Worm Breeder's Gazette 10(1): 48

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.

Activation of Transposable Elements in Mutator Strains

J. Collins, B. Forbes, A. Rushforth and P. Anderson

Figure 1

We have discovered transposable elements other than Tc1 that are 
active in the C.  elegans genome.  Some of this work was reported at 
the meetings last May (Collins, Forbes, and Anderson.  CSH Meetings 
Abstract, 1987).  This article will recap and update that report.  To 
hunt for new transposons, we isolated 60 independent, spontaneous unc-
22 mutations in mutator strain TR679 [mut-2(r459)].  We reported 
previously that mutator strains exhibit elevated germline activity of 
Tc1 (Collins, Saari, and Anderson.  Nature 328:726-728, 1987).  The 
gene structures of these spontaneous unc-22 alleles were analyzed on 
genomic Southern blots.  As expected, most contained Tc1 insertions.  
However, eleven alleles had blot patterns that looked like insertions, 
but did not look like Tc1 (based on differences in size and 
restriction enzyme sites).  Further genetic and molecular analysis of 
these alleles has identified two new transposable elements, Tc3 and 
Tc5, that are active in the mutator genetic background.  Most of what 
we know about Tc3 was presented at the meetings in May.  We isolated 
three independent Tc3 insertions.  All three insertions are 2.5 kb 
long.  Each is inserted at a unique site within unc-22.  The Tc3 
insertions revert at high frequency and the wild type on Southern 
blots.  This indicates that reversion of the Tc3-induced unc-22 
alleles occurs by precise or nearly precise excision of Tc3.  We 
cloned one of the Tc3 insertions.  It does not hybridize with Tc1, Tc2,
Tc4, Tc5, or the foldback sequence (Dreyfus and Emmons.  CSH Meetings 
Abstract, 1987).  The copy number of Tc3 is about 15 in wild type 
strains Bristol, Bergerac, DH424, and TR403; the number and pattern of 
bands is nearly identical in all of these strains.  However, the copy 
number is greater (approx. 20) in TR679.  Furthermore, different 
independent populations derived from TR679 differ from one another in 
number and pattern of Tc3 bands.  These results indicate that Tc3 is 
highly active only in the mut-2 background.  We have sequenced one end 
of the cloned Tc3 element and the corresponding region from the wild 
type unc-22 gene.  Tc3 is inserted into a TA dinucleotide (just like 
Tc1), but the rest of the unc-22 insertion site sequence is unlike the 
Tc1 insertion site consensus.  The terminal eight bases of Tc3 are 
identical to eight of the terminal nine bases of Tc1.  This is 
[See Figure 1]
This suggests that Tc1 and Tc3 are both activated in the mut-2 
background because both elements respond to the same transposase 
function(s).  Our sequencing data for the other end of Tc3 indicate 
that it is an inverted repeat of the end shown above.  We can not say 
for certain whethter or not it is a perfect repeat.
We have recently discovered yet another transposable element, which 
we designate Tc5.  We isolated two independent Tc5 insertions in unc-
22.  Both unstable and revertants of both appear wild type on Southern 
blots.  This demonstrates that reversion occurs by excision of Tc5 
from unc-22, as discussed above for Tc3.  Both insertions are 
approximately 3.2 kb long, and the two sites of insertion in unc-22 
are separated by several kb.  We have cloned both Tc5 insertions.  
They are indistinguishable from one another by restriction enzyme 
analysis.  Tc5 does not hybridize to Tc1 through Tc4 or the foldback 
sequence.  The copy number of Tc5 is about 4-6 in all the wild type 
strains (Bristol, Bergerac, DH424, and TR403) and the blot pattern is 
identical or nearly identical in these strains.  The copy number in 
TR679 and descendants is 10-15.  It looks like Tc5 is activated in the 
TR679 background, like Tc3.  Unlike Tc1, Tc3 and Tc5 do not exhibit 
high frequency somatic excision, since no somatic excision band is 
visible on genomic blots for either of these elements.  We are working 
on cloning three more novel unc-22 insertions -- candidates for Tc6, 
Tc7, and Tc8 ???

Figure 1