echinus, required for interommatidial cell sorting and cell death in the Drosophila pupal retina, encodes a protein with homology to ubiquitin-specific proteases

Background: Programmed cell death is used to remove excess cells between ommatidia in the Drosophila pupal retina. This death is required to establish the crystalline, hexagonal packing of ommatidia that characterizes the adult fly eye. In previously described echinus mutants, interommatidial cell sorting, which precedes cell death, occurred relatively normally. Interommatidial cell death was partially suppressed, resulting in adult eyes that contained excess pigment cells, and in which ommatidia were mildly disordered. These results have suggested that echinus functions in the pupal retina primarily to promote interommatidial cell death. Results: We generated a number of new echinus alleles, some of which are likely null mutants. Analysis of these alleles provides evidence that echinus has roles in cell sorting as well as cell death. echinus encodes a protein with homology to ubiquitin-specific proteases, which cleave ubiquitin-conjugated proteins at the ubiquitin C-terminus. The echinus locus encodes multiple splice forms, including two proteins that lack residues thought to be critical for deubiquitination activity. Surprisingly, ubiquitous expression in the eye of versions of Echinus that lack residues critical for ubiquitin specific protease activity, as well as a version predicted to be functional, rescue the echinus loss-of-function phenotype. Finally, genetic interactions were not detected between echinus loss and gain-of-function and a number of known apoptotic regulators. These include Notch, EGFR, the caspases Dronc, Drice, Dcp-1, Dream, the caspase activators, Rpr, Hid, and Grim, the caspase inhibitor DIAP1, and Lozenge or Klumpfuss. Conclusions: The echinus locus encodes multiple splice forms of a protein with homology to ubiquitin-specific proteases, but protease activity is unlikely to be required for echinus function, at least when echinus is overexpressed. Characterization of likely echinus null alleles and genetic interactions suggests that echinus acts at a novel point(s) to regulate interommatidial cell sorting and/or cell death in the fly eye

Repeatability and Reproducibility of Macular Thickness Measurements Using Fourier Domain Optical Coherence Tomography

Aim: To evaluate repeatability and reproducibility of macular thickness measurements in visually normal eyes using the Topcon 3D OCT-1000. Methods: Phase 1 investigated scan repeatability, the effect of age and pupil dilation. Two groups (6 younger and 6 older participants) had one eye scanned 5 times pre and post- dilation by 1 operator. Phase 2 investigated between-operator, within and between-visit reproducibility. 10 participants had 1 un-dilated eye scanned 3 times on 2 separate visits by 2 operators. Results: Phase 1: No significant difference existed between repeat scans (p=0.75) and no significant difference was found pre- and post-dilation (p=0.54). In the younger group variation was low (95% limits ± 3.62 m) and comparable across all retinal regions. The older group demonstrated greater variation (95% limits ± 7.6 m). Phase 2: For a given retinal location, 95% confidence limits for within-operator, within-visit reproducibility was 5.16 m. This value increased to 5.56 m for the same operator over two visits and to 6.18 m for two operators over two visits. Conclusion: A high level repeatability, close to 6 m, of macular thickness measurement is possible using the 3D OCT- 1000. Measured differences in macular thickness between successive visits that exceed 6 m in pre-presbyopic individuals are therefore likely to reflect actual structural change. OCT measures are more variable in older individuals and it is advisable to take a series of scans so that outliers can be more easily identified

An investigation of dendritic delay in octopus cells of the mammalian cochlear nucleus

