Chromosome territories have a highly nonspherical morphology and nonrandom positioning.

Interphase chromosomes are organized into discrete chromosome territories (CTs) that may occupy preferred sub-nuclear positions. While chromosome size and gene density appear to influence positioning, the biophysical mechanisms behind CT localization, especially the relationship between morphology and positioning, remain obscure. One reason for this has been the difficulty in imaging, segmenting, and analyzing structures with variable or imprecise boundaries. This prompted us to develop a novel approach, based on the two-dimensional (2D) wavelet-transform modulus maxima (WTMM) method, adapted to perform objective and rigorous CT segmentation from nuclear background. The wavelet transform acts as a mathematical microscope to characterize spatial image information over a continuous range of size scales. This multiresolution nature, combined with full objectivity of the formalism, makes it more accurate than intensity-based segmentation algorithms and more appropriate than manual intervention. Using the WTMM method in combination with numerical simulation models, we show that CTs have a highly nonspherical 3D morphology, that CT positioning is nonrandom, and favors heterologous CT groupings. We discuss potential relationships between morphology, positioning, chromosomal function, and instability.

The nonhomologous end joining factor Artemis suppresses multi-tissue tumor formation and prevents loss of heterozygosity.

Nonhomologous end joining (NHEJ) is a critical DNA repair pathway, with proposed tumor suppression functions in many tissues. Mutations in the NHEJ factor ARTEMIS cause radiation-sensitive severe combined immunodeficiency in humans and may increase susceptibility to lymphoma in some settings. We now report that deficiency for Artemis (encoded by Dclre1c/Art in mouse) accelerates tumorigenesis in several tissues in a Trp53 heterozygous setting, revealing tumor suppression roles for NHEJ in lymphoid and non-lymphoid cells. We also show that B-lineage lymphomas in these mice undergo loss of Trp53 heterozygosity by allele replacement, but arise by mechanisms distinct from those in Art Trp53 double null mice. These findings demonstrate a general tumor suppression function for NHEJ, and reveal that interplay between NHEJ and Trp53 loss of heterozygosity influences the sequence of multi-hit oncogenesis. We present a model where p53 status at the time of tumor initiation is a key determinant of subsequent oncogenic mechanisms. Because Art deficient mice represent a model for radiation-sensitive severe combined immunodeficiency, our findings suggest that these patients may be at risk for both lymphoid and non-lymphoid cancers.

Inhibition of Th1 responses prevents inflammatory bowel disease in scid mice reconstituted with CD45RBhi CD4+ T cells.

We have described a murine model of IBD that was induced in C.B-17 scid mice by transfer of the CD45RBhi subpopulation of CD4+ T cells from normal BALB/c mice and could be prevented by cotransfer of the CD45RBlo CD4+ T cell subset. Here we have dissected the mechanism of pathogenesis of IBD in this model and used this information for rational immunotherapy of the disease. CD4+ cells from diseased mice displayed a highly polarized Th1 pattern of cytokine synthesis upon polyclonal stimulation in vitro. The administration of anti-IFN gamma MAb to mice soon after T cell transfer prevented development of colitis for up to 12 weeks. Continual neutralization of TNF with anti-TNF MAbs reduced the incidence of severe disease; however, neutralization of TNF during only the first 3-4 weeks had no effect. Severe colitis was completely abrogated in mice treated systemically with rIL-10, but not with rIL-4.

Phenotypically distinct subsets of CD4+ T cells induce or protect from chronic intestinal inflammation in C. B-17 scid mice.

CD4+ T cells in the mouse can be subdivided into two fractions based on the level of expression of the CD45RB determinant. Previous studies have shown that these subsets are functionally distinct. We have further characterized the properties of these subpopulations in vivo by injecting them into C. B-17 scid mice. The animals restored with the CD45RBhighCD4+ T cell population developed a lethal wasting disease with severe mononuclear cell infiltrates into the colon and elevated levels of IFN-gamma mRNA. In contrast, animals restored with the reciprocal CD45RBlow subset or with unfractionated CD4+ T cells did not develop the wasting or colitis. Importantly, the co-transfer of the CD45RBlow population with the CD45RBhigh population prevented the wasting disease and colitis. These data indicate that important regulatory interactions occur between the CD45RBhigh and CD45RBlowCD4+ T cell subsets and that disruption of this mechanism has fatal consequences.