273 research outputs found
Investigating PAX6 and SOX2 dynamic interactions at the single molecule level in live cells
The abundance of transcription factor (TF) molecules in the nuclei of
eukaryotic cells are in the range of thousands. However, the functional binding
sites of most TFs lie in the range of hundreds. This suggests that there is a
surplus of the number of molecules for many TFs, relative to their binding sites
at any given time. Nevertheless, precise TF levels are instrumental for normal
development and maintenance, with haploinsufficiency (namely lowering the
dosage of a TF by half) being a hallmark of many TF-related human
developmental disorders. Qualitative methods assessing TF binding such as
chromatin immunoprecipitation, provide static information, from fixed cell
populations and so fail to provide insight into TF dynamic behaviour. Live-cell
imaging methodologies such as Fluorescence Correlation Spectroscopy
(FCS) offer the ability to measure kinetics of binding to chromatin, protein-protein interactions, absolute concentrations of molecules and the underlying
cell-to-cell variability.
SOX2 and PAX6 TFs exhibit haploinsufficiency in humans. Heterozygous point
mutations, deletions or insertions in these genes can lead to a plethora of
abnormal ocular developmental disorders (e.g. coloboma, aniridia,
microphthalmia, anopthalmia). SOX2 encodes a high-mobility group (HMG)
domain-containing TF, essential for maintaining self-renewal of embryonic
stem cells and is expressed in proliferating central nervous system (CNS)
progenitors. PAX6 contains two DNA binding domains; a PAIRED domain (PD)
and a homeodomain (HD). Both DNA binding domains present in PAX6 (PD
and HD) can function either jointly, or separately, to regulate a plethora of
genes implicated in the development and maintenance of the CNS, the eye
and the pancreas. Despite existing genetic and phenotypic evidence, it
remains unclear how PAX6 and SOX2 influence each other at the molecular
level and how sensitive their stoichiometry is during ocular development.
In this thesis I investigated the dynamic interplay between PAX6/SOX2 and
chromatin in live cells, at the molecular level. I compared wild-type protein
function with pathogenic missense variants using advanced fluorescence
microscopy techniques and assessed how these mutations quantitatively and
qualitatively affected molecular behaviour. My results showed that both SOX2
and PAX6 pathogenic missense mutants display differential subnuclear
localisation, as well as altered protein-protein and protein-chromatin
interactions, linking molecular diffusion to pathogenic phenotype in humans.
More importantly, I identified a novel role of SOX2 in stabilising PAX6-
chromatin complexes in live cells, providing further insight into the complex
and dynamic relation of PAX6 and SOX2 in ocular tissue specification,
maintenance and development
The strategic role of middle managers in the formulation and implementation of digital transformation projects
Middle managers play a crucial role in implementing digital strategy. They directly influence the success or failure of an organization's digital transformation. They are the enablers of digital strategy implementation and company transformation and sometimes can cast a sabotaging shadow side when not involved and engaged in the process. However, there are ways to engage them and divert their sabotaging behavior
Treatment controversies in a rare case of a simultaneous branch retinal artery and vein occlusion secondary to hyperhomocysteinemia and hypertension
Simulations of the Poynting--Robertson Cosmic Battery in Resistive Accretion Disks
We describe the results of numerical "2.5--dimensional" MHD simulations of an
initially unmagnetized disk model orbiting a central point--mass and responding
to the continual generation of poloidal magnetic field due to a secular source
that emulates the Poynting--Robertson (PR) drag on electrons in the vicinity of
a luminous stellar or compact accreting object. The fluid in the disk and in
the surrounding hotter atmosphere has finite electrical conductivity and allows
for the magnetic field to diffuse freely out of the areas where it is
generated, while at the same time, the differential rotation of the disk twists
the poloidal field and quickly induces a substantial toroidal--field component.
