This study describes comprehensive polling of transcription start and termination sites and analysis of previously unidentified full-length complementary DNAs derived from the mouse genome. We identify the 5' and 3' boundaries of 181,047 transcripts with extensive variation in transcripts arising from alternative promoter usage, splicing, and polyadenylation. There are 16,247 new mouse protein-coding transcripts, including 5154 encoding previously unidentified proteins. Genomic mapping of the transcriptome reveals transcriptional forests, with overlapping transcription on both strands, separated by deserts in which few transcripts are observed. The data provide a comprehensive platform for the comparative analysis of mammalian transcriptional regulation in differentiation and development.

RASSF1A is a potential tumor suppressor gene that undergoes epigenetic inactivation in lung and breast cancers through hypermethylation of its promoter region.

Loss of heterozygosity (LOH) involving several chromosome 3p regions accompanied by chromosome 3p deletions are detected in almost 100% of small (SCLCs) and more than 90% of non-small (NSCLCs) cell lung cancers. In addition, these changes appear early in the pathogenesis of lung cancer and are found as clonal lesions in the smoking damaged respiratory epithelium including histologically normal epithelium as well as in epithelium showing histologic changes of preneoplasia. These 3p genetic alterations lead to the conclusion that the short arm of human chromosome 3 contains several tumor suppressor gene(s) (TSG(s)). Although the first data suggesting that 3p alterations were involved in lung carcinogenesis were published more than 10 years ago, only recently has significant progress been achieved in identifying the candidate TSGs and beginning to demonstrate their functional role in tumor pathogenesis. Some of the striking results of these findings has been the discovery of multiple 3p TSGs and the importance of tumor acquired promoter DNA methylation as an epigenetic mechanism for inactivating the expression of these genes in lung cancer. This progress, combined with the well known role of smoking as an environmental causative risk factor in lung cancer pathogenesis, is leading to the development of new diagnostic and therapeutic strategies which can be translated into the clinic to combat and prevent the lung cancer epidemic. It is clear now that genetic and epigenetic abnormalities of several genes residing in chromosome region 3p are important for the development of lung cancers but it is still obscure how many of them exist and which of the numerous candidate TSGs are the key players in lung cancer pathogenesis. We review herein our current knowledge and describe the most credible candidate genes.

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Clear cell-type renal cell carcinomas (clear RCC) are characterized almost universally by loss of heterozygosity on chromosome 3p, which usually involves any combination of three regions: 3p25-p26 (harboring the VHL gene), 3p12-p14.2 (containing the FHIT gene), and 3p21-p22, implying inactivation of the resident tumorsuppressor genes (TSGs). For the 3p21-p22 region, the affected TSGs remain, at present, unknown. Recently, the RAS association family 1 gene (isoform RASSF1A), located at 3p21.3, has been identified as a candidate lung and breast TSG. In this report, we demonstrate aberrant silencing by hypermethylation of RASSF1A in both VHL-caused clear RCC tumors and clear RCC without VHL inactivation. We found hypermethylation of RASSF1A's GC-rich putative promoter region in most of analyzed samples, including 39 of 43 primary tumors (91%). The promoter was methylated partially or completely in all 18 RCC cell lines analyzed. Methylation of the GC-rich putative RASSF1A promoter region and loss of transcription of the corresponding mRNA were related causally. RASSF1A expression was reactivated after treatment with 5-aza-2 -deoxycytidine. Forced expression of RASSF1A transcripts in KRC͞Y, a renal carcinoma cell line containing a normal and expressed VHL gene, suppressed growth on plastic dishes and anchorage-independent colony formation in soft agar. Mutant RASSF1A had reduced growth suppression activity significantly. These data suggest that RASSF1A is the candidate renal TSG gene for the 3p21.3 region. L oss of heterozygosity (LOH) of chromosome 3p is the most common event in clear cell-type renal cell carcinomas (clear RCC). It involves one or more of the three major commonly deleted regions (1-3) that may be subdivided further by using pioneering stringent criteria for LOH analysis (4-6).The most frequent form of hereditary RCC is the familial VHL cancer syndrome (7). The VHL gene was mapped to 3p25 and was isolated by positional cloning (8). It was inactivated by intragenic and genomic deletions and various kinds of mutations in 100% of analyzed VHL families and in a large portion of sporadic clear RCC (9). Inactivation of VHL by promoter hypermethylation (10) in sporadic clear RCC was noted also and was observed quite frequently (20%). However, in other sporadic clear RCC (reaching in some studies 30-50%) and some non-VHL clear RCC families (11, 12), VHL was not affected (see review in ref. 13).Another commonly deleted region was found at 3p12-p14. Although the overall frequency of the specific LOH is not high, chromosome transfer experiments indicated that the 3p12-p14 region could suppress the tumorigenic properties of some clear RCC cell lines (14), implying the presence of a gene or genes involved in the origin and͞or development of clear RCC. This role may be attributed to the FHIT gene at 3p14.3 (15), and͞or other tumor-suppressor genes (TSGs) residing in 3p12.Multiple studies have identified LOH on 3p21-p22 as the most frequent 3p loss in renal tumor development (4,5,16,17). Recent data indicate that ...

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Using oligonucleotide microarray analysis, THY1, mapping close to a previously defined 11q22-23 nasopharyngeal carcinoma (NPC) critical region was identified as showing consistent downregulated expression in the tumour segregants, as compared to their parental tumour-suppressing microcell hybrids (MCHs). Gene expression and protein analyses show that THY1 was not expressed in the NPC HONE1 recipient cells, tumour segregants, and other NPC cell lines; THY1 was exclusively expressed in the non-tumourigenic MCHs. The mechanism of THY1 gene inactivation in these cell lines was attributed to hypermethylation. Clinical study showed that in 65% of NPC specimens there was either downregulation or loss of THY1 gene expression. Using a tissue microarray and immunohistochemical staining, 44% of the NPC cases showed downregulated expression of THY1 and 9% lost THY1 expression. The frequency of THY1 downregulated expression in lymph node metastatic NPC was 63%, which was significantly higher than in the primary tumour (33%). After transfection of THY1 gene into HONE1 cells, a dramatic reduction of colony formation ability was observed. These findings suggest that THY1 is a good candidate tumour suppressor gene in NPC, which is significantly associated with lymph node metastases.

Chromosome 3p21.3 region is frequently (>90%) deleted in lung and other major human carcinomas. We subdivided 3p21.3 into LUCA and AP20 subregions and discovered frequent homozygous deletions (10 -18%) in both subregions. This finding strongly implies that they harbor multiple tumor suppressor genes involved in the origin and͞or development of major epithelial cancers. In this study, we performed an initial analysis of RBSP3͞HYA22, a candidate tumor suppressor genes located in the AP20 region. Two sequence splice variants of RBSP3͞HYA22 (A and B) were identified, and we provide evidence for their tumor suppressor function. By sequence analysis RBSP3͞HYA22 belongs to a gene family of small C-terminal domain phosphatases that may control the RNA polymerase II transcription machinery. Expression of the gene was drastically (>20-fold) decreased in 11 of 12 analyzed carcinoma cell lines and in three of eight tumor biopsies. We report missense and nonsense mutations in tumors where RBSP3͞HYA22 was expressed, growth suppression with regulated transgenes in culture, suppression of tumor formation in severe combined immunodeficient mice, and dephosphorylation of ppRB by RBSP3͞HYA22, presumably leading to a block of the cell cycle at the G1͞S boundary.