The Conserved MAPK Site in E(spl)-M8, an Effector of Drosophila Notch Signaling, Controls Repressor Activity during Eye Development

The specification of patterned R8 photoreceptors at the onset of eye development depends on timely inhibition of Atonal (Ato) by the Enhancer of split (E(spl) repressors. Repression of Ato by E(spl)-M8 requires the kinase CK2 and is inhibited by the phosphatase PP2A. The region targeted by CK2 harbors additional conserved Ser residues, raising the prospect of regulation via multi-site phosphorylation. Here we investigate one such motif that meets the consensus for modification by MAPK, a well-known effector of Epidermal Growth Factor Receptor (EGFR) signaling. Our studies reveal an important role for the predicted MAPK site of M8 during R8 birth. Ala/Asp mutations reveal that the CK2 and MAPK sites ensure that M8 repression of Ato and the R8 fate occurs in a timely manner and at a specific stage (stage-2/3) of the morphogenetic furrow (MF). M8 repression of Ato is mitigated by halved EGFR dosage, and this effect requires an intact MAPK site. Accordingly, variants with a phosphomimetic Asp at the MAPK site exhibit earlier (inappropriate) activity against Ato even at stage-1 of the MF, where a positive feedback-loop is necessary to raise Ato levels to a threshold sufficient for the R8 fate. Analysis of deletion variants reveals that both kinase sites (CK2 and MAPK) contribute to ‘cis’-inhibition of M8. This key regulation by CK2 and MAPK is bypassed by the E(spl)D mutation encoding the truncated protein M8*, which potently inhibits Ato at stage-1 of R8 birth. We also provide evidence that PP2A likely targets the MAPK site. Thus multi-site phosphorylation controls timely onset of M8 repressor activity in the eye, a regulation that appears to be dispensable in the bristle. The high conservation of the CK2 and MAPK sites in the insect E(spl) proteins M7, M5 and Mγ, and their mammalian homologue HES6, suggest that this mode of regulation may enable E(spl)/HES proteins to orchestrate repression by distinct tissue-specific mechanisms, and is likely to have broader applicability than has been previously recognized

The Ser/Thr Phosphatase PP2A Regulatory Subunit Widerborst Inhibits Notch Signaling

Drosophila Enhancer of split M8, an effector of Notch signaling, is regulated by protein kinase CK2. The phosphatase PP2A is thought to play an opposing (inhibitory) role, but the identity of the regulatory subunit was unknown. The studies described here reveal a role for the PP2A regulatory subunit widerborst (wdb) in three developmental contexts; the bristle, wing and the R8 photoreceptors of the eye. wdb overexpression elicits bristle and wing defects akin to reduced Notch signaling, whereas hypomorphic mutations in this PP2A subunit elicit opposite effects. We have also evaluated wdb functions using mutations in Notch and E(spl) that affect the eye. We find that the eye and R8 defects of the well-known Nspl mutation are enhanced by a hypomorphic allele of wdb, whereas they are strongly rescued by wdb overexpression. Similarly, ectopic wdb rescues the eye and R8 defects of the E(spl)D mutation, which affects the m8 gene. In addition, wdb overexpression also rescues the bristle defects of ectopically expressed M8, or the eye and R8 defects of its CK2 phosphomimetic variant M8-S159D. The latter finding suggests that PP2A may target M8 at highly conserved residues in the vicinity of the CK2 site, whose phosphorylation controls repression of Atonal and the R8 fate. Together, the studies identify PP2A-Wdb as a participant in Notch signaling, and suggest that M8 activity is controlled by phosphorylation and dephosphorylation. The conservation of the phosphorylation sites between Drosophila E(spl) and the HES/HER proteins from mammals, reptiles, amphibians, birds and fish raises the prospect that this mode of regulation is widespread

Stealth-spectacles: the discursive waves of the nuclear Asian seascape

When compared to the bombast of nuclear tests, nuclear submarines come with the relatively quiet fantasy of victory-to-come against neighbouring nuclear adversaries. Such political expressions are making their mark in Indian popular culture that hitherto had little commentary to offer on submarines. Outlets such as film and digital media on submarines rest on an aporia that resonates across the pleats and folds of secrecy and publicity: there is a felt need to keep covert underwater vessels under wraps, yet also an irrepressible desire to glorify the technological achievement and political posturings enabled by thesecond strike capability of a nuclear armed and powered submarine. Highlighting the tensile allure of both stealth and spectacle, the article considers the ways submarines make a mark in Indian audio-visual and digital media alongside the affective resonance of submarines more widely. By understanding their hegemonic dynamics, we can begin to raise questions about the ongoing nuclearisation of the Asian region and neighbouring arterial seas described here as the Asian seascape

