphal growth occurs naturally on most solid medium and can occur over a range of nutrient conditions (26). It has been proposed that a gradient of nitrogen in the substrate is necessary to both initiate and direct hyphal growth in S. japonicus (26). In this report we establish conditions to induce hyphal growth in a microchamber in liquid media. In addition, we show that a low dose of the topoisomerase inhibitor camptothecin (CPT) induces hyphal differentiation under rich nutrient conditions and identify a role for the DNA damage checkpoint response in promoting the CPT-dependent transition from unicellular to hyphal growth. Genetic analysis demonstrates that this role of the checkpoint is distinct from checkpoint arrest, and we suggest it may provide an opportunity for S. japonicus to grow away from sources of genotoxic stress.     

Big,small or mezzo?

Lessons from science studies for the ongoing debate about ‘big'' versus ‘little'' research projectsDuring the past six decades, the importance of scientific research to the developed world and the daily lives of its citizens has led many industrialized countries to rebrand themselves as ‘knowledge-based economies''. The increasing role of science as a main driver of innovation and economic growth has also changed the nature of research itself. Starting with the physical sciences, recent decades have seen academic research increasingly conducted in the form of large, expensive and collaborative ‘big science'' projects that often involve multidisciplinary, multinational teams of scientists, engineers and other experts.Although laboratory biology was late to join the big science trend, there has nevertheless been a remarkable increase in the number, scope and complexity of research collaborations…Although laboratory biology was late to join the big science trend, there has nevertheless been a remarkable increase in the number, scope and complexity of research collaborations and projects involving biologists over the past two decades (Parker et al, 2010). The Human Genome Project (HGP) is arguably the most well known of these and attracted serious scientific, public and government attention to ‘big biology''. Initial exchanges were polarized and often polemic, as proponents of the HGP applauded the advent of big biology and argued that it would produce results unattainable through other means (Hood, 1990). Critics highlighted the negative consequences of massive-scale research, including the industrialization, bureaucratization and politicization of research (Rechsteiner, 1990). They also suggested that it was not suited to generating knowledge at all; Nobel laureate Sydney Brenner joked that sequencing was so boring it should be done by prisoners: “the more heinous the crime, the bigger the chromosome they would have to decipher” (Roberts, 2001).A recent Opinion in EMBO reports summarized the arguments against “the creeping hegemony” of ‘big science'' over ‘little science'' in biomedical research. First, many large research projects are of questionable scientific and practical value. Second, big science transfers the control of research topics and goals to bureaucrats, when decisions about research should be primarily driven by the scientific community (Petsko, 2009). Gregory Petsko makes a valid point in his Opinion about wasteful research projects and raises the important question of how research goals should be set and by whom. Here, we contextualize Petsko''s arguments by drawing on the history and sociology of science to expound the drawbacks and benefits of big science. We then advance an alternative to the current antipodes of ‘big'' and ‘little'' biology, which offers some of the benefits and avoids some of the adverse consequences.Big science is not a recent development. Among the first large, collaborative research projects were the Manhattan Project to develop the atomic bomb, and efforts to decipher German codes during the Second World War. The concept itself was put forward in 1961 by physicist Alvin Weinberg, and further developed by historian of science Derek De Solla Price in his pioneering book, Little Science, Big Science. “The large-scale character of modern science, new and shining and all powerful, is so apparent that the happy term ‘Big Science'' has been coined to describe it” (De Solla Price, 1963). Weinberg noted that science had become ‘big'' in two ways. First, through the development of elaborate research instrumentation, the use of which requires large research teams, and second, through the explosive growth of scientific research in general. More recently, big science has come to refer to a diverse but strongly related set of changes in the organization of scientific research. This includes expensive equipment and large research teams, but also the increasing industrialization of research activities, the escalating frequency of interdisciplinary and international collaborations, and the increasing manpower needed to achieve research goals (Galison & Hevly, 1992). Many areas of biological research have shifted in these directions in recent years and have radically altered the methods by which biologists generate scientific knowledge.Despite this long history of collaboration, laboratory biology remained ‘small-scale'' until the rising prominence of molecular biology changed the research