This article explores the role of telomerase in the immortalization of human cells, challenging the traditional view that telomerase activation is essential for this process. Researchers found that while telomerase activity is typically associated with cell immortalization, some immortalized cell lines lack detectable telomerase activity. These cells, however, maintain very long and heterogeneous telomeres, suggesting that telomere lengthening can occur through mechanisms other than telomerase activation. The study examined 35 immortalized cell lines, of which 15 were telomerase-negative but had extremely long telomeres. These cells were able to overcome telomere shortening through a novel, unidentified mechanism. The study also investigated the role of telomerase in hybrid cells formed by fusing telomerase-negative and telomerase-positive cells. Some of these hybrids senesced, but a few later regained proliferation and became immortal, either with or without telomerase activity. This suggests that telomerase activation is not the only path to immortalization. The findings support the hypothesis that telomere shortening leads to cellular senescence, and that maintaining or lengthening telomeres is necessary for cell immortalization. The study also analyzed telomere length in various cell lines using pulsed-field gel electrophoresis, revealing that telomerase-negative cells had very long and heterogeneous telomeres. These results indicate that telomere maintenance can occur through mechanisms other than telomerase, and that telomerase-negative cells may have a common mechanism for telomere elongation. The study highlights the complexity of telomere dynamics and the potential for alternative mechanisms of telomere lengthening in immortalized cells. Overall, the findings challenge the notion that telomerase activation is the sole pathway to cell immortalization and suggest that other mechanisms may also be involved.This article explores the role of telomerase in the immortalization of human cells, challenging the traditional view that telomerase activation is essential for this process. Researchers found that while telomerase activity is typically associated with cell immortalization, some immortalized cell lines lack detectable telomerase activity. These cells, however, maintain very long and heterogeneous telomeres, suggesting that telomere lengthening can occur through mechanisms other than telomerase activation. The study examined 35 immortalized cell lines, of which 15 were telomerase-negative but had extremely long telomeres. These cells were able to overcome telomere shortening through a novel, unidentified mechanism. The study also investigated the role of telomerase in hybrid cells formed by fusing telomerase-negative and telomerase-positive cells. Some of these hybrids senesced, but a few later regained proliferation and became immortal, either with or without telomerase activity. This suggests that telomerase activation is not the only path to immortalization. The findings support the hypothesis that telomere shortening leads to cellular senescence, and that maintaining or lengthening telomeres is necessary for cell immortalization. The study also analyzed telomere length in various cell lines using pulsed-field gel electrophoresis, revealing that telomerase-negative cells had very long and heterogeneous telomeres. These results indicate that telomere maintenance can occur through mechanisms other than telomerase, and that telomerase-negative cells may have a common mechanism for telomere elongation. The study highlights the complexity of telomere dynamics and the potential for alternative mechanisms of telomere lengthening in immortalized cells. Overall, the findings challenge the notion that telomerase activation is the sole pathway to cell immortalization and suggest that other mechanisms may also be involved.