<Emphasis Type="Italic">Drosophila</Emphasis> central brain formation requires Robo proteins

The Robo proteins have been extensively studied in the Drosophila embryonic ventral nerve cord, in which their expression level controls the midline crossing and optic lobe formation, but nothing is known about their activities during adult central brain formation. We have analyzed how Robo guidance cues influence central complex (CX) and mushroom body (MB) formation. Mutations of robo2 and robo3 confer a series of strong MB and CX defects. We found that the Robo2 and Robo3 proteins are expressed in two structures of the developing CX, the fan-shaped body (FB) and the noduli (NO), and by fibers across the central neuropile. We conclude that the Robo2 and Robo3 receptors play postembryonic roles during central brain formation.     

Morphological characterization of single fan-shaped body neurons in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The fan-shaped body is the largest substructure of the central complex in Drosophila melanogaster. Two groups of large-field neurons that innervate the fan-shaped body, viz., F1 and F5 neurons, have recently been found to be involved in visual pattern memory for “contour orientation” and “elevation” in a rut-dependent manner. The F5 neurons have been found to be responsible for the parameter “elevation” in a for-dependent manner. We have shown here that the F1 neuron also affects visual memory for “contour orientation” in a for-dependent way. With the help of Gal4/UAS and FLP-out techniques, we have characterized the morphological features of these two groups of neurons at single neuron resolution. We have observed that F1 or F5 neurons are groups of isomorphic individual neurons. Single F1 neurons have three main arborization regions: one in the first layer of the fan-shaped body, one in the ventral body, and another in the inferior medial protocerebrum. Single F5 neurons have two arborization regions: one in the fifth layer of the fan-shaped body and the other in the superior medial protocerebrum. The polarity of the F1 and F5 neurons has been studied with the Syt-GFP marker. Our results indicate the existence of presynaptic sites of both F1 and F5 neurons located in the fan-shaped body and postsynaptic sites outside of the fan-shaped body. This work was supported by the “973 Program” (2005CB522804 and 2009CB918702), the National Natural Sciences Foundation of China (30621004, 30625022, and 30770682), and the Knowledge Innovation Program of the Chinese Academy of Sciences (KSCX2-YW-R-28).     

Characterization of the PMT Gene Family in Cryptococcus neoformans

Background

Protein-O-mannosyltransferases (Pmt''s) catalyze the initial step of protein-O-glycosylation, the addition of mannose residues to serine or threonine residues of target proteins.

Methodology/Principal Findings

Based on protein similarities, this highly conserved protein family can be divided into three subfamilies: the Pmt1 sub-family, the Pmt2 sub-family and the Pmt4 sub-family. In contrast to Saccharomyces cerevisiae and Candida albicans, but similar to filamentous fungi, three putative PMT genes (PMT1, PMT2, and PMT4) were identified in the genome of the human fungal pathogen Cryptococcus neoformans. Similar to Schizosaccharomyces pombe and C. albicans, C. neoformans PMT2 is an essential gene. In contrast, the pmt1 and pmt4 single mutants are viable; however, the pmt1/pmt4 deletions are synthetically lethal. Mutation of PMT1 and PMT4 resulted in distinct defects in cell morphology and cell integrity. The pmt1 mutant was more susceptible to SDS medium than wild-type strains and the mutant cells were enlarged. The pmt4 mutant grew poorly on high salt medium and demonstrated abnormal septum formation and defects in cell separation. Interestingly, the pmt1 and pmt4 mutants demonstrated variety-specific differences in the levels of susceptibility to osmotic and cell wall stress. Delayed melanin production in the pmt4 mutant was the only alteration of classical virulence-associated phenotypes. However, the pmt1 and pmt4 mutants showed attenuated virulence in a murine inhalation model of cryptococcosis.

Conclusion/Significance

These findings suggest that C. neoformans protein-O-mannosyltransferases play a crucial role in maintaining cell morphology, and that reduced protein-O-glycosylation leads to alterations in stress resistance, cell wall composition, cell integrity, and survival within the host.     