Management of adults with primary frozen shoulder in secondary care (UK FROST): a multicentre, pragmatic, three-arm, superiority randomised clinical trial

Manipulation under anaesthesia and arthroscopic capsular release are costly and invasive treatments for frozen shoulder, but their effectiveness remains uncertain. We compared these two surgical interventions with early structured physiotherapy plus steroid injection. In this multicentre, pragmatic, three-arm, superiority randomised trial, patients referred to secondary care for treatment of primary frozen shoulder were recruited from 35 hospital sites in the UK. Participants were adults (≥18 years) with unilateral frozen shoulder, characterised by restriction of passive external rotation (≥50%) in the affected shoulder. Participants were randomly assigned (2:2:1) to receive manipulation under anaesthesia, arthroscopic capsular release, or early structured physiotherapy. In manipulation under anaesthesia, the surgeon manipulated the affected shoulder to stretch and tear the tight capsule while the participant was under general anaesthesia, supplemented by a steroid injection. Arthroscopic capsular release, also done under general anaesthesia, involved surgically dividing the contracted anterior capsule in the rotator interval, followed by manipulation, with optional steroid injection. Both forms of surgery were followed by postprocedural physiotherapy. Early structured physiotherapy involved mobilisation techniques and a graduated home exercise programme supplemented by a steroid injection. Both early structured physiotherapy and postprocedural physiotherapy involved 12 sessions during up to 12 weeks. The primary outcome was the Oxford Shoulder Score (OSS; 0-48) at 12 months after randomisation, analysed by initial randomisation group. We sought a target difference of 5 OSS points between physiotherapy and either form of surgery, or 4 points between manipulation and capsular release. The trial registration is ISRCTN48804508. Between April 1, 2015, and Dec 31, 2017, we screened 914 patients, of whom 503 (55%) were randomly assigned. At 12 months, OSS data were available for 189 (94%) of 201 participants assigned to manipulation (mean estimate 38·3 points, 95% CI 36·9 to 39·7), 191 (94%) of 203 participants assigned to capsular release (40·3 points, 38·9 to 41·7), and 93 (94%) of 99 participants assigned to physiotherapy (37·2 points, 35·3 to 39·2). The mean group differences were 2·01 points (0·10 to 3·91) between the capsular release and manipulation groups, 3·06 points (0·71 to 5·41) between capsular release and physiotherapy, and 1·05 points (-1·28 to 3·39) between manipulation and physiotherapy. Eight serious adverse events were reported with capsular release and two with manipulation. At a willingness-to-pay threshold of £20 000 per quality-adjusted life-year, manipulation under anaesthesia had the highest probability of being cost-effective (0·8632, compared with 0·1366 for physiotherapy and 0·0002 for capsular release). All mean differences on the assessment of shoulder pain and function (OSS) at the primary endpoint of 12 months were less than the target differences. Therefore, none of the three interventions were clinically superior. Arthoscopic capsular release carried higher risks, and manipulation under anaesthesia was the most cost-effective. The National Institute for Health Research Health Technology Assessment programme. [Abstract copyright: Copyright © 2020 The Author(s). Publishedx by Elsevier Ltd. This is an Open Access article under the CC BY-NC-ND 4.0 license. Published by Elsevier Ltd.. All rights reserved.

A Common Standard for Film Production Incentives Impact Analysis

Compares in detail two impact analysis reports done on Massachusetts film production incentives (FPIs) for the year 2011 to understand the different evaluation variables used by evaluators (key government agencies, industry stakeholders, and third parties) to analyze the impact of the FPIs on the state economy, job creation, etc. To understand the discrepancy in reports, this paper examines the origin and proliferation of FPIs in the light of two most important types of policy convergences namely convergence through emulation and convergence through penetration. This study, then compares nine impact analysis reports starting from 2002 to 2013 to list down variables used in the analysis. This study concludes with the author\u27s adjustments to the impact analysis reports by HR&A and Department of Revenue on Massachusetts FPIs using common variables