Controlled chemical modifications of single-walled carbon nanotubes (SWCNTs) that tune their useful properties have been sought for multiple applications. We found that beneficial optical changes in ...SWCNTs resulted from introducing low concentrations of oxygen atoms. Stable covalently oxygen-doped nanotubes were prepared by exposure to ozone and then light. Treated samples showed distinct structure-specific near-infrared fluorescence at wavelengths 10 to 15% longer than displayed by pristine semiconducting SWCNTs. Dopant sites harvest light energy absorbed in undoped nanotube regions by trapping mobile excitons. The oxygen-doped SWCNTs are much easier to detect and image than pristine SWCNTs because they give stronger near-infrared emission and do not absorb at the shifted emission wavelength.
The larvae of Drosophila melanogaster grow rapidly through use of a highly truncated cell cycle in which mitosis is entirely eliminated. The Drosophila homolog of the protooncogene transcription ...factor Myc plays a major role in promoting this endopolyploid (EP) growth. We have previously determined that the gene jim lovell (lov), which encodes a member of the BTB/POZ (Bric-a-brac, Tramtrack, Broad/Pox virus zinc finger) domain family of transcription factors, is also required for EP growth in one larval tissue, the trachea. Here we show that lov promotes EP growth in three further tissues indicating a fundamental role in this process. However, epistasis experiments revealed heterogeneity in lov's action in these tissues. Whereas in the tracheae and salivary glands lov acts downstream of Myc, in the fat body, reduced expression of lov does not impede the action of Myc, indicating an upstream action for the gene. We show here that lov's regulation of the gene uninflatable (uif) in the tracheae is a component of this difference. uif is required for tracheal EP growth downstream of Myc and lov but has no equivalent role in the fat body. Although Uif is a transmembrane component of the plasma membrane in the tracheae, its action downstream of Myc suggests an intracellular role for the protein in the tracheae. In addition to regulating uif expression in some tissues we also show that lov locates to the nucleolus, indicating it can function in both polymerase I and polymerase II transcriptional events. Our major finding is that tissue-specific mechanisms can interact with universal growth promotion by Myc to generate the individual endopolyploid organs of the larvae.
The Drosophila protein Jim Lovell (Lov) is a putative transcription factor of the BTB/POZ (Bric- a-Brac/Tramtrack/Broad/ Pox virus and Zinc finger) domain class that is expressed in many elements of ...the developing larval nervous system. It has roles in innate behaviors such as larval locomotion and adult courtship. In performing tissue-specific knockdown with the Gal4-UAS system we identified a new behavioral phenotype for lov: larvae failed to burrow into their food during their growth phase and then failed to tunnel into an agarose substratum during their wandering phase. We determined that these phenotypes originate in a previously unrecognized role for lov in the tracheae. By using tracheal-specific Gal4 lines, Lov immunolocalization and a lov enhancer trap line, we established that lov is normally expressed in the tracheae from late in embryogenesis through larval life. Using an assay that monitors food burrowing, substrate tunneling and death we showed that lov tracheal knockdown results in tracheal fluid-filling, producing hypoxia that activates the aberrant behaviors and inhibits development. We investigated the role of lov in the tracheae that initiates this sequence of events. We discovered that when lov levels are reduced, the tracheal cells are smaller, more numerous and show lower levels of endopolyploidization. Together our findings indicate that Lov is necessary for tracheal endoreplicative growth and that its loss in this tissue causes loss of tracheal integrity resulting in chronic hypoxia and abnormal burrowing and tunneling behavior.
The ability of near-infrared fluorescence imaging to detect single-walled carbon nanotubes (SWNTs) in organisms and biological tissues has been explored using Drosophila melanogaster (fruit flies). ...Drosophila larvae were raised on food containing approximately 10 ppm of disaggregated SWNTs. Their viability and growth were not reduced by nanotube ingestion. Near-IR nanotube fluorescence was imaged from intact living larvae, and individual nanotubes in dissected tissue specimens were imaged, structurally identified, and counted to estimate a biodistribution.
