Abstract. The intranuclear position of a set of genes was analyzed with respect to the territories occupied by the whole chromosomes in which these genes are localized. Genes and their respective chromosome territories were simultaneously visualized in three-dimensionally preserved nuclei applying dual color fluorescence in situ hybridization. Three coding (DMD, MYH7, and HBB) and two noncoding sequences (D1Z2 and an anonymous sequence) were analyzed in four different cell types, including cells where DMD and MYH7 are actively transcribed. Spatial analysis by confocal laser scanning microscopy revealed that the genes are preferentially located in the periphery of chromosome territories. This positioning was independent from the activity of the genes. In contrast, the nonexpressed anonymous fragment was found randomly distributed or localized preferentially in the interior of the corresponding chromosome territory. Furthermore, the distribution of the analyzed genes within the territorial peripheries was found to be highly characteristic for each gene, and, again, independent from its expression. The impact of these findings with regard to the three-dimensional arrangement of the linear DNA string within chromosome territories, as well as with respect to a putative nuclear subcompartment confining gene expression, are discussed.'~EtPuIrTeE intensive efforts to decode the linear strucof complex genomes, up to the present day little is known about the three-dimensional genome organization within the cell nuclei of higher organisms. A territorial organization of interphase chromosomes has already been postulated by early cytologists such as Carl Rabl and Theodor Boveri (for review see Cremer, 1985), but this concept was mainly ignored in the 1960's and 70's (see e.g., Comings, 1980). The territorial organization was first shown indirectly by experiments, in which damaged regions of microirradiated cell nuclei, visualized in the subsequently prepared metaphase chromosomes, were found to be locally clustered (Zorn et al., 1979;Cremer et al., 1982). The chromosome territories were later visualized directly by means of in situ hybridization in interspecies somatic hybrid cells (Manuelidis, 1985;Schardin et al., 1985). Application of chromosome painting protocols subsequently proved that territorial organization of chromosomes occurs also in nonhybrid cells of mammalian (Cremer et al., 1988; Lichter et al., 1988a,b; Please address all correspondence to P. Lichter, Abteilung Organisation komplexer Genome, DKFZ, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany. Pinkel et al., 1988) and plant (Schwarzacher et al., 1989;Leitch et al., 1990) origin. However, the spatial organization of the linear DNA molecule within a chromosome territory remains unclear. Based on the analysis of nuclear distances between DNA segments from the same chromosome, it has been suggested that the spatial position of chromatin segments within a chromosome territory follows a random distribution (van den Engh et al., 1992;Yokota et al., 1995;Sa...

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Deletions a ecting the interval between the RB1 gene and marker D13S25 at band 13q14 are the most frequent genetic abnormalities of B-cell chronic lymphocytic leukemia (B-CLL) and indicate the presence of a novel tumor suppressor gene in this region. In the current study, a high resolution physical map of fragments spanning one megabasepair (Mb) of genomic DNA at the critical 13q14 segment was constructed. To de®ne the minimal region of loss within the RB1 and D13S25 interval, we screened 322 B-CLLs for deletions at either of the two loci. Thirty mantle cell lymphomas (MCLs) were included in the analysis because we observed a 13q14 deletion pattern similar to B-CLL in this disease. The incidence of 13q14 deletions was 51% in B-CLL and 70% in MCL, respectively. No frequent loss of the BRCA2 gene at band 13q12 was found. Detailed deletion mapping at band 13q14 with probes from the RB1 ± D13S25 interval lead to the identi®cation of a critical deletion region 400 kb in size. Within this region two segments were most frequently a ected, one at D13S272 120 kb in size and another 240 kb distal of D13S272 80 kb in size. From these two segments expressed sequences were identi®ed as candidates for the putative 13q14 tumor suppressor gene involved in the pathogenesis of B-CLL and MCL.Keywords: deletion mapping; FISH; exon ampli®cation; DIRVISH The molecular pathogenesis of B-CLL, the most frequent leukemia in Western countries, is not well understood (Rozman and Montserrat, 1995). Although a number of recurrent chromosome aberrations have been identi®ed, no gene to date has been found to be consistently involved (Juliusson et al., 1990;DoÈ hner et al., 1997a).The most common structural chromosome aberrations in B-CLL involve band 13q14 (Juliusson et al., 1990), to which the RB1 tumor suppressor gene has been localized (Friend et al., 1986). Monoallelic RB1 deletion was frequently observed in B-CLL but disruption of both RB1 alleles occurred rarely (Liu et al., 1992(Liu et al., , 1993Stilgenbauer et al., 1993). Based on the observation that the distal marker D13S25 was deleted more frequently than RB1, a novel B-CLL tumor suppressor gene located in vicinity to this marker was postulated (Brown et al., 1993;Liu et al., 1993;Chapman et al., 1994).Subsequent studies aimed at the localization of this gene have been thus far inconclusive. Devilder et al.(1995) localized a minimal deletion segment distal of D13S25. However, comparing deletions of RB1 and D13S25 lead us to the hypothesis that the critical segment is localized proximal of D13S25 (Stilgenbauer et al., 1995). This was supported by Liu et al. (1995) who found loss of D13S319 located less than 680 kb proximal of D13S25 more frequently deleted than other 13q14 markers. Bullrich et al. (1996) recently assembled a YAC contig to order new markers in the region and identi®ed a minimal deletion between D13S25 and the marker 206XF12 located less than 550 kb proximal of D13S25. However, in this study deletions did not appear to cluster in a single critical region and the poten...