Detection and distribution patterns of telomerase activity in insects

Abstract

Telomeres of most insects consist of pentanucleotide (TTAGG) n repeats, although the repeats are absent in Diptera and some other insect species, where the telomere regions are perhaps maintained without telomerase. To understand various and unusual telomere formation in insects, we have studied the characteristic features of a putative insect telomerase that has not been previously described. Using a modified telomeric repeat amplification protocol (TRAP), we first detected the telomerase activity in crickets, cockroaches and two Lepidopteran insects. The telomerase from crickets and cockroaches required dATP, dGTP and dTTP but not dCTP as a substrate and sequence analyses of the products of TRAP revealed that the (TTAGG) n repeats are synthesized by telomerase. The cockroach telomerase was detected both in somatic (fat body, muscle and neural tissues) and germ line (testis) cells, suggesting that expression of this enzyme is not regulated in a tissue-specific manner at an adult stage. While we detected high levels of telomerase activity in crickets and cockroaches, we could not detect activity in all tissues and cell cultures of the silkworm, Bombyx mori and in two Drosophila and one Sarcophaga cell lines. This supports the theory that Dipteran insects maintain their telomeres without telomerase.

Telomerase-independent mechanisms of telomere elongation

“…Instead, D. melanogaster telomeres do hybridize to DNA fragments from the non-long-terminal-repeat (non-LTR) retrotransposons, HeT-A and TART [41,42,52,53]. Moreover, telomerase assays failed to detect any activity [54], and the genome lacks a gene resembling telomerase [55]. Clearly, several independent approaches are required to determine with reasonable confidence the absence of a telomerase-based elongation mechanism in any species.…”

“…Their telomeres failed to hybridize to the typical plant-type telomere probe (TTTAGGG) n [56 -58], but at least in Aloe, the telomeres were shown later to hybridize with the vertebrate-type (TTAGGG) n repeats [59]. Moreover, a variety of different oligonucleotide templates must be tested to demonstrate the absence of telomerase activity in an organism, as was done in several insect species [54]. A negative result does not exclude the possibility that an unusual telomeric repeat is synthesized by a telomerase in the species under investigation.…”

“…It is not known whether telomere elongation in these species is indeed independent of telomerase. The detection of telomerase activity in tissue extracts of crickets, cockroaches, potato hornworm and swallowtail butterfly strongly suggests that the pentanucleotide repeats that were observed by fluorescence in situ hybridization (FISH) are telomerase generated [54].…”

Reverse transcriptase-related proteins in telomeres and in certain chromosomal loci of Rhynchosciara (Diptera: Sciaridae)

“…Telomerase activity has not been detected in dipterans so far (Sasaki and Fujiwara 2000) and, except for D. melanogaster, mechanisms for telomere recovery are still unknown in dipterans. A recombination process was found to extend a transgene at the end of one chromosome of Anopheles gambiae terminating with complex repeats (Roth et al 1997) and might thus be an alternative process for telomeric DNA extension.…”

Telomeres in Chironomus thummi are characterized by different subfamilies of complex DNA repeats

“…However, it is possible to find exceptions to this model. Although several attempts to hybridize with short telomeric repeats have been made, all Diptera studied up to now have failed to show a positive signal (Okazaki et al 1993) and lack detectable telomerase (Sasaki and Fujiwara 2000). To terminate the chromosomes Drosophila melanogaster has instead two telomere-specific LINE-like retrotransposons, HeT-A and TART (reviewed in Mason and Biessmann 1995;Pardue and DeBaryshe 1999).…”