Mortal: A Mutant of White Clover Defective in Nodal Root Development

A monogenic dominant mutant of white clover (Trifolium repens L.), designated Mortal, which is defective in the formation of adventitious nodal roots, is described. Mortal plants grown at temperatures ranging from 10 to 25°C do not initiate nodal root primordium development. However, all other aspects of plant development are normal, including the formation of lateral roots and wound-induced adventitious roots. In some genetic backgrounds, the Mortal mutation has a temperature-sensitive conditional phenotype. Mortal plants shifted from growing conditions of 20 to 30°C for 2 to 3 d form nodal root meristems. However, new nodes that develop after plants are returned to 20°C exhibit the mutant phenotype. The capacity to form nodal roots on cuttings placed in water is also influenced by the genetic background of the Mortal mutation. Genetic analysis established that the physiological reversion of Mortal to nodal root formation is controlled by at least two separate dominant genetic loci, one for Nodal water response (Now) and one for Nodal temperature response (Not); the Now locus has a dominant epistatic interaction with the Not locus. The conditional nature of Mortal should provide opportunities for the identification of genetic and physiological mechanisms that influence the development of nodal roots.Whereas the basic structure of angiosperms is established during embryogenesis, most organs are formed by postembryonic development (Esau, 1977). Generally, all of the shoot structures (leaves, nodes, internodes, axillary shoot meristems, and flowers) are derived from the primary shoot apical meristem. However, adventitious shoot-borne roots are an exception because they develop endogenously from differentiated parenchyma cells close to the vascular tissues (Lovell and White, 1986).Little is known about the genes that control adventitious shoot-borne root morphogenesis, despite their importance for anchorage, nutrient acquisition, and water uptake from the soil in a wide range of plant species. One approach to understanding the genetic mechanisms that underlie adventitious root initiation and development is to identify and characterize mutants altered in the process. At present, few mutants with defects in adventitious root development are known (Schiefelbein and Benfey, 1991). There are mutants of tomato that produce few or no adventitious roots (Butler, 1954; Zobel, 1991) and mutants of maize that are defective in the formation of lateral seminal roots, crown roots, or both lateral seminal and crown roots (Jenkins, 1930; de Miranda, 1980; Hetz et al., 1996).The general unpredictability in the formation of secondary roots on shoots complicates the analysis of the genetic and molecular mechanisms controlling adventitious root development. This can be minimized by characterizing the genetic control of the formation of adventitious nodal root primordia. In some plant species, adventitious root primordia arise in a precise and ordered manner during node development. One such example is the nodal roots that form on the prostrate stolons of white clover (Trifolium repens L.; Thomas, 1987). In T. repens, the nodes of each stolon alternate in orientation so that successive nodes produce leaves and axillary buds on opposite sides of the stolon (Erith, 1924).The organization of nodes and nodal roots in white clover is illustrated in Figure ​Figure1.1. Root primordia are typically absent from the first four nodes of wild-type white clover stolons, and nodes bearing the first five leaf primordia are enclosed within the leaf sheath of the first visible node (Erith, 1924; Thomas, 1987). The first nodal root primordium is initiated below the axillary shoot bud of the fifth node, and a second primordium forms above the axillary shoot of the sixth node. In the seventh node, the lowermost of each pair of nodal root primordia matures into a root apical meristem that grows out through both the stolon epidermis and the stipular sheath to form a visible root, whereas development of the uppermost nodal root meristem is normally arrested such that it remains within the stipule. Further growth of the uppermost nodal root meristem usually occurs only in very moist conditions. Figure 1Stolon morphology of a wild-type white clover plant. A, Underside view of the apical portion of a stolon showing the nodes (N), leaf stipule (S), and petiole (P). B, Schematic representation of nodal development. SAM, Shoot apical meristem; LRP, lower ...White clover (2n = 4× = 32) is predominantly an obligate outcrossing species with disomic inheritance. Therefore, populations are a heterogeneous mixture of highly heterozygous individuals. This heterogeneity and the associated plasticity in environmental response complicates genetic analysis of some developmental traits in white clover. However, there are dominant self-compatible alleles of the gametophytic S locus system of sexual incompatibility, which can be used to self plants for the genetic analysis of traits (Williams, 1987).To determine the genetic control of adventitious root formation in white clover, we have identified and characterized a spontaneous mutant, designated Mortal, which is defective in nodal root primordium initiation. When grown at 20°C, Mortal plants lacked nodal root primordia but were normal in other aspects of shoot and root morphology. However, in some genetic backgrounds, Mortal was conditional, responding to either a temperature shift to 30°C or to the placing of stolon cuttings in water, by developing nodal roots. Here we describe Mortal and provide a genetic model for responses of the mutant to these temperature-shift and water treatments.     