The secular PR term has dual purpose in these simulations as the source of the
magnetic field and the trigger of a magnetorotational instability (MRI) in the
disk. The MRI is especially mild and does not destroy the disk because a small
amount of resistivity dampens the instability efficiently. In simulations with
moderate resistivities (diffusion timescales up to 16 local dynamical
times) and after 100 orbits, the MRI has managed to transfer outward
substantial amounts of angular momentum and the inner edge of the disk, along
with azimuthal magnetic flux, has flowed toward the central point--mass where a
new, magnetized, nuclear disk has formed. The toroidal field in this nuclear
disk is amplified by differential rotation and it cannot be contained; when it
approaches equipartition, it unwinds vertically and produces episodic jet--like
outflows. The poloidal field in the inner region cannot diffuse back out if the
characteristic diffusion time is of the order of or larger than the dynamical
time; it continues to grow linearly in time undisturbed and without saturation,
as the outer sections of many poloidal loops are being drawn radially outward.Comment: 27 pages, 55 figure
Intrafamilial Phenotype Variability in Two Male Siblings, With X-linked Juvenile Retinoschisis and Dorzolamide Treatment Effect in the Natural History of the Disease
To investigate how genotype is related to phenotype and document correlations of genotype-phenotype with response of topical administration of dorzolamide in siblings affected with X-linked juvenile retinoschisis (XLRS). We performed a retrospective study on two male siblings (four eyes) with XLRS, who were treated with topical installation of dorzolamide. Clinical diagnosis was supported with familial genetic analysis with bi-directional Sanger sequencing of RS1 pathogenic variant. Optical coherence tomography (OCT), fundus fluorescein angiography (FFA), ultrasound scan (U/S) and electroretinogram (ERG) were used in the evaluation. Central macular thickness (CMT) and best corrected visual acuity (BCVA) were recorded monthly for eighteen months. We performed genetic analysis in their family for mutations in the gene that encodes the protein retinoschisin, responsible for retinoschisis (RS1). It was proved that phenotype variability might be related to the same pathogenic variant. While there was an improvement in BCVA and OCT central macular thickness in the patient with the mild form of disease, the visual acuity and the OCT scans of the patient with severe form of disease did not improve. Intrafamilial phenotypic variability between individuals sharing identical pathogenic variant was documented. Both our patients had a pathogenic variant in a hemizygous state at a genomic location in exon 6 of the RS1 gene; Frameshift mutation that is likely to cause protein truncation was identified which is suggested to result in greater clinical severity. Consequently, it was found that response to dorzolamide is correlated to phenotypic severity
BrainNetVis: An Open-Access Tool to Effectively Quantify and Visualize Brain Networks
This paper presents BrainNetVis, a tool which serves brain network modelling
and visualization, by providing both quantitative and qualitative network measures
of brain interconnectivity. It emphasizes the needs that led to the creation of this
tool by presenting similar works in the field and by describing how our tool contributes
to the existing scenery. It also describes the methods used for the calculation
of the graph metrics (global network metrics and vertex metrics), which carry
the brain network information. To make the methods clear and understandable, we
use an exemplar dataset throughout the paper, on which the calculations and the
visualizations are performed. This dataset consists of an alcoholic and a control
group of subjects
Intrafamilial Phenotype Variability in Two Male Siblings, With X-linked Juvenile Retinoschisis and Dorzolamide Treatment Effect in the Natural History of the Disease
To investigate how genotype is related to phenotype and document correlations of genotype-phenotype with response of topical administration of dorzolamide in siblings affected with X-linked juvenile retinoschisis (XLRS). We performed a retrospective study on two male siblings (four eyes) with XLRS, who were treated with topical installation of dorzolamide. Clinical diagnosis was supported with familial genetic analysis with bi-directional Sanger sequencing of RS1 pathogenic variant. Optical coherence tomography (OCT), fundus fluorescein angiography (FFA), ultrasound scan (U/S) and electroretinogram (ERG) were used in the evaluation. Central macular thickness (CMT) and best corrected visual acuity (BCVA) were recorded monthly for eighteen months. We performed genetic analysis in their family for mutations in the gene that encodes the protein retinoschisin, responsible for retinoschisis (RS1). It was proved that phenotype variability might be related to the same pathogenic variant. While there was an improvement in BCVA and OCT central macular thickness in the patient with the mild form of disease, the visual acuity and the OCT scans of the patient with severe form of disease did not improve. Intrafamilial phenotypic variability between individuals sharing identical pathogenic variant was documented. Both our patients had a pathogenic variant in a hemizygous state at a genomic location in exon 6 of the RS1 gene; Frameshift mutation that is likely to cause protein truncation was identified which is suggested to result in greater clinical severity. Consequently, it was found that response to dorzolamide is correlated to phenotypic severity
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