Octopus cells, located in the mammalian auditory brainstem, receive their excitatory synaptic input exclusively from auditory nerve fibers (ANFs). They respond with accurately timed spikes but are broadly tuned for sound frequency. Since the representation of information in the auditory nerve is well understood, it is possible to pose a number of questions about the relationship between the intrinsic electrophysiology, dendritic morphology, synaptic connectivity, and the ultimate functional role of octopus cells in the brainstem. This study employed a multi-compartmental Hodgkin-Huxley model to determine whether dendritic delay in octopus cells improves synaptic input coincidence detection in octopus cells by compensating for the cochlear traveling wave delay. The propagation time of post-synaptic potentials from synapse to soma was investigated. We found that the total dendritic delay was approximately 0.275 ms. It was observed that low-threshold potassium channels in the dendrites reduce the amplitude dependence of the dendritic delay of post-synaptic potentials. As our hypothesis predicted, the model was most sensitive to acoustic onset events, such as the glottal pulses in speech when the synaptic inputs were arranged such that the model's dendritic delay compensated for the cochlear traveling wave delay across the ANFs. The range of sound frequency input from ANFs was also investigated. The results suggested that input to octopus cells is dominated by high frequency ANFs

BAFFLES: Bayesian Ages for Field Lower-mass Stars

Funding: R.D. acknowledges support from the Fonds de Recherche du Québec. Supported by NSF grants AST-1411868 (E.L.N., B.M.), and AST-1518332 (R.J.D.R.). Supported by NASA grants NNX14AJ80G (E.L.N., B.M.), NNX15AC89G and NNX15AD95G (B.M., R.J.D.R.), 80NSSC17K0535 (B.M., E.L.N., R.J.D.R), and NASA Hubble Fellowship grant HST-HF2-51405.001-A (I.C.).Age is a fundamental parameter of stars, yet in many cases, ages of individual stars are presented without robust estimates of the uncertainty. We have developed a Bayesian framework, BAFFLES, to produce the age posterior for a star from its calcium emission strength (log(R′HK)) or lithium abundance (Li EW) and B − V color. We empirically determine the likelihood functions for calcium and lithium as functions of age from literature measurements of stars in benchmark clusters with well-determined ages. We use a uniform prior on age, which reflects a uniform star formation rate. The age posteriors we derive for several test cases are consistent with literature ages found from other methods. BAFFLES represents a robust method to determine the age posterior probability distribution for any field star with 0.45 ≤ B − V ≤ 0.9 and a measurement of R′HK and/or 0.35 ≤ B − V ≤ 1.9 and measured Li EW. We compile colors, R′HK, and Li EW from over 2630 nearby field stars from the literature, and present the derived BAFFLES age posterior for each star.PostprintPeer reviewe

Li zoning in zircon as a potential geospeedometer and peak temperature indicator

Zircon Li concentrations and δ[superscript 7]Li values may potentially trace crustal recycling because continental and mantle-derived zircons yield distinct values. The usefulness of these differences may depend upon the retentivity of zircon to Li concentrations and isotopic ratios. Given the relatively high Li diffusivities measured by Cherniak and Watson (Contrib Mineral Petrol 160: 383–390, 2010), we sought to discover the scenarios under which Li mobility might be inhibited by charge-compensating cations. Toward this end, we conducted “in” diffusion experiments in which Li depth profiles of synthetic Lu-doped, P-doped, and undoped zircon were determined by nuclear reaction analysis. In separate experiments, Li was ion-implanted at depth within polished natural zircon slabs to form a Gaussian Li concentration profile. Diffusively relaxed concentration profiles were measured after heating the slabs to determine diffusivities. In all experiments, which ranged from 920 to 650 °C, calculated diffusivities are in agreement with a previously established Arrhenius relationship calibrated on trace-element-poor Mud Tank zircon. Our revised Arrhenius relationship that includes both datasets is: D[subscript Li] = 9.60 x 10[superscript -7] exp [-278 ± 8 kJ mol[superscript - 1]/RT] m[superscript 2] s[superscript - 1] We also observed that synthetic sector-zoned zircon exhibits near-step-function Li concentration profiles across sectors that correlate with changes in the rare earth element (REE) and P concentrations. This allowed us to examine how Li diffusion might couple with REE diffusion in a manner different than that described above. In particular, re-heating these grains revealed significant Li migration, but no detectable migration of the rare earth elements. Thus, unlike most elements in zircon which are not mobile at the micrometer scale under most time–temperature paths in the crust, Li zoning, relaxation of zoning, or lack of zoning altogether could be used to reveal time–temperature information. Discrete ~10 μm concentration zones of Li within zircon may be partially preserved at 700 °C for tens to hundreds of years, and at 450 °C for millions of years. In this regard, Li zoning in zircon holds significant potential as a geospeedometer, and in some instances as a qualitative indicator of the maximum temperature experienced by the zircon

