Mapping the dynamics of the glucocorticoid receptor within the nuclear landscape

Funding: The work was supported by ANPCyT (PICT 2012-0899 to V.L., PICT 2011-1321 and PICT 2014-0630 to A.P.), UBACyT (20020110100074 to V.L.), NIH (P41-GM103540 and P50-GM076516 to E.G. and P.A.) and the Intramural Research Program of the National Institutes of Health (NIH), the National Cancer Institute (NCI) and the Center for Cancer Research (CCR) (to D.M.P.) grants. M.S., L.B., M.V.D., G.B., A.P. and V.L. are CONICET members. M.S. was supported by IUBMB with a Wood-Whelan research fellowship.The distribution of the transcription machinery among different sub-nuclear domains raises the question on how the architecture of the nucleus modulates the transcriptional response. Here, we used fluorescence fluctuation analyses to quantitatively explore the organization of the glucocorticoid receptor (GR) in the interphase nucleus of living cells. We found that this ligand-activated transcription factor diffuses within the nucleus and dynamically interacts with bodies enriched in the coregulator NCoA-2, DNA-dependent foci and chromatin targets. The distribution of the receptor among the nuclear compartments depends on NCoA-2 and the conformation of the receptor as assessed with synthetic ligands and GR mutants with impaired transcriptional abilities. Our results suggest that the partition of the receptor in different nuclear reservoirs ultimately regulates the concentration of receptor available for the interaction with specific targets, and thus has an impact on transcription regulation.Publisher PDFPeer reviewe

Algorithmic Analysis Techniques for Molecular Imaging

This study addresses image processing techniques for two medical imaging modalities: Positron Emission Tomography (PET) and Magnetic Resonance Imaging (MRI), which can be used in studies of human body functions and anatomy in a non-invasive manner. In PET, the so-called Partial Volume Effect (PVE) is caused by low spatial resolution of the modality. The efficiency of a set of PVE-correction methods is evaluated in the present study. These methods use information about tissue borders which have been acquired with the MRI technique. As another technique, a novel method is proposed for MRI brain image segmen- tation. A standard way of brain MRI is to use spatial prior information in image segmentation. While this works for adults and healthy neonates, the large variations in premature infants preclude its direct application. The proposed technique can be applied to both healthy and non-healthy premature infant brain MR images. Diffusion Weighted Imaging (DWI) is a MRI-based technique that can be used to create images for measuring physiological properties of cells on the structural level. We optimise the scanning parameters of DWI so that the required acquisition time can be reduced while still maintaining good image quality. In the present work, PVE correction methods, and physiological DWI models are evaluated in terms of repeatabilityof the results. This gives in- formation on the reliability of the measures given by the methods. The evaluations are done using physical phantom objects, correlation measure- ments against expert segmentations, computer simulations with realistic noise modelling, and with repeated measurements conducted on real pa- tients. In PET, the applicability and selection of a suitable partial volume correction method was found to depend on the target application. For MRI, the data-driven segmentation offers an alternative when using spatial prior is not feasible. For DWI, the distribution of b-values turns out to be a central factor affecting the time-quality ratio of the DWI acquisition. An optimal b-value distribution was determined. This helps to shorten the imaging time without hampering the diagnostic accuracy.Siirretty Doriast

Bayesian Model Based Tracking with Application to Cell Segmentation and Tracking

The goal of this research is to develop a model-based tracking framework with biomedical imaging applications. This is an interdisciplinary area of research with interests in machine vision, image processing, and biology. This thesis presents methods of image modeling, tracking, and data association applied to problems in multi-cellular image analysis, especially hematopoietic stem cell (HSC) images at the current stage. The focus of this research is on the development of a robust image analysis interface capable of detecting, locating, and tracking individual hematopoietic stem cells (HSCs), which proliferate and differentiate to different blood cell types continuously during their lifetime, and are of substantial interest in gene therapy, cancer, and stem-cell research. Such a system can be potentially employed in the future to track different groups of HSCs extracted from bone marrow and recognize the best candidates based on some biomedical-biological criteria. Selected candidates can further be used for bone marrow transplantation (BMT) which is a medical procedure for the treatment of various incurable diseases such as leukemia, lymphomas, aplastic anemia, immune deficiency disorders, multiple myeloma and some solid tumors. Tracking HSCs over time is a localization-based tracking problem which is one of the most challenging tracking problems to be solved. The proposed cell tracking system consists of three inter-related stages: i) Cell detection/localization, ii) The association of detected cells, iii) Background estimation/subtraction. that will be discussed in detail

The doctoral research abstracts. Vol:12 2017 / Institute of Graduate Studies, UiTM

