FGF ligands in Drosophila have distinct activities required to support cell migration and differentiation

Fibroblast growth factor (FGF) signaling controls a vast array of biological processes including cell differentiation and migration, wound healing and malignancy. In vertebrates, FGF signaling is complex, with over 100 predicted FGF ligand-receptor combinations. Drosophila melanogaster presents a simpler model system in which to study FGF signaling, with only three ligands and two FGF receptors (FGFRs) identified. Here we analyze the specificity of FGFR [Heartless (Htl) and Breathless (Btl)] activation by each of the FGF ligands [Pyramus (Pyr), Thisbe (Ths) and Branchless (Bnl)] in Drosophila. We confirm that both Pyr and Ths can activate Htl, and that only Bnl can activate Btl. To examine the role of each ligand in supporting activation of the Htl FGFR, we utilize genetic approaches that focus on the earliest stages of embryonic development. When pyr and ths are equivalently expressed using the Gal4 system, these ligands support qualitatively different FGFR signaling responses. Both Pyr and Ths function in a non-autonomous fashion to support mesoderm spreading during gastrulation, but Pyr exhibits a longer functional range. pyr and ths single mutants exhibit defects in mesoderm spreading during gastrulation, yet only pyr mutants exhibit severe defects in dorsal mesoderm specification. We demonstrate that the Drosophila FGFs have different activities and that cell migration and differentiation have different ligand requirements. Furthermore, these FGF ligands are not regulated solely by differential expression, but the sequences of these linked genes have evolved to serve different functions. We contend that inherent properties of FGF ligands make them suitable to support specific FGF-dependent processes, and that FGF ligands are not always interchangeable

Quantitative imaging of the collective cell movements shaping an embryo

The recent development of imaging and image processing techniques, such as 4D microscopy and 3D cell tracking, enables analysis through quantification of the movement of large cell populations in vivo. These imaging approaches provide an opportunity to study embryonic morphogenesis during development from the level of cellular processes to the scale of entire organism. Image analysis reveals cell collective behaviors that shape an embryo and offers some surprising insights into the cell-cell interactions involved in concerted movements. We illustrate the power of this approach by studying the early development of Drosophila embryos

Dynamic Analyses of Drosophila Gastrulation Provide Insights into Collective Cell Migration

The concerted movement of cells from different germ layers contributes to morphogenesis during early embryonic development. Using an optimized imaging approach and quantitative methods, we analyzed the trajectories of hundreds of ectodermal cells and internalized mesodermal cells within Drosophila embryos over 2 hours during gastrulation. We found a high level of cellular organization, with mesoderm cell movements correlating with some but not all ectoderm movements. During migration, the mesoderm population underwent two ordered waves of cell division and synchronous cell intercalation, and cells at the leading edge stably maintained position. Fibroblast growth factor (FGF) signaling guides mesodermal cell migration; however, we found some directed dorsal migration in an FGF receptor mutant, which suggests that additional signals are involved. Thus, decomposing complex cellular movements can provide detailed insights into collective cell migration

Quantitative imaging of collective cell migration during Drosophila gastrulation: multiphoton microscopy and computational analysis

This protocol describes imaging and computational tools to collect and analyze live imaging data of embryonic cell migration. Our five-step protocol requires a few weeks to move through embryo preparation and four-dimensional (4D) live imaging using multiphoton microscopy, to 3D cell tracking using image processing, registration of tracking data and their quantitative analysis using computational tools. It uses commercially available equipment and requires expertise in microscopy and programming that is appropriate for a biology laboratory. Custom-made scripts are provided, as well as sample datasets to permit readers without experimental data to carry out the analysis. The protocol has offered new insights into the genetic control of cell migration during Drosophila gastrulation. With simple modifications, this systematic analysis could be applied to any developing system to define cell positions in accordance with the body plan, to decompose complex 3D movements and to quantify the collective nature of cell migration

Novel Approaches to Investigate One-Carbon Metabolism and Related B-Vitamins in Blood Pressure.

