Spatial quantitation of FISH signals in diploid versus aneuploid nuclei

Abstract

Fluorescence in situ hybridization (FISH) is the most widely used molecular technique to visualize chromosomal abnormalities. Here, we describe a novel 3D modeling approach to allow precise shape estimation and localization of FISH signals in the nucleus of human embryonic stem cells (hES) undergoing progressive but defined aneuploidy. The hES cell line WA09 acquires an extra copy of chromosome 12 in culture with increasing passages. Both diploid and aneuploid nuclei were analyzed to quantitate the differences in the localization of centromeric FISH signals for chromosome 12 as it transitions from euploidy to aneuploidy. We employed superquadric modeling primitives coupled with principal component analysis to determine the 3D position of FISH signals within the nucleus. A novel aspect of our modeling approach is that it allows comparison of FISH signals across multiple cells by normalizing the position of the centromeric signals relative to a reference landmark in oriented nuclei. Using this model we present evidence of changes in the relative positioning of centromeres in trisomy-12 cells when compared to diploid cells from the same population. Our analysis also suggests a significant change in the spatial distribution of at least one of the FISH signals in the aneuploid chromosome complements implicating that an overall change in centromere position may occur in trisomy-12 due to the addition of an extra chromosome. These studies underscore the unique utility of our modeling algorithms in quantifying FISH signals in three dimensions.

Chromosomal aneuploidies induced upon Lamin B2 depletion are mislocalized in the interphase nucleus

“…Human X chromosome is altered from a predominantly central to a more peripheral location in X chromosome aneuploidies (XXXXY) (Petrova et al 2007). Interestingly, while two copies of chromosome 21 territory are in closer proximity as compared to the third copy in cells derived from Down’s syndrome patients (Paz et al 2013), spontaneous trisomy for Chr.12 in human embryonic stem cell line (WA09) also shows an altered position of the trisomic chromosome (Shete et al 2014). However, given the overarching function of Lamins in regulating ploidy and genome organization, the specific role of Lamins in the spatial organization of aneuploid CTs is largely unclear.…”

Dendritic cell targeted HIV‐1 gag protein vaccine provides help to a recombinant Newcastle disease virus vectored vaccine including mobilization of protective CD8+ T cells

“…Multiple vaccine combination strategies have been exploited to augment viral vectored vaccines mediated T cell immunity (reviewed in Ref. [36][37][38][39][40][41]), including heterologous vector prime protein boost or vice versa [42][43][44].…”

Cutting the edge

“…Signal overlap of multiple signals in a single nucleus may lead to erroneous results. One approach is to analyze the spatial distribution of signals in 3D images captured by confocal microscopy . In their study, Du and colleagues approached the problem by mathematically modeling of flattened nuclei.…”