Two receptor-like kinases required together for the establishment of Arabidopsis cotyledon primordia

Inter-regional signaling coordinates pattern formation in Arabidopsis thaliana embryos. However, little is known regarding the cells and molecules involved in inter-regional communication. We have characterized two related leucine-rich repeat receptor-like kinases (LRR-RLKs), RECEPTOR-LIKE PROTEIN KINASE1 (RPK1) and TOADSTOOL2 (TOAD2), which are required together for patterning the apical embryonic domain cell types that generate cotyledon primordia. Central domain protoderm patterning defects were always observed subjacent to the defective cotyledon primordia cell types in mutant embryos. In addition, RPK1-GFP and TOAD2-GFP translational fusions were both localized to the central domain protodermal cells when cotyledon primordia were first recognizable. We propose that RPK1 and TOAD2 are primarily required to maintain central domain protoderm cell fate and that the loss of this key embryonic cell type in mutant embryos results in patterning defects in other regions of the embryo including the failure to initiate cotyledon primordia.     

Standardized,Systemic Phenotypic Analysis of Umod C93F and Umod A227T Mutant Mice

Uromodulin-associated kidney disease (UAKD) summarizes different clinical features of an autosomal dominant heritable disease syndrome in humans with a proven uromodulin (UMOD) mutation involved. It is often characterized by hyperuricemia, gout, alteration of urine concentrating ability, as well as a variable rate of disease progression inconstantly leading to renal failure and histological alterations of the kidneys. We recently established the two Umod mutant mouse lines Umod ^C93F and Umod ^A227T on the C3H inbred genetic background both showing kidney defects analogous to those found in human UAKD patients. In addition, disease symptoms were revealed that were not yet described in other published mouse models of UAKD. To examine if further organ systems and/or metabolic pathways are affected by Umod mutations as primary or secondary effects, we describe a standardized, systemic phenotypic analysis of the two mutant mouse lines Umod ^A227T and Umod ^C93F in the German Mouse Clinic. Different genotypes as well as different ages were tested. Beside the already published changes in body weight, body composition and bone metabolism, the influence of the Umod mutation on energy metabolism was confirmed. Hematological analysis revealed a moderate microcytic and erythropenic anemia in older Umod mutant mice. Data of the other analyses in 7-10 month-old mutant mice showed single small additional effects.     

