Genes that can be mutated to unmask hidden antigenic determinants in the cuticle of the nematode Caenorhabditis elegans.

Abstract

Rabbit antisera directed against a mixture of proteins solubilized from the wild-type adult Caenorhabdiis ekgans cuticle were used to isolate mutants, induced by ethyl methanesulfonate treatment, that exhibit alterations in surface antigenicity by immunofluorescence. Genetic mapping and complementation data for four such mutations define two genes, srf-2(I) and srf-3(IV). The nematode surface is antigenic in the infected host. The appearance of specific surface antigens is dynamic, with changes occurring at and between molts (4, 5). However, little is understood of the genetic mechanisms underlying surface antigen expression.The cuticle of the free-living nematode Caenorhabditis elegans has been studied by classical genetics. Mutations affecting cuticle structure include those that alter gross morphology (6-9), some of which are in collagen genes (10-13), and heterochronic mutations that cause expression of adult-specific cuticle features in larvae or vice versa (14-16). In addition, an adult-specific surface antigenic class is missing in certain C. elegans varietal strains; this marker maps to a locus designated srfl(II) (17).Little is known genetically about a characteristic feature of the nematode cuticle-i.e., its layered ultrastructural organization (e.g., ref. 18). We have isolated and analyzed several C. elegans surface antigen mutants that may have implications for understanding the layering pattern and the immune response to nematode parasites. MATERIALS AND METHODSPreparation of Immunogen and Antisera. Rabbit antisera directed against wild-type adult C. elegans cuticle proteins solubilized by SDS and 2-mercaptoethanol (2-ME) were prepared as described (19) Up to 10,000 synchronous F2 larvae were stained for immunofluorescence with L4-adsorbed antiserum and examined; only mutant larvae with alterations in surface antigenicity were expected to stain (17). Immunofluorescent Li to L4 larvae were individually picked. False positive clones were eliminated by rescreening. Only independent mutants were saved.Strains. Wild-type (strain N2) and mutant strains (6) of C. elegans var. Bristol were obtained from the Caenorhabditis elegans Genetics Center, Columbia, MO. sqt-J(scJOJ)II, sqt-J(e1350), sqt-J(sc97), sqt-1(sclOO), sqt-2(sc3)II, sqt-3(sc8)V, sqt-3(sc24), sqt-3(sc63), rol-8(scJ5)II, and rol-8(sc98) were obtained from R. S. Edgar (University of California, Santa Cruz) and were described in ref. 9. lin-JJ(n389)I was described in ref. 20. A strain containing sDf2(IV) (21) was genotypically sDf2/nTJ(IV); +/nTl(V) (22,23).Abbreviations: 2-ME, 2-mercaptoethanol; LI, L3, and L4, first, third, and fourth larval stage, respectively; EMS, ethyl methanesulfonate; PI, protease inhibitors. tTo whom reprint requests should be addressed.

Caenorhabditis elegans Mutants Resistant to Attachment of Yersinia Biofilms

“…We confirmed this phenotype for srf-2(yj262), srf-3(yj10), and srf-5(ct115), each of which is believed to have a surface-specific primary defect (Politz et al 1990;Silverman et al 1997). In contrast, biofilm attachment was normal for the pleiotropic srf mutations srf-4(ct109), srf-8(dv38), and srf-9(dv4) (Link et al 1992).…”

“…srf-1 was defined as an antigenic polymorphism between N2 and wild isolates of C. elegans (Politz et al 1987). srf-2 and srf-3 were initially defined by ectopic binding of a polyclonal antibody that failed to bind wildtype adults (Politz et al 1990). Subsequently, a different phenotype was observed in staining with monoclonal antibodies: these reagents recognized an epitope on the surface of wild-type L1 larvae that is absent from srf-2 and srf-3 animals (Hemmer et al 1991).…”

“…Stage-specific phenotypes of bah-3 and srf-6 mutants: Antibody M37 binds to L1 larvae but not to other stages of wild-type C. elegans under normal growth conditions (Politz et al 1990;Hemmer et al 1991). Alone among the mutants recovered in the Bah screen, bah-1 and bah-2 had the wild-type antibody phenotype ( Table 2).…”

“…The surface composition has been examined genetically using phenotypes of antibody binding (to unidentified epitopes) or lectin binding to whole animals. This identified three genes whose mutants appear to have primary defects in the surface: srf-2, srf-3, and srf-5 (Politz et al 1990;Link et al 1992). Mutations in srf-6 result in stage-specific defects in surface antigen display (Hemmer et al 1991;Grenache et al 1996).…”

Glycosylation Genes Expressed in Seam Cells Determine Complex Surface Properties and Bacterial Adhesion to the Cuticle of Caenorhabditis elegans

“…Both bus-8 and bus-17 affect cuticle integrity, and mutants exhibit increased drug sensitivity, but there are no obvious changes in cuticle integrity or permeability for mutants of the other four genes. Further diversity is revealed by surface lectin and antibody staining (Politz et al 1990;Link et al 1992;Gravato-Nobre et al 2005). …”