A 3-Mb High-Resolution BAC/PAC Contig of 12q22 Encompassing the 830-kb Consensus Minimal Deletion in Male Germ Cell Tumors

Cytogenetic and molecular genetic analyses have shown that the 12q22 region is recurrently deleted in male germ cell tumors (GCTs), suggesting that this site may harbor a tumor suppressor gene (TSG). Previous loss of heterozygosity (LOH) analyses identified a consensus minimal deleted region between the markers D12S377 and D12S296, and a YAC clone contig covering the region was generated. Here, we describe a high-resolution sequence-ready physical map of this contig covering a 3-Mb region. The map comprised of 52 cosmids, 49 PACs, and 168 BACs that were anchored to the previous YAC contig; 99 polymorphic, nonpolymorphic, EST, and gene-based markers are now placed on this map in a unique order. Of these, 61 markers were isolated in the present study, including one that was polymorphic. In addition, we have narrowed the minimal deletion to ∼830 kb between D12S1716 (proximal) and P382A8-AG (distal) by LOH analysis of 108 normal-tumor DNAs from GCT patients using 21 polymorphic STSs. These physical and deletion maps should prove useful for identification of the candidate TSG in GCTs, provide framework to generate complete DNA sequence, and ultimately generate a gene map of this segment of the chromosome 12

Characteristic promoter hypermethylation signatures in male germ cell tumors

BACKGROUND: Human male germ cell tumors (GCTs) arise from undifferentiated primordial germ cells (PGCs), a stage in which extensive methylation reprogramming occurs. GCTs exhibit pluripotentality and are highly sensitive to cisplatin therapy. The molecular basis of germ cell (GC) transformation, differentiation, and exquisite treatment response is poorly understood. RESULTS: To assess the role and mechanism of promoter hypermethylation, we analyzed CpG islands of 21 gene promoters by methylation-specific PCR in seminomatous (SGCT) and nonseminomatous (NSGCT) GCTs. We found 60% of the NSGCTs demonstrating methylation in one or more gene promoters whereas SGCTs showed a near-absence of methylation, therefore identifying distinct methylation patterns in the two major histologies of GCT. DNA repair genes MGMT, RASSF1A, and BRCA1, and a transcriptional repressor gene HIC1, were frequently methylated in the NSGCTs. The promoter hypermethylation was associated with gene silencing in most methylated genes, and reactivation of gene expression occured upon treatment with 5-Aza-2' deoxycytidine in GCT cell lines. CONCLUSIONS: Our results, therefore, suggest a potential role for epigenetic modification of critical tumor suppressor genes in pathways relevant to GC transformation, differentiation, and treatment response

TRIPSACUM AS A POSSIBLE AMPHIDIPLOID OF WILD MAIZE AND MANISURIS

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Alternative Translocation Breakpoint Cluster Region 5' to BCL-6 in B-cell Non-Hodgkin’s Lymphoma

Chromosomal translocations involving band 3q27 with various different partner chromosomes represent a recurrent cytogenetic abnormality in B-cell non-Hodgkin’s lymphoma. In a fraction of these translocations, the chromosomal breakpoint is located within the 5' noncoding region of the BCL-6 proto-oncogene where the BCL-6 major breakpoint region (MBR) maps. As a result of the translocation, BCL-6 expression is deregulated by promoter substitution. However, between 30 and 50% of lymphomas with cytogenetically detectable translocations affecting band 3q27 retain a germ-line configuration at the BCL-6 locus. To identify possible additional breakpoint clusters within 3q27, we cloned a t(3;14)(q27;q32) lymphoma without MBR rearrangement and found a novel breakpoint site located between 245 and 285 kb 5' to BCL-6. Breakpoints within this newly described region, which we called the alternative breakpoint region (ABR), were found to be recurrent in lymphomas carrying t(3q27) chromosomal translocations but devoid of BCL-6 MBR rearrangements. Comparative analysis of multiple lymphomas carrying rearrangements within the ABR showed that the breakpoints cluster within a 20-kb distance. Translocations involving the ABR may juxtapose BCL-6 to distantly acting, heterologous transcriptional regulatory elements which cause deregulation of the proto-oncogene. The identification of BCL-6 ABR provides new tools for the diagnosis of lymphomas carrying aberrations at 3q27 and deregulated BCL-6 genes

Genomic and expression analysis of the 12p11-p12 amplicon using EST arrays identifies two novel amplified and overexpressed genes

We performed parallel array comparative genomic hybridization and array expression analysis of the 12p11-p12 amplicon in human testicular seminomas and an ovarian carcinoma cell line using an expressed sequence tags (ESTs) array spotted with 8254 ESTs. The data were normalized using a robust statistical modeling and the significance inferred from the local SD. We identified two ESTs within the chromosomal amplicon that were amplified and overexpressed in > or =75-100 percent of analyzed tumors with the 12p11-p12 amplicon. These sequences, belonging to coding regions of two novel genes designated here as GCT1 and GCT2, were broadly expressed in a panel of human tissues, including testis and ovary. GCT1 and GCT2 were overexpressed in 92 and 71 percent, respectively, of a panel of seminomas tested. Combined array comparative genomic hybridization and array expression analysis is a valid approach for gene discovery in large chromosomal amplicons

Differential expression patterns of c-REL protein in classic and nodular lymphocyte predominant Hodgkin lymphoma

Classic Hodgkin lymphoma (cHL) is characterized by numerical gains of the short arm of chromosome 2. The high frequency of 2p overrepresentation including REL, particularly in the nodular sclerosis subtype suggests that constitutive activation of nuclear factor kappaB/REL is a hallmark of Reed-Sternberg (RS) cells. The aim of this study was to investigate c-Rel protein expression patterns in cHL and nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) cases by immunohistochemical analysis. A total of 79 cases of HL were analyzed, which included 59 cases of cHL (49 nodular sclerosis; 8 mixed cellularity; 2 lymphocyte-rich) and 20 cases of NLPHL. Positive staining was defined in this study as a reaction seen in the nuclei or nuclei and cytoplasm of RS or lymphocytic and histiocytic (L&H) cells in cHL and NLPHL cases, respectively. The percent positivity of c-REL staining of RS cells in cHL was seen in 51 of 59 cases (86.4%). No significant difference in c-REL expression was seen between nodular sclerosis (42 of 49, 85.7%) and mixed cellularity subtypes (7 of 8 cases, 87.5%; P = 1). In comparison, positive c-REL protein expression in L&H cells was seen in 5 of 20 NLPHL cases (25.0%). Therefore, significantly higher positivity of RS cells in cHL was seen compared with positivity of L&H cells in NLPHL; 86.4% vs. 25.0%; P = 0). Expression of Epstein-Barr virus latent membrane protein was seen in 6 of 30 cases (19.0%; 25 cHL, 5 NLPHL) and EBER1 in 5 of 27 cases (18.5%; 24 cHL, 3 NLPHL). The presence of Epstein-Barr virus did not correlate with c-REL protein expression (P = 1). Our results demonstrate that there is differential c-REL protein expression in cHL in comparison with NLPHL and suggest that c-REL may play a role in the pathogenesis of classic Hodgkin lymphoma