Analysis of a transposable element in Caenorhabditis elegans ( nematode genome organization / repetitive DNA / inverted repeat / strain variation ) LOUISE

Abstract

A transposable element, designated Tcl, has been characterized in Caenorhabditis elegans. Tcl is 1.7 kilobases long, has an inverted terminal repeat of <100 base pairs, and is repeated as a highly conserved element. The copy number and genomic positions of Tcl are extremely variable among strains, implying that Tcl is mobile. However, progeny of interstrain crosses did not show hybrid dysgenic traits that might be due to Tcl transposition. Transposable elements are discrete genetic units capable of integrating into many sites in the genome (for review, see refs. 1 and 2). Since their discovery (3), several eukaryotic transposable elements have been described. The copia, P, and FB elements in Drosophila and the Ty elements in yeast all have terminal repeats and are members of dispersed repetitive families (4-7). Evidence for their transposition initially came from analyses of their distributions in the genome by the Southern blot technique (8), which showed that the copy numbers and sites of these elements are hypervariable among different strains of the same species and, to a lesser degree, among individuals of the same strain (4, 9-11). Direct evidence for transposition of Ty, copia, and P elements came from analyses of mutations caused by their insertions into nonhomologous genomic sites (6, 12). The P elements are exceptional for their remarkably high frequency of transposition during interbreeding of certain Drosophila strains (13). Transposition of P elements is the most likely basis of P-M hybrid dysgenesis, in which progeny of hybrid crosses exhibit sterility, high rates of mutation, and chromosomal aberrations (14). In this paper, we report the characterization of a transposable element in the nematode Caenorhabditis elegans and compare it with other known eukaryotic transposable elements. Previous studies in this laboratory have shown that the two strains of C. elegans, Bristol and Bergerac, give occasional differences in restriction endonuclease cleavage patterns on Southern blots when probed with randomly selected cloned fragments (15). One such DNA polymorphism was found to be a 1.7-kilobasepair (kb) difference adjacent to the actin gene cluster (16). Here, we demonstrate that the observed polymorphism near the actin genes is due to the presence of the transposable element "Tcl" at that site in the Bergerac strain. Another polymorphism between the Bristol and Bergerac strains has been characterized by Emmons et al. (17) and it also was found to be due to Tci. MATERIALS AND METHODS Nematodes. All C. elegans var. Bristol worms used in this study are descendants of a single Bristol N2 hermaphrodite (18). All laboratory stocks are derived from a culture of Bristol N2 frozen in 1972 in separate vials. Stocks stored frozen since 1972 were obtained from this laboratory and the Medical Research Council laboratories in Cambridge, England, and a stock stored frozen since 1974 was obtained from the Caenorhabditis Genetics Center. Laboratory stocks that were passaged regularly for the past 3-8 years were obtained from D. Baillie, R. Edgar, D. Riddle, and S. Ward. These cultures were generally propagated for a few generations after being thawed, then allowed to survive starvation conditions as dauer larvae for a few weeks. Aliquots of the starved cultures were then transferred to fresh medium, repeating the continuous cycle of growth and starvation. Two laboratory strains of C. elegans var. Bergerac were used. DNA analysis was done on the Bergerac LY strain from this laboratory, obtained from J. Brun in 1977 and originally isolated in France in 1949 (19). For the hybrid dysgenesis tests, we used the Bergerac FR strain, which the Caenorhabditis Genetics Center obtained independently from Brun in 1980. The Bergerac FR strain has male fertility levels and brood sizes comparable with Bristol N2, while the Bergerac LY strain has low male fertility and brood sizes. Bergerac stocks were propagated from single hermaphrodites thawed in recent months and maintained similarly to Bristol stocks. Wild strains of C. elegans obtained from R. Russell included CL2a, GA-1, PA-2, and PaC-1. EPC-4, also called DH424, is a wild isolate that we collected. All wild strains were collected independently from soil samples in or near Pasadena, CA, and found to be fertile with the Bristol N2 strain. DNA Analysis. Procedures for the isolation of C. elegans DNA from first-stage (LI) larvae, restriction endonuclease digestion, gel fractionation of DNA fragments, and hybridization to Southern blots have been described (15). Electron microscopy of renatured single-stranded molecules and heteroduplex molecules was by published procedures (20, 21). All spreads contained linearized pBR325 DNA, which can form stem-loop structures, as a size marker (22). Phage and Plasmid Recombinants. Recombinant phage containing C. elegans DNA were isolated from an EcoRI partial digest library in A Charon 10 or a Sau3A partial digest library in A 1059 (16, 23). EcoRI fragments were subcloned into pBR325 by published methods (15). Isolation of recombinant DNA carrying C. elegans sequences containing actin gene I, actin gene IV, collagen gene col-1, and ribosomal DNA have been described (16, 24, 25). Hybrid Dysgenesis Tests. For each mating, three males from a given stock were transferred to a plate with a single fourthstage (L4) larval hermaphrodite; matings giving 45-50% male progeny were analyzed further. Five to 10 F1 hermaphrodites per mating were transferred individually as larvae to fresh plates to propagate by self-fertilization. F1 sterility was scored by counting both the number of eggs laid and the number of progAbbreviations: bp, base pairs; kb, kilobase pair(s). * To whom reprint requests should be addressed. 3585 The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. Proc. Natl Acad. Sci. USA 80 (1983) eny hatched. Crosses were carried out at 20'C because the Bergerac FR, PA-2, and EPC-4 strains are not viable at or above 250C. Stocks were maintained at 16'C.

Cite this paper

@inproceedings{LIAO2003AnalysisOA, title={Analysis of a transposable element in Caenorhabditis elegans ( nematode genome organization / repetitive DNA / inverted repeat / strain variation ) LOUISE}, author={LOUISE W. LIAO and BRADLEY ROSENZWEIG and David A. Hirsh}, year={2003} }