En/Spm transposable element system of maize
Enhancer (En)/Suppressor-mutator (Spm)
13 bp perfect terminal inverted repeat
P
ATG
ATG
8.3 kb
tnpD
tnpA
Makes a 3-bp footprints (target site duplication)
En/Spm Transposable elements
cis Determinants for Excision: Approx. 180 bp at the 5’ end and 300 bp of 3’
end represent cis determinants. Contained in these regions are reiterations
of a 12-bp sequence motif that is recognized by TNPA protein with 6 motifs
present at 5’ end and 8 at 3’ end.
Trans-factors: TNPA and TNPD (transposase complex)
TNPA brings the two ends together and
causes DNA bending, TNPD cleaves
TNPA
TNPD
The “suppressor” function of En/Spm Transposable elements
En/Spm have a unique feature: they act as suppressor of gene function
The non-autonomous derivative of En/Spm (dSpm) when inserted into a
gene causes reduced gene expression of that gene instead of knocking it
out. The residual gene activity is due to the spicing of dSpm from pre-mRNA.
However, if trans-factors TNPA is present then gene activity is knocked out
i.e. pre-mRNA is not formed. TNPA binding with dSpm probably causes
steric hindrance for RNA polymerase.
Mutator Transposable element system of maize
Mutator (Mu) trait was first identified by Robertson (1978) as a heritable high
forward-mutation rate exhibited by maize lines. Many of these de novo
mutations exhibited somatic instability, primarily apparent reversion to wildtype. This phenomenon was found to be associated with Mu transposable
elements.
Autonomous MuDR element (4942-bp): 200-bp TIR, create 9-bp target site
duplication.
TIR
P
2.8-kb RNA (823 aa peptide)
1-kb RNA
(207 aa peptide)
Both proteins tightly bind to TIRs
P
TIR
Several Mu elements (subfamilies) exist that contain variable internal sequence.
These are non-autonomous derivatives of MuDR.
Applications:
1. Mu elements are known to transpose to any locus, especially genes, therefore it
is very useful for creating tagged mutations.
2. Mutator’s frequent transposition activity (even to unlinked locus) is reminiscent of
P element system of Drosophila. In Drosophila, P elements have been used as
vectors to increase the efficiency of transgene integration in the injected oocytes.
Therefore, Mu is an attractive system for increasing transformation efficiency of
maize. The strategy would involve introducing gene-of-interest in a Mu element
and then injecting that into maize cells containing MuDR (MuDR based cloning in
impossible because MuDR is unstable in E. coli). However, no report has so far
been published that describes the success for this strategy.