landscapeUnderstanding the implications of this change begins with an appreciation of the history of collaborations in the life sciences—biology has long been a collaborative effort. Natural scientists accompanied the great explorers in the grand alliance between science and exploration during the sixteenth and seventeenth centuries (Capshew & Rader, 1992), which not only served to map uncharted territories, but also contributed enormously to knowledge of the fauna and flora discovered. These early expeditions gradually evolved into coordinated, multidisciplinary research programmes, which began with the International Polar Years, intended to concentrate international research efforts at the North and South Poles (1882–1883; 1932–1933). The Polar Years became exemplars of large-scale life science collaboration, begetting the International Geophysical Year (1957–1958) and the International Biological Programme (1968–1974).For Weinberg, the potentially negative consequences associated with big science were “adminstratitis, moneyitis, and journalitis”…Despite this long history of collaboration, laboratory biology remained ‘small-scale'' until the rising prominence of molecular biology changed the research landscape. During the late 1950s and early 1960s, many research organizations encouraged international collaboration in the life sciences, spurring the creation of, among other things, the European Molecular Biology Organization (1964) and the European Molecular Biology Laboratory (1974). In addition, international mapping and sequencing projects were developed around model organisms such as Drosophila and Caenorhabditis elegans, and scientists formed research networks, exchanged research materials and information, and divided labour across laboratories. These new ways of working set the stage for the HGP, which is widely acknowledged as the cornerstone of the current ‘post-genomics era''. As an editorial on ‘post-genomics cultures'' put it in the journal Nature, “Like it or not, big biology is here to stay” (Anon, 2001).Just as big science is not new, neither are concerns about its consequences. As early as 1948, the sociologist Max Weber worried that as equipment was becoming more expensive, scientists were losing autonomy and becoming more dependent on external funding (Weber, 1948). Similarly, although Weinberg and De Solla Price expressed wonder at the scope of the changes they were witnessing, they too offered critical evaluations. For Weinberg, the potentially negative consequences associated with big science were “adminstratitis, moneyitis, and journalitis”; meaning the dominance of science administrators over practitioners, the tendency to view funding increases as a panacea for solving scientific problems, and progressively blurry lines between scientific and popular writing in order to woo public support for big research projects (Weinberg, 1961). De Solla Price worried that the bureaucracy associated with big science would fail to entice the intellectual mavericks on which science depends (De Solla Price, 1963). These concerns remain valid and have been voiced time and again.As big science represents a major investment of time, money and manpower, it tends to determine and channel research in particular directions that afford certain possibilities and preclude others (Cook & Brown, 1999). In the worst case, this can result in entire scientific communities following false leads, as was the case in the 1940s and 1950s for Soviet agronomy. Huge investments were made to demonstrate the superiority of Lamarckian over Mendelian theories of heritability, which held back Russian biology for decades (Soyfer, 1994). Such worst-case scenarios are, however, rare. A more likely consequence is that big science can diminish the diversity of research approaches. For instance, plasma fusion scientists are now under pressure to design projects that are relevant to the large-scale International Thermonuclear Experimental Reactor, despite the potential benefits of a wide array of smaller-scale machines and approaches (Hackett et al, 2004). Big science projects can also involve coordination challenges, take substantial time to realize success, and be difficult to evaluate (Neal et al, 2008).Importantly, big science projects allow for the coordination and activation of diverse forms of expertise across disciplinary, national and professional boundariesAnother danger of big science is that researchers will lose the intrinsic satisfaction that arises from having personal control over their work. Dissatisfaction could lower research productivity (Babu & Singh, 1998) and might create the concomitant danger of losing talented young researchers to other, more engaging callings. Moreover, the alienation of scientists from their work as a result of big science enterprises can lead to a loss of personal responsibility for research. In turn, this can increase the likelihood of misconduct, as effective social control is eroded and “the satisfactions of science are overshadowed by organizational demands, economic calculations, and career strategies” (Hackett, 1994).Practicing scientists are aware of these risks. Yet, they remain engaged in large-scale projects because they must, but also because of the real benefits these projects offer. Importantly, big science projects allow for the coordination and