c-Myc-Dependent Formation of Robertsonian Translocation Chromosomes in Mouse Cells12

Robertsonian (Rb) translocation chromosomes occur in human and murine cancers and involve the aberrant joining of two acrocentric chromosomes in humans and two telocentric chromosomes in mice. Mechanisms leading to their generation remain elusive, but models for their formation have been proposed. They include breakage of centromeric sequences and their subsequent fusions, centric misdivision, misparing between highly repetitive sequences of p-tel or p-arm repeats, and recombinational joining of centromeres and/or centromeric fusions. Here, we have investigated the role of the oncoprotein c-Myc in the formation of Rb chromosomes in mouse cells harboring exclusively telocentric chromosomes. In mouse plasmacytoma cells with constitutive c-Myc deregulation and in immortalized mouse lymphocytes with conditional c-Myc expression, we show that positional remodeling of centromeres in interphase nuclei coincides with the formation of Rb chromosomes. Furthermore, we demonstrate that c-Myc deregulation in a myc box II-dependent manner is sufficient to induce Rb translocation chromosomes. Because telomeric signals are present at all joined centromeres of Rb chromosomes, we conclude that c-Myc mediates Rb chromosome formation in mouse cells by telomere fusions at centromeric termini of telocentric chromosomes. Our findings are relevant to the understanding of nuclear chromosome remodeling during the initiation of genomic instability and tumorigenesis

Telomere-Centromere-Driven Genomic Instability Contributes to Karyotype Evolution in a Mouse Model of Melanoma1,2

Aneuploidy and chromosomal instability (CIN) are hallmarks of most solid tumors. These alterations may result from inaccurate chromosomal segregation during mitosis, which can occur through several mechanisms including defective telomere metabolism, centrosome amplification, dysfunctional centromeres, and/or defective spindle checkpoint control. In this work, we used an in vitro murine melanoma model that uses a cellular adhesion blockade as a transforming factor to characterize telomeric and centromeric alterations that accompany melanocyte transformation. To study the timing of the occurrence of telomere shortening in this transformation model, we analyzed the profile of telomere length by quantitative fluorescent in situ hybridization and found that telomere length significantly decreased as additional rounds of cell adhesion blockages were performed. Together with it, an increase in telomere-free ends and complex karyotypic aberrations were also found, which include Robertsonian fusions in 100% of metaphases of the metastatic melanoma cells. These findings are in agreement with the idea that telomere length abnormalities seem to be one of the earliest genetic alterations acquired in the multistep process of malignant transformation and that telomere abnormalities result in telomere aggregation, breakage-bridge-fusion cycles, and CIN. Another remarkable feature of this model is the abundance of centromeric instability manifested as centromere fragments and centromeric fusions. Taken together, our results illustrate for this melanoma model CIN with a structural signature of centromere breakage and telomeric loss

Differences in Nuclear DNA Organization Between Lymphocytes, Hodgkin and Reed–Sternberg Cells Revealed by Structured Illumination Microscopy

Advances in light microscopy have enabled the visualization of DNA in the interphase nucleus with more detail than is visible with conventional light microscopy. The nuclear architecture is assumed to be different in cancer cells compared to normal cells. In this paper we have studied, for the first time, the organization of nuclear DNA and that of DNA-free space in control lymphocytes, Hodgkin cells and Reed–Sternberg cells using 3D structured illumination microscopy (SIM). We have observed detail in these SIM images that was not observed in conventional widefield images. We have measured the size distribution of the DNA structure using granulometry and noted a significant, progressive increase in the amount of sub-micron structures from control lymphocytes to Hodgkin cells to Reed–Sternberg cells. The DNA-free space changes as well; “holes” in the DNA distribution start to appear in the malignant cells. We have studied whether these “holes” are nucleoli by staining for upstream binding factor (UBF), a protein associated with the nucleolus. We have found that the relative UBF content progressively and significantly decreases—or is absent—in the DNA-free space when measured as either the Pearson correlation coefficient with the DNA-free space or as the number of “holes” that contain UBF. Similar differences exist within the population of Reed–Sternberg cells between binucleated and multinucleated cells with four or more subnuclei. To our knowledge, this is the first study that investigates the changes of the nuclear DNA structure in any disease with superresolution light microscopy. J. Cell. Biochem. 115: 1441–1448, 2014. © 2014 The Authors. Journal of Cellular Biochemistry published by Wiley Periodicals, Inc. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.ImPhys/Imaging PhysicsApplied Science