Hypertension, a major risk factor for heart disease and stroke, is the world's leading cause of preventable, premature death. A common polymorphism (677C→T) in the gene encoding the folate metabolizing enzyme methylenetetrahydrofolate reductase (MTHFR) is associated with increased blood pressure, and there is accumulating evidence demonstrating that this phenotype can be modulated, specifically in individuals with the MTHFR 677TT genotype, by the B-vitamin riboflavin, an essential co-factor for MTHFR. The underlying mechanism that links this polymorphism, and the related gene-nutrient interaction, with hypertension is currently unknown. Previous research has shown that 5-methyltetrahydrofolate, the product of the reaction catalysed by MTHFR, appears to be a positive allosteric modulator of endothelial nitric oxide synthase (eNOS) and may thus increase the production of nitric oxide, a potent vasodilator. Blood pressure follows a circadian pattern, peaking shortly after wakening and falling during the night, a phenomenon known as 'dipping'. Any deviation from this pattern, which can only be identified using ambulatory blood pressure monitoring (ABPM), has been associated with increased cardiovascular disease (CVD) risk. This review will consider the evidence linking this polymorphism and novel gene-nutrient interaction with hypertension and the potential mechanisms that might be involved. The role of ABPM in B-vitamin research and in nutrition research generally will also be reviewed.The PhD studentship of A.M. was funded by the Northern Ireland Department for Employment and Learning. DSM Nutritional Products Ltd. partly supported project costs associated with this work. The funders had no role in the design, analysis or writing of this paper

A Comparison of Male and Female Title I ADA Discrimination Allegations in Relation to Employer Characteristics

This investigation determined if there is a difference in the proportion of Title I workplace discrimination allegations filed by females in comparison to males under the Americans with Disabilities Act (ADA) in relation to the employer characteristics. Findings are reported and implications for future research and vocational rehabilitation practice

Levels of autotrophy and heterotrophy in mesophotic corals near the end photic zone

Mesophotic corals live at ~30-150 m depth and can sustain metabolic processes under light-limited conditions by enhancing autotrophy through specialized photoadaptations or increasing heterotrophic nutrient acquisition. These acclimatory processes are often species-specific, however mesophotic ecosystems are largely unexplored and acclimation limits for most species are unknown. This study examined mesophotic coral ecosystems using a remotely operated vehicle (Ashmore Reef, Western Australia at 40 – 75m depth) to investigate the trophic ecology of five species of scleractinian coral (from genera Leptoseris, Pachyseris, and Craterastrea) using stable isotope analyses (δ13C and δ15N) of host and symbiont tissues and protein concentration. Trophic strategies were analyzed between species and between overall corals sampled above and below the end-photic point, where light is only 1% of surface irradiance. Results showed species-specific differences in resource use. Leptoseris hawaiiensis, L. scabra, and P. speciosa had similar Δ13C values (δ13C host - δ13C symbiont) approaching zero ( \u3c 0.5 ‰) which indicated greater dependence on symbiont autotrophy. In contrast, Leptoseris glabra and Craterastrea levis had higher Δ13C values (1.4 to 3.5 ‰) which indicated a greater reliance on external carbon sources. The latter two species also demonstrated tight nitrogen recycling within the holobiont, exhibiting low Δ15N values (host δ15N - symbiont δ15N = \u3c 0.5 ‰), compared to more autotrophic species (Δ15N = \u3e 1.2 ‰). Some species demonstrated the ability to maintain metabolic processes despite substantially reduced light availability (0.5 – 2% of surface irradiance). This research challenges our knowledge of acclimation limits for many scleractinian corals and contributes novel information for Ashmore Reef, the Western Australia region and mesophotic ecosystems in general, and critically examines common methods used to interpretate trophic ecology with bulk stable isotopes δ13C and δ15N