145 research outputs found
Drosophila Homeodomain-Interacting Protein Kinase (Hipk) Phosphorylates the Hippo/Warts Signalling Effector Yorkie
Developmental growth and patterning are regulated by an interconnected signalling
network of several pathways. In Drosophila, the Warts (Wts) kinase, a component of the Hippo
signalling pathway, plays an essential role in regulating transcription and growth by phosphorylating
its substrate Yorkie (Yki). The phosphorylation of Yki critically influences its localisation and
activity as a transcriptional coactivator. In this study, we identified the homeodomain-interacting
protein kinase (Hipk) as another kinase that phosphorylates Yki and mapped several sites of Yki
phosphorylated by Hipk, using in vitro analysis: Ser168, Ser169/Ser172 and Ser255. These sites
might provide auxiliary input for Yki regulation in vivo, as transgenic flies with mutations in these
show prominent phenotypes; Hipk, therefore, represents an additional upstream regulator of Yki
that works in concert with Wts
Analyse von Wechselwirkungen der Homeodomänen Interagierenden Proteinkinase (Hipk) mit Genen der Augenentwicklung von Drosophila melanogaster
Ziel der Arbeit war die Untersuchung des einzigen bei Drosophila vorkommenden Vertreters der HIPK-Familie im Kontext der Augenentwicklung. Die retinale Determination wird bei Drosophila von den Faktoren des Retinalen Determinations Gen Netzwerkes (RDGN) gesteuert. Während einer Interaktionsstudie mit Hilfe der Bimolekularen Fluoreszenzkomplementation (BiFC) konnten die RDGN-Faktoren Twin of Eyeless (Toy) und Eyeless (Ey) als in vivo Interaktionspartner der Hipk bestätigt werden. Außerdem konnten beide Proteine durch in vitro Phosphorylierungsanalysen als Substrat der Hipk identifiziert werden und die vollständige Kartierung von Toy ergab vier Hipk-Phosphorylierungsstellen. Um die Auswirkungen der Hipk-Phosphorylierung von Toy in vivo zu untersuchen, wurden ausgehend von den kartierten Phosphorylierungsstellen transgene phosphorylierungsmutante Fliegenstämme erzeugt und diese zur ektopischen Expression im Rahmen von Fehlexpressionsanalysen eingesetzt. Zur Auffindung weiterer Wechselwirkungen der Hipk mit Faktoren der Augenentwicklung wurden Kreuzungsanalysen bei variierender Hipk-Dosis durchgeführt. Außerdem konnten bei einer Untersuchung regulatorischer Regionen von hipk drei potentiell augenspezifische hipk-Enhancer identifiziert werden. Zur funktionellen Analyse wurde ein entsprechender transgener Fliegenstamm erzeugt (hipkEGT).HIPKs are well conserved in different signaling pathways and developmental processes throughout the animal kingdom. Purpose of this work was the investigation of the only representative of the Hipk family in Drosophila within the context of eye development. Retinal determination in Drosophila is molecularly controlled by the Retinal Determination Gene Network (RDGN). Using a Bimolecular Fluorescence Complementation (BiFC), the two RDGN factors and Paired box protein 6 (PAX6) homologues Twin of eyeless (Toy) and Eyeless (Ey) were confirmed to be in vivo interaction partners of Hipk. In addition, Toy and Ey were set in an enzyme-substrate relationship with Hipk by in vitro phosphorylation assays. Full mapping of Toy revealed four Hipk phosphorylation sites. In order to investigate the effects of Hipk phosphorylation in vivo transgenic phosphorylation mutant fly strains were generated based on the mapped Hipk-phosphorylation sites and used for ectopic expression. In order to detect further interactions of the Hipk with factors of eye development, genetic analyzes were performed at varying Hipk dose. In addition, an analysis of hipk regulatory regions by larval reporter gene expression identified three specific hipk enhancers for expression in developing eye tissue. For the functional analysis a corresponding transgenic fly strain was generated (hipkEGT)
Drosophila Homeodomain-Interacting Protein Kinase (Hipk) Phosphorylates the Homeodomain Proteins Homeobrain, Empty Spiracles, and Muscle Segment Homeobox