activation of diverse forms of expertise across disciplinary, national and professional boundaries to solve otherwise intractable basic and applied problems. Although calling for international and interdisciplinary collaboration is popular, practicing it is notably less popular and much harder (Weingart, 2000). Big science projects can act as a focal point that allows researchers from diverse backgrounds to cooperate, and simultaneously advances different scientific specialties while forging interstitial connections among them. Another major benefit of big science is that it facilitates the development of common research standards and metrics, allowing for the rapid development of nascent research frontiers (Fujimura, 1996). Furthermore, the high profile of big science efforts such as the HGP and CERN draw public attention to science, potentially enhancing scientific literacy and the public''s willingness to support research.Rather than arguing for or against big science, molecular biology would best benefit from strategic investments in a diverse portfolio of big, little and ‘mezzo'' research projectsBig science can also ease some of the problems associated with scientific management. In terms of training, graduate students and junior researchers involved in big science projects can gain additional skills in problem-solving, communication and team working (Court & Morris, 1994). The bureaucratic structure and well-defined roles of big science projects also make leadership transitions and researcher attrition easier to manage compared with the informal, refractory organization of most small research projects. Big science projects also provide a visible platform for resource acquisition and the recruitment of new scientific talent. Moreover, through their sheer size, diversity and complexity, they can also increase the frequency of serendipitous social interactions and scientific discoveries (Hackett et al, 2008). Finally, large-scale research projects can influence scientific and public policy. Big science creates organizational structures in which many scientists share responsibility for, and expectations of, a scientific problem (Van Lente, 1993). This shared ownership and these shared futures help coordinate communication and enable researchers to present a united front when advancing the potential benefits of their projects to funding bodies.Given these benefits and pitfalls of big science, how might molecular biology best proceed? Petsko''s response is that, “[s]cientific priorities must, for the most part, be set by the free exchange of ideas in the scientific literature, at meetings and in review panels. They must be set from the bottom up, from the community of scientists, not by the people who control the purse strings.” It is certainly the case, as Petsko also acknowledges, that science has benefited from a combination of generous public support and professional autonomy. However, we are less sanguine about his belief that the scientific community alone has the capacity to ascertain the practical value of particular lines of inquiry, determine the most appropriate scale of research, and bring them to fruition. In fact, current mismatches between the production of scientific knowledge and the information needs of public policy-makers strongly suggest that the opposite is true (Sarewitz & Pielke, 2007).Instead, we maintain that these types of decision should be determined through collective decision-making that involves researchers, governmental funding agencies, science policy experts and the public. In fact, the highly successful HGP involved such collaborations (Lambright, 2002). Taking into account the opinions and attitudes of these stakeholders better links knowledge production to the public good (Cash et al, 2003)—a major justification for supporting big biology. We do agree with Petsko, however, that large-scale projects can develop pathological characteristics, and that all programmes should therefore undergo regular assessments to determine their continuing worth.Rather than arguing for or against big science, molecular biology would best benefit from strategic investments in a diverse portfolio of big, little and ‘mezzo'' research projects. Their size, duration and organizational structure should be determined by the research question, subject matter and intended goals (Westfall, 2003). Parties involved in making these decisions should, in turn, aim at striking a profitable balance between differently sized research projects to garner the benefits of each and allow practitioners the autonomy to choose among them.This will require new, innovative methods for supporting and coordinating research. An important first step is ensuring that funding is made available for all kinds of research at a range of scales. For this to happen, the current funding model needs to be modified. The practice of allocating separate funds for individual investigator-driven and collective research projects is a positive step in the right direction, but it does not discriminate between projects of different sizes at a sufficiently fine resolution. Instead, multiple funding pools should be made available for projects of different sizes and scales, allowing for greater accuracy in project planning, funding and evaluation.It is up to scientists and policymakers to discern how to