Comparison of three RNA amplification methods as sources of DNA for sequencing

DNA products generated from a region of the measles virus genome by three RNA reverse transcription and amplification methods were cloned and sequenced, and the results were compared in order to evaluate the methods' relative fidelities. The methods were: (i) reverse transcription followed by a nested polymerase chain reaction (RT-nPCR), (ii) a combined RT-PCR using rTth polymerase and (iii) nucleic acid sequence-based amplification (NASBA). NASBA was followed by RT-PCR with rTth polymerase or RT using AMV reverse transcriptase to generate DNA products for cloning. Products from all three sets of reactions were cloned into a vector, pT7Blue, and 790 bp of cloned DNA were sequenced and analyzed for base changes to determine the error rates for each amplification method. Sequence analysis of cloned RT-nPCR products showed no errors, whereas cloned rTth mediated RT-PCR products possessed an error rate of 0.38 and cloned NASBA products 0.38. Products generated by NASBA followed by RT-PCR with rTth polymerase possessed an error rate of 1.9. The results indicated that cloned DNA products generated by RT-nPCRs possessed least errors and that for NASBA, RT of reaction products before cloning and sequencing was preferable to using RT-PCR

Comparison of three RNA amplification methods as sources of DNA for sequencing

DNA products generated from a region of the measles virus genome by three RNA reverse transcription and amplification methods were cloned and sequenced, and the results were compared in order to evaluate the methods' relative fidelities. The methods were: (i) reverse transcription followed by a nested polymerase chain reaction (RT-nPCR), (ii) a combined RT-PCR using rTth polymerase and (iii) nucleic acid sequence-based amplification (NASBA). NASBA was followed by RT-PCR with rTth polymerase or RT using AMV reverse transcriptase to generate DNA products for cloning. Products from all three sets of reactions were cloned into a vector, pT7Blue, and 790 bp of cloned DNA were sequenced and analyzed for base changes to determine the error rates for each amplification method. Sequence analysis of cloned RT-nPCR products showed no errors, whereas cloned rTth mediated RT-PCR products possessed an error rate of 0.38 and cloned NASBA products 0.38. Products generated by NASBA followed by RT-PCR with rTth polymerase possessed an error rate of 1.9. The results indicated that cloned DNA products generated by RT-nPCRs possessed least errors and that for NASBA, RT of reaction products before cloning and sequencing was preferable to using RT-PCR

Pleural Fluid Mesothelin as an Adjunct to the Diagnosis of Pleural Malignant Mesothelioma

Rationale:. The diagnosis of pleural malignant mesothelioma (MM) by effusion cytology may be difficult and is currently controversial. Effusion mesothelin levels are increased in patients with MM but the clinical role of this test is uncertain. Objectives:. To determine the clinical value of measuring mesothelin levels in pleural effusion supernatant to aid diagnosis of MM. Methods and Measurements. Pleural effusion samples were collected prospectively from 1331 consecutive patients. Mesothelin levels were determined by commercial ELISA in effusions and their relationship to concurrent pathology reporting and final clinical diagnosis was determined. Results:. 2156 pleural effusion samples from 1331 individuals were analysed. The final clinical diagnosis was 183 MM, 436 non-MM malignancy, and 712 nonmalignant effusions. Effusion mesothelin had a sensitivity of 67% for MM at 95% specificity. Mesothelin was elevated in over 47% of MM cases in effusions obtained before definitive diagnosis of MM was established. In the setting of inconclusive effusion cytology, effusion mesothelin had a positive predictive value of 79% for MM and 94% for malignancy. Conclusions:. A mesothelin-positive pleural effusion, irrespective of the identification of malignant cells, indicates the likely presence of malignancy and adds weight to the clinical rationale for further investigation to establish a malignant diagnosis