Biology subjects

Shachar, S.

Publications and source records attributed to Shachar, S..

3 recordsLinked to original sources

bioRxiv2018

Effects of Human Sex Chromosome Dosage on Spatial Chromosome Organization

Sex chromosome aneuploidies (SCAs) are common genetic syndromes characterized by the presence of an aberrant number of X and Y chromosomes due to meiotic defects. These conditions impact structure and function of diverse tissues, but the proximal effects of SCA on genome organization are unknown. Here, to determine the consequences of SCAs on global genome organization, we have analyzed multiple architectural features of chromosome organization in a comprehensive set of primary cells from SCA patients with various ratios of X and Y chromosomes by use of imaging-based high-throughput Chromosome Territory Mapping (HiCTMap). We find that X chromosome supernumeracy does not affect the size, volume or nuclear position of the Y chromosome or an autosomal chromosome. In contrast, the active X chromosome undergoes architectural changes as a function of increasing X copy number, as measured by a decrease in size and an increase in circularity, which is indicative of chromatin compaction. With Y chromosome supernumeracy, Y chromosome size is reduced suggesting higher chromatin condensation. The radial positioning of chromosomes is unaffected in SCA karyotypes. Taken together, these observations document changes in genome architecture in response to alterations in sex chromosome numbers and point to trans-effects of dosage compensation on chromosome organization.

cell biology↗

bioRxiv2017

HiCTMap: Detection and Analysis of Chromosome Territory Structure and Position by High-throughput Imaging

The spatial organization of chromosomes in the nuclear space is an extensively studied field that relies on measurements of structural features and 3D positions of chromosomes with high precision and robustness. However, no tools are currently available to image and analyze chromosome territories in a high-throughput format. Here, we have developed High-throughput Chromosome Territory Mapping (HiCTMap), a method for the robust and rapid analysis of 2D and 3D chromosome territory positioning in mammalian cells. HiCTMap is a high-throughput imaging-based chromosome detection method which enables routine analysis of chromosome structure and nuclear position. Using an optimized FISH staining protocol in a 384-well plate format in conjunction with a bespoke automated image analysis workflow, HiCTMap faithfully detects chromosome territories and their position in 2D and 3D in a large population of cells per experimental condition. We apply this novel technique to visualize chromosomes 18, X, and Y in male and female primary human skin fibroblasts, and show accurate detection of the correct number of chromosomes in the respective genotypes. Given the ability to visualize and quantitatively analyze large numbers of nuclei, we use HiCTMap to measure chromosome territory area and volume with high precision and determine the radial position of chromosome territories using either centroid or equidistant-shell analysis. The HiCTMap protocol is also compatible with RNA FISH as demonstrated by simultaneous labeling of X chromosomes and Xist RNA in female cells. We suggest HiCTMap will be a useful tool for routine precision mapping of chromosome territories in a wide range of cell types and tissues.

cell biology↗

bioRxiv2017

Causes and consequences of nuclear gene positioning

The eukaryotic genome is organized in a manner that allows folding of the genetic material in the confined space of the cell nucleus, while at the same time enabling its physiological function. A major principle of spatial genome organization is the non-random position of genomic loci relative to other loci and to nuclear bodies. The mechanisms that determine the spatial position of a locus, and how position affects function, are just beginning to be characterized. Initial results suggest multiple, gene-specific mechanisms and the involvement of a wide range of cellular machineries. In this Commentary, we review recent findings from candidate approaches and unbiased screening methods that provide initial insight into the cellular mechanisms of positioning and their functional consequences. We highlight several specific mechanisms including tethering to the nuclear periphery, passage through replication and histone modifications that contribute to gene positioning in yeast, plants and mammals.

molecular biology↗