Characterization of a class of Mu elements present in normal lines

Normal stocks of maize contain sequences homologous to Mu transposable elements. One class of Mu-homologous sequence has Mu terminal inverted repeats unassociated with the Mu1 internal sequence. This class is present in multiple copies in normal maize lines. Two examples, termed Mu4 and Mu5, were cloned from the maize inbred line B37 based on homology to the Mu1 termini and sequenced. A structural comparison of Mu4 and Mu5 with Mu1 (Figure 1) illustrates the following points.

1. Mu4 and Mu5 are 2.0 and 1.3kb in length, respectively, and both have Mu terminal inverted repeats which are approximately 90% similar to those of Mu1.

2. Mu4 and Mu5 are flanked by short (9bp) direct repeats as are other Mu elements.

3. The terminal inverted repeats of Mu4 and Mu5 are longer than those of Mu1, extending internally from the Mu1-homologous termini by 280 and 130bp, respectively, as shown in Figure 1.

4. Except for the sequence similarity of the Mu termini, Mu1, Mu4, and Mu5 have no sequences in common.

Our sequencing results suggest that Mu4 and Mu5 have the structures of transposable elements, and the apparent 9bp target site duplications suggest that they inserted into their current genomic locations in B37. In addition, Southern blot analysis using a 200bp probe made from the DNA immediately adjacent to the cloned Mu4 element demonstrates that Mu4 elements are not in the same genomic locations in all maize lines. The DNA flanking the cloned Mu4 element hybridizes strongly to a single BglII restriction fragment in all maize lines we have examined, but only in B37 and B73 does the BglII restriction fragment also hybridize to the Mu4 internal probe. Other lines do not have a Mu4 element adjacent to the flanking probe. However, all lines examined have 2 to 4 Mu4-homologous sequences elsewhere in the genome. Similar experiments are in progress for the cloned Mu5 element.

Southern blot analysis reveals multiple restriction fragments homologous to Mu1 termini in all related Zea species we have examined, including Zea mexicana, Zea luxurians and Zea diploperennis. A few of these sequences are also homologous to internal probes for Mu4 and Mu5. In contrast, Tripsacum dactyloides contains no sequences homologous to the Mu1 termini or to the Mu4 and Mu5 internal probes shown in Figure 1. Our hybridization conditions would detect sequences that are at least 86% homologous to the Mu1 termini. These results suggest that Mu4 and Mu5 elements have been in the genome during the evolution of the genus Zea. Experiments are in progress to determine if Mu4 and Mu5 may still contribute to genomic variation in either Mutator or non-Mutator lines of maize.

Mu elements in addition to Mu1 and the closely related Mu1.7 have been shown to transpose in Mutator stocks. These include the Mu3 element (Chen et al., Genetics 116:469, 1987) and an element cloned by Sue Wessler from the wx-mum5 allele isolated by Don Robertson. In addition, we have characterized two other elements, termed Mu6 and Mu7, which were cloned from a Mutator stock based on homology to the Mu1 termini. All of these elements share the Mu terminal inverted repeats, but have no other sequences in common. Internal probes specific for each element also hybridize to restriction fragments in nonMutator lines. We estimate that the 8 characterized elements account for approximately 50-70% of the restriction fragments homologous to Mu1 termini found in non-Mutator maize lines. Thus, additional Mu elements remain to be characterized.

Figure 1. Comparison of Mu4 and Mu5 with the transposable element Mu1. The Mu terminal inverted repeats which are the only regions of homology are indicated by thickened bars. The arrowheads flanking the elements represent the 9 bp direct repeats. Restriction sites indicated are H:Hinfl, T:Tth111-I, N:NcoI, B:BstEII, S:SalI, R:EcoRI. There are additional Hinfl sites in Mu4 which are not shown.

L.E. Talbert, A.J. Kelly and V.L. Chandler

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