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.
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 summarized below: [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 ???