Worm Breeder's Gazette 14(1): 89 (October 1, 1995)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.


Laurent Magnenat1, Timothy W. Nilsen2, Heinz Tobler1, Fritz Mueller1

1 Institute of Zoology, University of Fribourg, Perolles, CH-1700 Fribourg, Switzerland
2 School of Medicine, C.W.R.U., Cleveland, Ohio 44106, USA

Telomerases are ribonucleoproteins that act as RNA-dependent DNA
polymerases. Their function is to maintain the protecting telomeric
extremities of the chromosomes after each round of replication and to
heal broken chromosomes by adding telomeric repeats to them.
Strong indirect evidences indicate that telomerase activity is involved
in the process of chromatin diminution in Ascaris suum. This mechanism
involves developmentally-programmed chromosomal cleavage at specific
breakage regions (CBRs), new telomere formation (chromosomal healing)
and degradation of the eliminated chromatin (F. Mueller et al., 1991.
Cell 67: 815-822). Spontaneous chromosomal healing has been described
for the C. elegans X-chromosome mutation me8, which disrupts meiotic
crossing over and segregation (A. M. Villeneuve, 1994. Genetics 136:
887-902). Molecular analysis of the me8 mutation revealed that it
represents a terminal chromosomal truncation healed by de novo addition
of telomeric repeats directly to the site of breakage, most likely by
the action of a telomerase (C. Wicky et al., submitted).

Chromosomal healing in both nematodes share the same molecular
characteristics, arguing for a similar molecular mechanism (S. Jentsch,
pers. comm.; C. Wicky et al., in prep.) Developmentally-programmed
healing in A. suum , however, is highly efficient and occurs in all
presomatic cells, whereas the spontaneous chromosomal healing in C.
elegans takes place randomly in germ cells with low efficiency. Thus,
both nematode species A. suum and C. elegans represent interesting model
systems to study telomerase activity and chromosome healing processes in

The aim of the present project is to investigate an in vitro telomerase
activity in both nematode species. Therefore, the cell-free extracts
were incubated with dNTPs and an oligonucleotide primer designed to
serve as substrate for the telomerase enzyme. The elongated reaction
products were detected by PCR, according to a modified TRAP (telomere
repeat amplification protocol) assay (N.W. Kim et al., 1994. Science
266:2011-2015), using an oligonucleotide complementary to the telomeric
repeats. The PCR products were separated on a denaturing polyacrylamide
gel and visualised by autoradiography.

In our Ascaris 4-cell stage extracts, the telomeric primer was elongated
by up to 30 repeats of 6 bases. Because this activity was sensitive to
heat inactivation, proteinase K or RNase A digestions, and depends on
the time of incubation and the concentration of the extract in the
reaction, we propose that it represents telomerase activity. Our data
thus provide further strong evidence that telomerase  is responsible for
the developmentally-programmed healing process during chromatin
diminution in Ascaris. Currently, different C. elegans extracts are
tested for telomerase in vitro and preliminary results indicate a
similar activity.

Fractionation of the telomerase extracts will be useful to purify and
characterize the protein and RNA components of the telomerase enzyme and
specific co-factors involved in developmentally-programmed or
spontaneous chromosomal healing. The cloning of the corresponding genes
will accelerate the research on telomere function in multicellular