Candida albicans PEP12 Is Required for Biofilm Integrity and In Vivo Virulence

To investigate the role of the prevacuolar secretion pathway in biofilm formation and virulence in Candida albicans, we cloned and analyzed the C. albicans homolog of the Saccharomyces cerevisiae prevacuolar trafficking gene PEP12. C. albicans PEP12 encodes a deduced t-SNARE that is 28% identical to S. cerevisiae Pep12p, and plasmids bearing C. albicans PEP12 complemented the abnormal vacuolar morphology and temperature-sensitive growth of an S. cerevisiae pep12 null mutant. The C. albicans pep12 Δ null mutant was defective in endocytosis and vacuolar acidification and accumulated 40- to 60-nm cytoplasmic vesicles near the plasma membrane. Secretory defects included increased extracellular proteolytic activity and absent lipolytic activity. The pep12Δ null mutant was more sensitive to cell wall stresses and antifungal agents than the isogenic complemented strain or the control strain DAY185. Notably, the biofilm formed by the pep12Δ mutant was reduced in overall mass and fragmented completely upon the slightest disturbance. The pep12Δ mutant was markedly reduced in virulence in an in vitro macrophage infection model and an in vivo mouse model of disseminated candidiasis. These results suggest that C. albicans PEP12 plays a key role in biofilm integrity and in vivo virulence.In Saccharomyces cerevisiae, distinct secreted marker proteins are trafficked differentially through a prevacuolar compartment (PVC) prior to exocytosis (14). Furthermore, prevacuolar protein sorting genes play an important role in cargo transport in the prevacuolar branch of the exocytic pathway in S. cerevisiae (13, 15). By isolating dense- and light-vesicle populations in S. cerevisiae vps1 sec6-4, vps4 sec6-4, and pep12 sec6-4 mutants, it was observed that mutants blocked in this prevacuolar pathway missort marker proteins that are normally found in high-density post-Golgi compartment vesicles into low-density vesicles (15). Gurunathan et al. (13) also demonstrated these findings for vps1 and pep12 mutants with a late secretory mutant (snc1) background similar to that of the sec6-4 strains. These results indicate that some exocytic cargo, including the conditionally regulated soluble secretory proteins invertase and acid phosphatase, are differentially sorted through a PVC prior to exocytosis in the model yeast S. cerevisiae.To study the prevacuolar branch of exocytosis in Candida albicans and its role in virulence, we have previously cloned and analyzed the C. albicans prevacuolar trafficking genes VPS1 and VPS4. We demonstrated that C. albicans VPS4 is required for extracellular secretion of Sap2p and Sap4-6p and for virulence in an in vivo model of disseminated candidiasis (19, 20). C. albicans VPS1 is required for Sap2p secretion and biofilm formation (4). Interestingly, although the C. albicans null mutant lacking VPS4 forms a biofilm that is denser than that formed by the isogenic reintegrant strain, the conditional mutant lacking VPS1 expression forms a patchy biofilm of reduced density (4, 34). Thus, it appears that interference with normal prevacuolar trafficking affects both the secretion of virulence-associated proteins and biofilm formation.S. cerevisiae PEP12 encodes a 288-amino-acid syntaxin which regulates docking of Golgi compartment-derived transport vesicles at the PVC (3). Pep12p interacts with the v-SNARE Vti1p, and overexpression of Pep12p suppresses extracellular missorting of carboxypeptidase in the vti1 mutant (37). The S. cerevisiae pep12 null mutant displays a temperature-sensitive growth defect and is characterized by an enlarged vacuole with morphology defined as class D (3). A search of the C. albicans genome database identified a structural homolog of S. cerevisiae PEP12. Thus, the experiments described below were designed to determine whether the C. albicans PEP12 homolog is functionally homologous to S. cerevisiae PEP12 and to investigate its role in secretion, biofilm formation, and virulence.     

Peptidoglycan-Modifying Enzyme Pgp1 Is Required for Helical Cell Shape and Pathogenicity Traits in Campylobacter jejuni

The impact of bacterial morphology on virulence and transmission attributes of pathogens is poorly understood. The prevalent enteric pathogen Campylobacter jejuni displays a helical shape postulated as important for colonization and host interactions. However, this had not previously been demonstrated experimentally. C. jejuni is thus a good organism for exploring the role of factors modulating helical morphology on pathogenesis. We identified an uncharacterized gene, designated pgp1 (peptidoglycan peptidase 1), in a calcofluor white-based screen to explore cell envelope properties important for C. jejuni virulence and stress survival. Bioinformatics showed that Pgp1 is conserved primarily in curved and helical bacteria. Deletion of pgp1 resulted in a striking, rod-shaped morphology, making pgp1 the first C. jejuni gene shown to be involved in maintenance of C. jejuni cell shape. Pgp1 contributes to key pathogenic and cell envelope phenotypes. In comparison to wild type, the rod-shaped pgp1 mutant was deficient in chick colonization by over three orders of magnitude and elicited enhanced secretion of the chemokine IL-8 in epithelial cell infections. Both the pgp1 mutant and a pgp1 overexpressing strain – which similarly produced straight or kinked cells – exhibited biofilm and motility defects. Detailed peptidoglycan analyses via HPLC and mass spectrometry, as well as Pgp1 enzyme assays, confirmed Pgp1 as a novel peptidoglycan DL-carboxypeptidase cleaving monomeric tripeptides to dipeptides. Peptidoglycan from the pgp1 mutant activated the host cell receptor Nod1 to a greater extent than did that of wild type. This work provides the first link between a C. jejuni gene and morphology, peptidoglycan biosynthesis, and key host- and transmission-related characteristics.     

