Analysis of Telomerase RNA Gene Expression by In Situ Hybridization

The regulation of telomerase activity is likely to be a complex issue, involving the transcriptional activity of the telomerase RNA component gene, (hTR) and the telomerase catalytic component gene (hTRT), as well as the interaction of telomerase with other telomere-associated proteins (1 -7 ), (see Fig. 1 ). The use of telomerase as a diagnostic marker and target for cancer therapy relies on the development of reliable assays and technologies to detect telomeres and telomerase expression (8 –14 ), ( Table 1 ). Molecular techniques can be roughly broken down into two groups: lysate analysis and in situ analysis (10 ,15 ). With lysate methods, tumor biopsies are homogenized and the spatial relationships between tumor cells are destroyed (Southern blot analysis and polymerase chain reaction [PCR]). This leads to a loss of information on heterogeneity and small subpopulations and presents an averaging of changes. How ever, quantitation can be simpler and more accurate than in situ approaches. In comparison, in situ techniques, such as RNA in situ hybridization (ISH), allow visualization of gene expression in individual cells within their histological context (10 ,11 ,16 ,17 ). This is an important issue in examining the role of telomerase in the development of immortal clones of cancer cells from a telomerase-negative normal tissue. Also, for telomerase and telomerase component genes to be useful biomarkers for disease or as a therapeutic targets, differential expression is required between normal and cancerous tissue (11 ). However, in both normal and cancerous tissue, admixture of cell types may confound interpretation of many assays. The in situ approach described here is ideally placed to solve this problem (10 ,11 ).

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