Alternative lengthening of telomeres (ALT) in cancer cells

Telomeres are specialized chromosomal structures located at the very end of each chromosomal arm consisting of a tandemly repeated DNA sequence. They serve a variety of fundamental biological functions with the most important one as a non-coding DNA buffer for the end replication problem. Due to the inability of DNA polymerase to replicate the very end of a linear chromosome telomeric DNA erodes with each round of cell division. There has been proof to the concept that telomeric attrition serves as a mitotic clock for the cell since critically shortened telomeres trigger mechanisms that initiate cessation of cell growth and induce senescence. One way to circumvent the fate of cell death due to the gradual loss of telomeric DNA is accomplished by expression of telomerase activity. Although most somatic cells lack expression of the enzyme, tumor cells in vivo and in vitro exhibit telomerase activity in a significant frequency. The other way for maintaining telomere homeostasis represents the way of alternative lengthening of telomeres (ALT) characterized by the absence of telomerase and a highly characteristic pattern of telomere length. Presumably ALT is a recombination based mechanism but little is known about the mechanisms that induce or repress ALT and whether it is a defect in the recombination apparatus or a dysfunction of telomere binding proteins. The objective of my studies will be to investigate the ALT pathway in human tumor cell lines. The specific aims pursued for the project will include inhibition of telomerase activity prior to engagement of the ALT pathway, knock out of the telomerase catalytic subunit in a telomerase positive tumor cell line in order to engage ALT and measuring the frequency of recombination events in ALT and telomerase positive cell lines.