Circadian Control of Dendrite Morphology in the Visual System of Drosophila melanogaster

Background

In the first optic neuropil (lamina) of the fly''s visual system, monopolar cells L1 and L2 and glia show circadian rhythms in morphological plasticity. They change their size and shape during the day and night. The most pronounced changes have been detected in circadian size of the L2 axons. Looking for a functional significance of the circadian plasticity observed in axons, we examined the morphological plasticity of the L2 dendrites. They extend from axons and harbor postsynaptic sites of tetrad synaptic contacts from the photoreceptor terminals.

Methodology/Principal Findings

The plasticity of L2 dendrites was evaluated by measuring an outline of the L2 dendritic trees. These were from confocal images of cross sections of L2 cells labeled with GFP. They were in wild-type and clock mutant flies held under different light conditions and sacrified at different time points. We found that the L2 dendrites are longest at the beginning of the day in both males and females. This rhythm observed under a day/night regime (LD) was maintained in constant darkness (DD) but not in continuous light (LL). This rhythm was not present in the arrhythmic per⁰¹ mutant in LD or in DD. In the clock photoreceptor cry^b mutant the rhythm was maintained but its pattern was different than that observed in wild-type flies.

Conclusions/Significance

The results obtained showed that the L2 dendrites exhibit circadian structural plasticity. Their morphology is controlled by the per gene-dependent circadian clock. The L2 dendrites are longest at the beginning of the day when the daytime tetrad presynaptic sites are most numerous and L2 axons are swollen. The presence of the rhythm, but with a different pattern in cry^b mutants in LD and DD indicates a new role of cry in the visual system. The new role is in maintaining the circadian pattern of changes of the L2 dendrite length and shape.     

The fan-shaped and ellipsoid bodies of the brain central complex are involved in the control of courtship behavior and communicative sound production in Drosophila melanogaster males

To elucidate the role of the fan-shaped and ellipsoid bodies (FB and EB) of the central complex of Drosophila melanogaster brain in the control of male courtship behavior and singing, we analyzed characteristics of the courtship behavior and parameters of the communicative sound signals accompanying it in wild type flies and in flies from 5 mutant strains with various anatomical defects in FB and EB. The following strains of flies were used for experiments: Canton S (wild type, the control), eboKS263 with defects only in EB and ebo1041, ceb849, ceb892 and cbdKS96 with both the FB and EB damaged in different manner. The data obtained suggest that the FB and EB are involved: 1) in maintenance of a high courtship activity level, 2) in the control of accuracy of male following movements when courting a female, 3) in the control of the form and stability of sound elements in courtship songs, 4) in the control of the rhythmic structure of courtship songs determined by the stability of the pacemakers, and 5) in setting up a correspondence between the current behavior and the external situation.     

Cyclin G Functions as a Positive Regulator of Growth and Metabolism in Drosophila