In the first in vivo demonstration of spectral triangulation, biocompatible composites of single-walled carbon nanotubes in Matrigel have been surgically implanted into mouse ovaries and then ...noninvasively detected and located. This optical method deduces the three-dimensional position of a short-wave IR emission source from the wavelength-dependent attenuation of fluorescence in tissues. Measurements were performed with a second-generation optical scanner that uses a light-emitting diode matrix emitting at 736 nm for diffuse specimen excitation. The intrinsic short-wave IR fluorescence of the nanotubes was collected at various positions on the specimen surface, spectrally filtered, and detected by a photon-counting InGaAs avalanche photodiode. Sensitivity studies showed a detection limit of ∼120 pg of nanotubes located beneath ∼3 mm of tissue. In addition, the mass and location of implanted nanotubes could be deduced through spectral triangulation with sub-millimeter accuracy, as validated with the aid of magnetic resonance imaging (MRI) data. Dual-modality imaging combining spectral triangulation with computed tomography or MRI will allow accurate registration of emission centers with anatomical features. These results are a step toward the future use of probes with targeting agents such as antibodies linked to nanotube tags for the noninvasive detection and imaging of tumors in preclinical research on small animals. Translation to the clinic could aid in early detection of ovarian cancer and identification of metastases for resection during primary surgery.
Space travel presents unlimited opportunities for exploration and discovery, but requires better understanding of the biological consequences of long-term exposure to spaceflight. Immune function in ...particular is relevant for space travel. Human immune responses are weakened in space, with increased vulnerability to opportunistic infections and immune-related conditions. In addition, microorganisms can become more virulent in space, causing further challenges to health. To understand these issues better and to contribute to design of effective countermeasures, we used the Drosophila model of innate immunity to study immune responses in both hypergravity and spaceflight. Focusing on infections mediated through the conserved Toll and Imd signaling pathways, we found that hypergravity improves resistance to Toll-mediated fungal infections except in a known gravitaxis mutant of the yuri gagarin gene. These results led to the first spaceflight project on Drosophila immunity, in which flies that developed to adulthood in microgravity were assessed for immune responses by transcription profiling on return to Earth. Spaceflight alone altered transcription, producing activation of the heat shock stress system. Space flies subsequently infected by fungus failed to activate the Toll pathway. In contrast, bacterial infection produced normal activation of the Imd pathway. We speculate on possible linkage between functional Toll signaling and the heat shock chaperone system. Our major findings are that hypergravity and spaceflight have opposing effects, and that spaceflight produces stress-related transcriptional responses and results in a specific inability to mount a Toll-mediated infection response.
A detailed analysis is presented of the diffractive deep-inelastic scattering process ep→eXY, where Y is a proton or a low mass proton excitation carrying a fraction 1-xIP>0.95 of the incident proton ...longitudinal momentum and the squared four-momentum transfer at the proton vertex satisfies |t|<1 GeV2. Using data taken by the H1 experiment, the cross section is measured for photon virtualities in the range 3.5≤Q2≤1600 GeV2, triple differentially in xIP, Q2 and β=x/xIP, where x is the Bjorken scaling variable. At low xIP, the data are consistent with a factorisable xIP dependence, which can be described by the exchange of an effective pomeron trajectory with intercept αIP(0)=1.118±0.008(exp.)+0.029-0.010(model). Diffractive parton distribution functions and their uncertainties are determined from a next-to-leading order DGLAP QCD analysis of the Q2 and β dependences of the cross section. The resulting gluon distribution carries an integrated fraction of around 70% of the exchanged momentum in the Q2 range studied. Total and differential cross sections are also measured for the diffractive charged current process e+p→ν̄eXY and are found to be well described by predictions based on the diffractive parton distributions. The ratio of the diffractive to the inclusive neutral current ep cross sections is studied. Over most of the kinematic range, this ratio shows no significant dependence on Q2 at fixed xIP and x or on x at fixed Q2 and β.