The Drosophila homeodomain-interacting protein kinase (Hipk) is the fly representative of the well-conserved group of HIPKs in vertebrates. It was initially found through its characteristic interactions with homeodomain proteins. Hipk is involved in a variety of important developmental processes, such as the development of the eye or the nervous system. In the present study, we set Hipk and the Drosophila homeodomain proteins Homeobrain (Hbn), Empty spiracles (Ems), and Muscle segment homeobox (Msh) in an enzyme-substrate relationship. These homeoproteins are transcription factors that function during Drosophila neurogenesis and are, at least in part, conserved in vertebrates. We reveal a physical interaction between Hipk and the three homeodomain proteins in vivo using bimolecular fluorescence complementation (BiFC). In the course of in vitro phosphorylation analysis and subsequent mutational analysis we mapped several Hipk phosphorylation sites of Hbn, Ems, and Msh. The phosphorylation of Hbn, Ems, and Msh may provide further insight into the function of Hipk during development of the Drosophila nervous system
Vestibular paroxysmia: clinical characteristics and long-term course
In 2016, the Bárány Society defined new diagnostic criteria for the neurovascular compression syndrome of the eighth nerve, called “vestibular paroxysmia” (VP), differentiating between definite (dVP) and probable (pVP) forms. The aim of this study was (1) to describe clinical symptoms and laboratory findings in a well-diagnosed large patient cohort according to those criteria, and (2) to evaluate the long-term course over years in dVP. We identified 146 patients (73 dVP, 73 pVP) from our tertiary dizziness center registry. Data of structured history-taking, clinical neurological, neuro-ophthalmological/-otological examinations as well as MRI imaging were extracted for analyses. Overall, attack frequency ranged between 5 and 30 attacks per day; spinning vertigo was the most frequent type. In two-thirds of patients, attacks occurred spontaneously; in one-quarter, they were triggered by head movements. The majority (approximately 70%) reported no accompanying symptoms; in those with symptoms, mild unilateral cochlear symptoms prevailed. One-third of patients initially showed hyperventilation-induced nystagmus without specific direction, and a deviation of the subjective visual vertical between 3° and 6°. Complete loss of peripheral vestibular function was never evident. dVP and pVP significantly differed concerning the vertigo type, e.g., spinning vertigo was more frequent in dVP. Fortunately, three-quarters of dVP patients remained attack-free during follow-up (mean 4.8 years, standardized questionnaire), more than half of them even without any medication. Patients with ongoing attacks showed significantly higher attack frequency at baseline, but reported persistent frequency reduction. Overall, the long-term prognosis of VP appears favorable, not necessarily requiring ongoing treatment. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00415-022-11151-6
Enhancer analysis of the Drosophila zinc finger transcription factor Earmuff by gene targeting
Background
Many transcription factors are involved in the formation of the brain during the development of Drosophila melanogaster. The transcription factor Earmuff (Erm), a member of the forebrain embryonic zinc finger family (Fezf), is one of these important factors for brain development. One major function of Earmuff is the regulation of proliferation within type II neuroblast lineages in the brain; here, Earmuff is expressed in intermediate neural progenitor cells (INPs) and balances neuronal differentiation versus stem cell maintenance. Erm expression during development is regulated by several enhancers.
Results
In this work we show a functional analysis of erm and some of its enhancers. We generated a new erm mutant allele by gene targeting and reintegrated Gal4 to make an erm enhancer trap strain that could also be used on an erm mutant background. The deletion of three of the previously analysed enhancers showing the most prominent expression patterns of erm by gene targeting resulted in specific temporal and spatial defects in defined brain structures. These defects were already known but here could be assigned to specific enhancer regions.