In multicellular organisms, growth and proliferation is adjusted to nutritional conditions by a complex signaling network. The Insulin receptor/target of rapamycin (InR/TOR) signaling cascade plays a pivotal role in nutrient dependent growth regulation in Drosophila and mammals alike. Here we identify Cyclin G (CycG) as a regulator of growth and metabolism in Drosophila. CycG mutants have a reduced body size and weight and show signs of starvation accompanied by a disturbed fat metabolism. InR/TOR signaling activity is impaired in cycG mutants, combined with a reduced phosphorylation status of the kinase Akt1 and the downstream factors S6-kinase and eukaryotic translation initiation factor 4E binding protein (4E-BP). Moreover, the expression and accumulation of Drosophila insulin like peptides (dILPs) is disturbed in cycG mutant brains. Using a reporter assay, we show that the activity of one of the first effectors of InR signaling, Phosphoinositide 3-kinase (PI3K92E), is unaffected in cycG mutants. However, the metabolic defects and weight loss in cycG mutants were rescued by overexpression of Akt1 specifically in the fat body and by mutants in widerborst (wdb), the B''-subunit of the phosphatase PP2A, known to downregulate Akt1 by dephosphorylation. Together, our data suggest that CycG acts at the level of Akt1 to regulate growth and metabolism via PP2A in Drosophila.     

acr-23 Encodes a Monepantel-Sensitive Channel in Caenorhabditis elegans

Monepantel is a member of the recently identified class of anthelmintics known as the amino-acetonitrile derivatives (AADs). Monepantel controls all major gastro-intestinal nematodes in sheep including those that are resistant to the classical anthelmintics. Previous studies have shown that the Caenorhabditis elegans acr-23 and the Haemonchus contortus Hco-mptl-1 genes may be prominent targets of monepantel. With this discovery it became possible to investigate the mode of action of monepantel in nematodes at the molecular level. In the present study, we show that a C. elegans mutant acr-23 strain is fully rescued by expressing the wild-type acr-23 gene. Moreover, we present a new mutant allele, and characterize acr-23 alleles genetically. We also show that acr-23 is expressed in body wall muscle cells, and provide therefore a possible explanation for the paralysis caused by monepantel. Furthermore, genetic evidence suggests that the chaperone RIC-3 is required for expression of full monepantel resistance. Finally, we present reconstitution of the C. elegans ACR-23 receptor in Xenopus laevis oocytes and provide direct evidence of its modulation by monepantel. Conversely, co-injection of the chaperone RIC-3 had no impact for channel reconstitution in X. laevis oocytes. These results reinforce the involvement of the ACR-23 family in the mode of action of monepantel and advance our understanding of this new class of anthelmintics.     

Unc-45 Mutations in Caenorhabditis elegans Implicate a CRO1/She4p-like Domain in Myosin Assembly

The Caenorhabditis elegans unc-45 locus has been proposed to encode a protein machine for myosin assembly. The UNC-45 protein is predicted to contain an NH₂-terminal domain with three tetratricopeptide repeat motifs, a unique central region, and a COOH-terminal domain homologous to CRO1 and She4p. CRO1 and She4p are fungal proteins required for the segregation of other molecules in budding, endocytosis, and septation. Three mutations that lead to temperature-sensitive (ts) alleles have been localized to conserved residues within the CRO1/She4p-like domain, and two lethal alleles were found to result from stop codon mutations in the central region that would prevent translation of the COOH-terminal domain. Electron microscopy shows that thick filament accumulation in vivo is decreased by ∼50% in the CB286 ts mutant grown at the restrictive temperature. The thick filaments that assemble have abnormal structure. Immunofluorescence and immunoelectron microscopy show that myosins A and B are scrambled, in contrast to their assembly into distinct regions at the permissive temperature and in wild type. This abnormal structure correlates with the high degree of instability of the filaments in vitro as reflected by their extremely low yields and shortened lengths upon isolation. These results implicate the UNC-45 CRO1/She4p-like region in the assembly of myosin isoforms in C. elegans and suggest a possible common mechanism for the function of this UCS (UNC-45/CRO1/She4p) protein family.     

Early stages of leaf development in <Emphasis Type="Italic">has</Emphasis> mutant of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The elucidation of molecular mechanisms underlying the leaf development can be facilitated by the detailed anatomical study of leaf development mutants. We present an analysis of leaf anatomy and morphogenesis during early developmental stages in has mutant of Arabidopsis thaliana. The recessive has mutation affects a number of aspects in plant development, including the shape and size of both cotyledons and leaves. The earliest developmental observations suggest almost synchronous growth of the first two leaf primordia of has mutant. No significant disruption of the cell division pattern in the internal tissue is observed at the earliest stages of development, with the major anatomical difference compared to wild type primordia being the untimely maturation of mesophyll tissue cells in has mutant. At the stage of leaf blade formation, structure disruption becomes clearly evident, by irregular arrangement of the cell layers and the lack of polarity in juvenile has leaves. One distinguishing feature of the mutant leaf anatomy is the absence of mesophyll tissue differentiation. Altered has mutant leaf morphology could be at least partially accounted for by the ectopic STM activity that was found at the base of leaf primordia during early stages of leaf development in has plants.     

