Manipulating the Caenorhabditis elegans genome using mariner transposons

Abstract

Tc1, one of the founding members of the Tc1/mariner transposon superfamily, was identified in the nematode Caenorhabditis elegans more than 25 years ago. Over the years, Tc1 and other endogenous mariner transposons became valuable tools for mutagenesis and targeted gene inactivation in C. elegans. However, transposition is naturally repressed in the C. elegans germline by an RNAi-like mechanism, necessitating the use of mutant strains in which transposition was globally derepressed, which causes drawbacks such as uncontrolled proliferation of the transposons in the genome and accumulation of background mutations. The more recent mobilization of the Drosophila mariner transposon Mos1 in the C. elegans germline circumvented the problems inherent to endogenous transposons. Mos1 transposition strictly depends on the expression of the Mos transposase, which can be controlled in the germline using inducible promoters. First, Mos1 can be used for insertional mutagenesis. The mobilization of Mos1 copies present on an extrachromosomal array results in the generation of a small number of Mos1 genomic insertions that can be rapidly cloned by inverse PCR. Second, Mos1 insertions can be used for genome engineering. Triggering the excision of a genomic Mos1 insertion causes a chromosomal break, which can be repaired by transgene-instructed gene conversion. This process is used to introduce specific changes in a given gene, such as point mutations, deletions or insertions of a tag, and to create single-copy transgenes.

Target site selection by the mariner-like element, Mos1

“…Moreover, the method used to produce Mos1 insertions in C. elegans was not based on the selected expression of any gene. No bias towards favorable specific integration sites was introduced, reinforcing the hypothesis that rDNA is attractive for Mos1 (Robert and Bessereau 2009). Secondly, analysis of the distribution of the Mos1 insertions over the sequence of the 7-kb unit was also found not to be randomly distributed (P \ 0.05; Fig.…”

The cyclin-dependent kinase inhibitors, cki-1 and cki-2, act in overlapping but distinct pathways to control cell-cycle quiescence duringC. elegansdevelopment

“…Recent data obtained from replacing the p57 locus with p27 indicate that these two related mammalian CKIs perform both specific and shared roles. 29 As technologies emerge that would allow high efficiency homologous recombination in C. elegans , 30 it will be possible to uncover the specific molecular functions that differentiate cki-1 and cki-2 .…”

From genes to function: theC.elegansgenetic toolbox

“…39 To date, the only efficient transposon mutagenesis strategy using a heterologous transposon is based on the mobilization of the Drosophila mauritiana element Mos1 . 40, 41 …”