Innate behaviors have their origins in the specification of neural fates during development. Within Drosophila, BTB (Bric-a-brac,Tramtrack, Broad) domain proteins such as Fruitless are known to play ...key roles in the neural differentiation underlying such responses. We previously identified a gene, which we have termed jim lovell (lov), encoding a BTB protein with a role in gravity responses. To understand more fully the behavioral roles of this gene we have investigated its function through several approaches. Transcript and protein expression patterns have been examined and behavioral phenotypes of new lov mutations have been characterized. Lov is a nuclear protein, suggesting a role as a transcriptional regulator, as for other BTB proteins. In late embryogenesis, Lov is expressed in many CNS and PNS neurons. An examination of the PNS expression indicates that lov functions in the late specification of several classes of sensory neurons. In particular, only two of the five abdominal lateral chordotonal neurons express Lov, predicting functional variation within this highly similar group. Surprisingly, Lov is also expressed very early in embryogenesis in ways that suggests roles in morphogenetic movements, amnioserosa function and head neurogenesis. The phenotypes of two new lov mutations that delete adjacent non-coding DNA regions are strikingly different suggesting removal of different regulatory elements. In lov(47) , Lov expression is lost in many embryonic neurons including the two lateral chordotonal neurons. lov(47) mutant larvae show feeding and locomotor defects including spontaneous backward movement. Adult lov(47) males perform aberrant courtship behavior distinguished by courtship displays that are not directed at the female. lov(47) adults also show more defective negative gravitaxis than the previously isolated lov(91Y) mutant. In contrast, lov(66) produces largely normal behavior but severe female sterility associated with ectopic lov expression in the ovary. We propose a negative regulatory role for the DNA deleted in lov(66) .
In Drosophila testis, myosin VI plays a special role, distinct from its motor function, by anchoring components to the unusual actin-based structures (cones) that are required for spermatid ...individualization. For this, the two calmodulin (CaM) light-chain molecules of myosin VI are replaced by androcam (ACaM), a related protein with 67% identity to CaM. Although ACaM has a similar bi-lobed structure to CaM, with two EF hand-type Ca
binding sites per lobe, only one functional Ca
binding site operates in the amino-terminus. To understand this light chain substitution, we used hydrogen-deuterium exchange mass spectrometry (HDX-MS) to examine dynamic changes in ACaM and CaM upon Ca
binding and interaction with the two CaM binding motifs of myosin VI (insert2 and IQ motif). HDX-MS reveals that binding of Ca
to ACaM destabilizes its N-lobe but stabilizes the entire C-lobe, whereas for CaM, Ca
binding induces a pattern of alternating stabilization/destabilization throughout. The conformation of this stable holo-C-lobe of ACaM seems to be a "prefigured" version of the conformation adopted by the holo-C-lobe of CaM for binding to insert2 and the IQ motif of myosin VI. Strikingly, the interaction of holo-ACaM with either peptide converts the holo-N-lobe to its Ca
-free, more stable, form. Thus, ACaM in vivo should bind the myosin VI light chain sites in an apo-N-lobe/holo-C-lobe state that cannot fulfill the Ca
-related functions of holo-CaM required for myosin VI motor assembly and activity. These findings indicate that inhibition of myosin VI motor activity is a precondition for transition to an anchoring function.
Androcam replaces calmodulin as a tissue-specific myosin VI light chain on the actin cones that mediate D. melanogaster spermatid individualization. We show that the androcam structure and its ...binding to the myosin VI structural (Insert 2) and regulatory (IQ) light chain sites are distinct from those of calmodulin and provide a basis for specialized myosin VI function. The androcam N lobe noncanonically binds a single Ca ²⁺ and is locked in a “closed” conformation, causing androcam to contact the Insert 2 site with its C lobe only. Androcam replacing calmodulin at Insert 2 will increase myosin VI lever arm flexibility, which may favor the compact monomeric form of myosin VI that functions on the actin cones by facilitating the collapse of the C-terminal region onto the motor domain. The tethered androcam N lobe could stabilize the monomer through contacts with C-terminal portions of the motor or recruit other components to the actin cones. Androcam binds the IQ site at all calcium levels, constitutively mimicking a conformation adopted by calmodulin only at intermediate calcium levels. Thus, androcam replacing calmodulin at IQ will abolish a Ca ²⁺-regulated, calmodulin-mediated myosin VI structural change. We propose that the N lobe prevents androcam from interfering with other calmodulin-mediated Ca ²⁺ signaling events. We discuss how gene duplication and mutations that selectively stabilize one of the many conformations available to calmodulin support the molecular evolution of structurally and functionally distinct calmodulin-like proteins.
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