Worm Breeder's Gazette 10(3): 72

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Sequence of Tc1 Tightly Linked to a Mutator

Ikue Mori, Don Moerman and Bob Waterston

Figure 1

We have been investigating the molecular basis of a mutator, which 
is required for germline transpositional activity of Tc1.  The 
parallel of the genetic properties of the Tc1 mutator to autonomous 
elements in other eukaryotic element systems led us to look for a Tc1 
co-segregating with the mutator.  As we reported previously, we 
identified and cloned three 1.6 kb Tc1s (#40, #55, and #118) which 
cosegregate with mut-5 II, one of three mutators we identified.  Three 
Tc1's map under mnDf88, and further, Tc1#40 and #55 map to a contig 
near lin-5 (determined by A.  Coulson and J.  Sulston), indicating the 
agreement of positions between cloned Tc1's and mut-5 locus.
We further mapped mut-5 against Tc1#40, #55, and #118, by 
determining the presence or absence of these Tc1's as well as the 
mutator activity in Unc-4 recombinant lines derived from the mut-5 (
Tc1#55, #40, #118) rol-1/dpy-10 -22(st136::Tc1)/+ 
heterozygotes.  Our data suggest that mut-5 is tightly linked to 
Tc1#55 and #40, but probably segregates out from Tc1#118.  Because of 
the proximity of Tc1#55 and #40 (about 200 kb), we did not get 
recombination events between these two Tc1's to allow us to map 
further the position of mut-5.  Phil Carter in Kemphues lab was 
interested in using Tc1#40 and #55 to map zyg-11 on a contig, and 
constructed Dpy-10 and Unc-4 recombinant lines from the similar cross 
described above.  Fortunately, he got one Dpy-10 strain containing 
only Tc1#40 and one Unc-4 strain containing only Tc1#55.  Phil kindly 
sent us his recombinant strains, and our analysis of the mutator 
activities of his strains suggests that the Unc-4 strain with Tc1#55 
maintains mut-5 but the Dpy-10 strain with Tc1#40 does not.  However, 
we have to mention here that the activity of the Unc-4 strain with 
Tc1#55 is somewhat lower than other mut-5 strains (about 10% of other 
mut-5 strains).
When we constructed Unc-4 recombinant lines, we replaced the unc-22(
st136::Tc1) region, which was always maintained in the mut-5 strains, 
by the N2 homolog.  This replacement resulted in reducing the Tc1 copy 
number dramatically from 65 to 45.  The examination of Tc1 patterns 
from these low copy number strains convincingly suggests that three 
Tc1's we cloned are the only Tc1's co-segregating with mut-5.  
Therefore, we propose that if the mutator is an active Tc1 element, it 
should be one of three, and most probably, is Tc1#55.
We sequenced Tc1#55, #40, #118, and #150, an anonymous Tc1 from 
elsewhere, and compared them with two previously sequenced Tc1's, Ce(
Be)Tl (Rosenzweig et al, 1983) and st137::Tc1 (Plasterk, 1987).  The 
six Tc1's are different in 15 positions, but 5 of the differences come 
from st137::Tc1.  When the polypeptide sequences of TcA, a putative 
transposase of Tc1, are compared (Table 1), Tc1#40, #55, and #150 have 
identical TcAs.  The only specific change which distinguishes Tc1#55, 
the most probable candidate of the mutator, from other Tc1's is a G to 
T substitution at position 218, 5' of TcA.  If Tc1#55 is the mutator, 
the sequence analysis provides two possible explanations for its 
mutator activity: one is a single nucleotide substitution, which would 
result in, for example, an elevated level of transcription of TcA in 
germline, and the other is a position effect.
[See Figure 1]

Figure 1