Foreword: Congratulation to IGS on your continuous efforts to publish the 12th issue of the Doctoral Research Abstracts which highlights research in various disciplines from science and technology, business and administration to social sciences and humanities. This research abstract features the abstracts from 71 PhD doctorates who will receive their scrolls in this 87th UiTM momentous convocation ceremony. To the 71 doctorates, you have most certainly done UiTM proud by journeying through the scholarly world with its endless challenges and obstacles, and by persevering right till the very end. Graduands, your success in achieving the highest academic qualification has demonstrated that you have indeed engineered your destiny well. The action of registering for a PhD program was not by chance but by choice. It was a choice made to realise your self-actualization level that is the highest level in Maslow’s Hierarchy of Needs, while at the same time unleashing your potential in scholarly research. Again, congratulations to all PhD graduates. As you leave the university as alumni we hope a new relationship will be fostered between you and the faculty in soaring UiTM to greater heights. I wish you all the best in your future endeavor. Keep UiTM close to your heart and be our ambassador wherever you go. / Prof Emeritus Dato’ Dr Hassan Said Vice Chancellor Universiti Teknologi MAR

Molecular and behavioral changes associated with adult hippocampus-specific SynGAP1 knockout

The synaptic Ras/Rap-GTPase-activating protein (SynGAP1) plays a unique role in regulating specific downstream intracellular events in response to N-methyl-D-aspartate receptor (NMDAR) activation. Constitutive heterozygous loss of SynGAP1 disrupts NMDAR-mediated physiological and behavioral processes, but the disruptions might be of developmental origin. Therefore, the precise role of SynGAP1 in the adult brain, including its relative functional significance within specific brain regions, remains unexplored. The present study constitutes the first attempt in achieving adult hippocampal-specific SynGAP1 knockout using the Cre/loxP approach. Here, we report that this manipulation led to a significant numerical increase in both small and large GluA1 and NR1 immunoreactive clusters, many of which were non-opposed to presynaptic terminals. In parallel, the observed marked decline in the amplitude of spontaneous excitatory currents (sEPSCs) and inter-event intervals supported the impression that SynGAP1 loss might facilitate the accumulation of extrasynaptic glutamatergic receptors. In addition, SynGAP1-mediated signaling appears to be critical for the proper integration and survival of newborn neurons. The manipulation impaired reversal learning in the probe test of the water maze and induced a delay-dependent impairment in spatial recognition memory. It did not significantly affect anxiety or reference memory acquisition but induced a substantial elevation in spontaneous locomotor activity in the open field test. Thus, the present study demonstrates the functional significance of SynGAP1 signaling in the adult brain by capturing several changes that are dependent on NMDAR and hippocampal integrity

Single-molecule Characterisation of DNA Hybridization via Fluorescence Microscopy and Optoplasmonic Sensing Approaches

The single-molecule study provides an unprecedented deep view into the biological process, unveiling the hidden heterogeneity that is hard to observe in ensemble methods. Recently, various techniques have shown the detection of biomolecules with single-molecule level sensitivity. However, each technique has its unique advantages and drawbacks. Single-molecule techniques can influence the molecular systems that they intend to detect in different ways. For example, fluorescence labels may affect the kinetics and dynamics of biomolecular interactions, while plasmonic-based approaches use local field enhancements that are not uniform across the involved plasmonic nanostructures, both of which can play a significant role in the observed statistics. Before one can combine the information obtained from fluorescence and optoplasmonic techniques, single-molecule experiments must be compared and cross-validated. Here we first compare the detection of DNA hybridization on fluorescence-based imaging technique and optoplasmionic sensors. We investigate the impact of (i) the presence of labels, and (ii) the potential influence of the plasmonic nanoparticle surface. Our measurements reveal that the dissociation rates of hybridized DNA strands are approximately the same for both techniques. Our study establishes the equivalence of both techniques for this DNA molecular test system and can serve as the basis for combining these techniques in other single-molecule studies. With optoplasmonic sensing, our results indicate that one may benefit from the larger binding efficiency of fluorescence imaging while the impact of the label is checked with the optoplasmonic sensing platform. To further combine the two single-molecule characterisation systems, we developed the first optical setup that integrates optoplasmonic and fluorescence-based detection. These sub-platforms provide complementary insights into single-molecule processes. We record the fluorescence and optoplasmonic sensor signals for individual, transient DNA hybridisation events. The hybridisation events are observed in the same sample cell and over a prolonged time (i.e. towards high binding site occupancies). A decrease in the association rate over the measurement duration is reported. Our dual optoplasmonic sensing and imaging platform offers insight into the observed phenomenon, revealing that irreversible hybridisation events accumulate over detected step signals in optoplasmonic sensing. Our results point to novel physicochemical mechanisms that result in the stabilisation of DNA hybridisation on optically-excited plasmonic nanoparticles.Operating Budge