Conclusion
This analysis is to our knowledge the first systematic analysis of several large enhancer deletions of a Drosophila gene by gene targeting and will enable deeper analysis of erm enhancer functions in the future
The equivocal image of young social enterprises : How self-versus other-oriented values influence external perceptions
Underway spectrophotometry in the Fram Strait (European Arctic Ocean): a highly resolved chlorophyll a data source for complementing satellite ocean color
Satellite remote sensing of chlorophyll a concentration (Chl-a) in the Arctic Ocean
is spatially and temporally limited and needs to be supplemented and validated with
substantial volumes of in situ observations. Here, we evaluated the capability of obtaining
highly resolved in situ surface Chl-a using underway spectrophotometry operated during two
summer cruises in 2015 and 2016 in the Fram Strait. Results showed that Chl-a measured
using high pressure liquid chromatography (HPLC) was well related (R2 = 0.90) to the
collocated particulate absorption line height at 676 nm obtained from the underway
spectrophotometry system. This enabled continuous surface Chl-a estimation along the cruise
tracks. When used to validate Chl-a operational products as well as to assess the Chl-a
algorithms of the aqua moderate resolution imaging spectroradiometer (MODIS-A) and
Sentinel-3 Ocean Land Color Imager (OLCI) Level 2 Chl-a operational products, and from
OLCI Level 2 products processed with Polymer atmospheric correction algorithm (version
4.1), the underway spectrophotometry based Chl-a data sets proved to be a much more
sufficient data source by generating over one order of magnitude more match-ups than those
obtained from discrete water samples. Overall, the band ratio (OCI, OC4) Chl-a operational
products from MODIS-A and OLCI as well as OLCI C2RCC products showed acceptable
results. The OLCI Polymer standard output provided the most reliable Chl-a estimates, and
nearly as good results were obtained from the OCI algorithm with Polymer atmospheric
correction method. This work confirms the great advantage of the underway
spectrophotometry in enlarging in situ Chl-a data sets for the Fram Strait and improving
satellite Chl-a validation and Chl-a algorithm assessment over discrete water sample analysis
in the laboratory
New distances to RAVE stars
Probability density functions are determined from new stellar parameters for
the distance moduli of stars for which the RAdial Velocity Experiment (RAVE)
has obtained spectra with S/N>=10. Single-Gaussian fits to the pdf in distance
modulus suffice for roughly half the stars, with most of the other half having
satisfactory two-Gaussian representations. As expected, early-type stars rarely
require more than one Gaussian. The expectation value of distance is larger
than the distance implied by the expectation of distance modulus; the latter is
itself larger than the distance implied by the expectation value of the
parallax. Our parallaxes of Hipparcos stars agree well with the values measured
by Hipparcos, so the expectation of parallax is the most reliable distance
indicator. The latter are improved by taking extinction into account. The
effective temperature absolute-magnitude diagram of our stars is significantly
improved when these pdfs are used to make the diagram. We use the method of
kinematic corrections devised by Schoenrich, Binney & Asplund to check for
systematic errors for general stars and confirm that the most reliable distance
indicator is the expectation of parallax. For cool dwarfs and low-gravity
giants tends to be larger than the true distance by up to 30 percent. The
most satisfactory distances are for dwarfs hotter than 5500 K. We compare our
distances to stars in 13 open clusters with cluster distances from the
literature and find excellent agreement for the dwarfs and indications that we
are over-estimating distances to giants, especially in young clusters.Comment: 20 pages accepted by MNRAS. Minor changes to the submitted versio
Generation of Mutants from the 57B Region of Drosophila melanogaster
The 57B region of Drosophila melanogaster includes a cluster of the three homeobox genes
orthopedia (otp), Drosophila Retinal homeobox (DRx), and homeobrain (hbn). In an attempt to isolate mu tants for these genes, we performed an EMS mutagenesis and isolated lethal mutants from the 57B
region, among them mutants for otp, DRx, and hbn. With the help of two newly generated deletions
from the 57B region, we mapped additional mutants to specific chromosomal intervals and identi fied several of these mutants from the 57B region molecularly. In addition, we generated mutants
for CG15651 and RIC-3 by gene targeting and mutants for the genes CG9344, CG15649, CG15650,
and ND-B14.7 using the CRISPR/Cas9 system. We determined the lethality period during develop ment for most isolated mutants. In total, we analysed alleles from nine different genes from the 57B
region of Drosophila, which could now be used to further explore the functions of the corresponding
genes in the future
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