benefit from the advantages that ‘bigness'' has to offer, while avoiding the pitfalls inherent in doing soSecond, science policy should consciously facilitate the ‘scaling up'', ‘scaling down'' and concatenation of research projects when needed. For instance, special funds might be established for supporting small-scale but potentially transformative research with the capacity to be scaled up in the future. Alternatively, small-scale satellite research projects that are more nimble, exploratory and risky, could complement big science initiatives or be generated by them. This is also in line with Petsko''s statement that “the best kind of big science is the kind that supports and generates lots of good little science.” Another potentially fruitful strategy we suggest would be to fund independent, small-scale research projects to work on co-relevant research with the later objective of consolidating them into a single project in a kind of building-block assembly. By using these and other mechanisms for organizing research at different scales, it could help to ameliorate some of the problems associated with big science, while also accruing its most important benefits.Within the life sciences, the field of ecology perhaps best exemplifies this strategy. Although it encompasses many small-scale laboratory and field studies, ecologists now collaborate in a variety of novel organizations that blend elements of big, little and mezzo science and that are designed to catalyse different forms of research. For example, the US National Center for Ecological Analysis and Synthesis brings together researchers and data from many smaller projects to synthesize their findings. The Long Term Ecological Research Network consists of dozens of mezzo-scale collaborations focused on specific sites, but also leverages big science through cross-site collaborations. While investments are made in classical big science projects, such as the National Ecological Observatory Network, no one project or approach has dominated—nor should it. In these ways, ecologists have been able to reap the benefits of big science whilst maintaining diverse research approaches and individual autonomy and still being able to enjoy the intrinsic satisfaction associated with scientific work.Big biology is here to stay and is neither a curse nor a blessing. It is up to scientists and policy-makers to discern how to benefit from the advantages that ‘bigness'' has to offer, while avoiding the pitfalls inherent in so doing. The challenge confronting molecular biology in the coming years is to decide which kind of research projects are best suited to getting the job done. Molecular biology itself arose, in part, from the migration of physicists to biology; as physics research projects and collaborations grew and became more dependent on expensive equipment, appreciating the saliency of one''s own work became increasingly difficult, which led some to seek refuge in the comparatively little science of biology (Dev, 1990). The current situation, which Petsko criticizes in his Opinion article, is thus the result of an organizational and intellectual cycle that began more than six decades ago. It would certainly behoove molecular biologists to heed his warnings and consider the best paths forward.? Open in a separate windowNiki VermeulenOpen in a separate windowJohn N. ParkerOpen in a separate windowBart Penders     

Functional divergence within class B MADS-box genes TfGLO and TfDEF in Torenia fournieri Lind

Homeotic class B genes GLOBOSA (GLO)/PISTILLATA (PI) and DEFICIENS (DEF)/APETALA3 (AP3) are involved in the development of petals and stamens in Arabidopsis. However, functions of these genes in the development of floral organs in torenia are less well known. Here, we demonstrate the unique floral phenotypes of transgenic torenia formed due to the modification of class B genes, TfGLO and TfDEF. TfGLO-overexpressing plants showed purple-stained sepals that accumulated anthocyanins in a manner similar to that of petals. TfGLO-suppressed plants showed serrated petals and TfDEF-suppressed plants showed partially decolorized petals. In TfGLO-overexpressing plants, cell shapes on the surfaces of sepals were altered to petal-like cell shapes. Furthermore, TfGLO- and TfDEF-suppressed plants partially had sepal-like cells on the surfaces of their petals. We isolated putative class B gene-regulated genes and examined their expression in transgenic plants. Three xyloglucan endo-1,4-beta-d-glucanase genes were up-regulated in TfGLO- and TfDEF-overexpressing plants and down-regulated in TfGLO- and TfDEF-suppressed plants. In addition, 10 anthocyanin biosynthesis-related genes, including anthocyanin synthase and chalcone isomerase, were up-regulated in TfGLO-overexpressing plants and down-regulated in TfGLO-suppressed plants. The expression patterns of these 10 genes in TfDEF transgenic plants were diverse and classified into several groups. HPLC analysis indicated that sepals of TfGLO-overexpressing plants accumulate the same type of anthocyanins and flavones as wild-type plants. The difference in phenotypes and expression patterns of the 10 anthocyanin biosynthesis-related genes between TfGLO and TfDEF transgenic plants indicated that TfGLO and TfDEF have partial functional divergence, while they basically work synergistically in torenia.