Labeling proteins with fluorine-18 using N-succinimidyl 4-[18F]fluorobenzoate

Two methods were investigated for the no-carrier-added synthesis of N-succinimidyl 4-[¹⁸F]fluorobenzoate (S[¹⁸F]FB). The first, an attempted nucleophilic aromatic substitution by [¹⁸F]fluoride on N-succinimidyl 4-nitrobenzoate was unsuccessful. The second method involved three steps; [¹⁸F]fluoride for trimethylammonium substitution on 4-formyl-N,N,N-trimethylanilinium triflate, oxidation to 4-[¹⁸F]fluorobenzoic acid, followed by reaction with N-hydroxysuccinimide and dicyclohexylcarbodiimide to form S[¹⁸F]FB. Total synthesis and purification time was 100 min and the overall radiochemical yield was 25% (decay corrected). A monoclonal antibody F(ab′)₂ fragment could be labeled in 40–60% yield by reaction with S[¹⁸F]FB for 15–20 min. The tissue distribution in normal mice and in vitro tumor binding of the antibody F(ab′)₂ labeled by reaction with S[¹⁸F]FB were comparable to those observed for the fragment after radioiodination using N-succinimidyl 4-[¹²⁵I]iodobenzoate.     

Effect of Age on Body Wall Cuticle Morphology of Heterodera schachtii Schmidt females

Fine structure of the body wall cuticle of Heterodera schachtii is compared with respect to age and body region of the female. The cuticle is more complex than previously reported. In newly molted females only layers A, B, and C are present, but 4 weeks after the final molt a thin D layer is present between the midbody and base of the cone. This D layer is absent in the cone of H. schachtii, regardless of age. As females age, an additional layer E is produced and includes zones E₁ and E₂. Zone El apparently is unique to H. schachtii, whereas E₂ is likely to be homologous with a similar layer in Atalodera. In the cone of old females (ca. 8 weeks after the final molt) of H. schachtii, the two zones become irregular in shape and comprise bullae. The presence of a thin D layer in Heterodera strengthens the previous hypothesis of a single ancestor of cyst nematodes.     

Serial Examination of an Inducible and Reversible Dilated Cardiomyopathy in Individual Adult Drosophila

Recent work has demonstrated that Drosophila can be used as a model of dilated cardiomyopathy, defined as an enlarged cardiac chamber at end-diastole when the heart is fully relaxed and having an impaired systolic function when the heart is fully contracted. Gene mutations that cause cardiac dysfunction in adult Drosophila can result from abnormalities in cardiac development or alterations in post-developmental heart function. To clarify the contribution of transgene expression to post-developmental cardiac abnormalities, we applied strategies to examine the temporal and spacial effects of transgene expression on cardiac function. We engineered transgenic Drosophila based on the well-characterized temperature-sensitive Gal80 protein in the context of the bipartite Gal4/UAS transgenic expression system in Drosophila employing the cardiac specific driver, tinCΔ4-Gal4. Then, we developed a strategy using optical coherence tomography to serially measure cardiac function in the individual flies over time course of several days. As a proof of concept we examined the effects of the expression of a human mutant delta-sarcoglycan associated with familial heart failure and observed a reversible, post-developmental dilated cardiomyopathy in Drosophila. Our results show that the unique imaging strategy based on the non-destructive, non-invasive properties of optical coherence tomography can be applied to serially examine cardiac function in individual adult flies. Furthermore, the induction and reversal of cardiac transgene expression can be investigated in adult flies thereby providing insight into the post-developmental effects of transgene expression.