The cuticle of the nematode Caenorhabditis elegans is a multi-layered, collagenous extracellular "skeleton" that plays crucial roles in both motility and morphogenesis. The ultrastructure of the adult cuticle of the worm has been characterized by transmission electron microscopy (1). The cuticle is circumferentially indented at intervals of 1 mm by annular furrows. The cortical and basal layers sandwich a medial layer that links the other two via struts. Struts are unique in several respects. They are a stage-specific structure found only in adult animals. Unlike the evenly distributed sub-structures found in all other layers of the cuticle, the struts are periodically distributed. They are found in transverse rows along either side of the annulae, spaced about 0.3 mm apart. There are also some struts between these rows. The struts are oriented in a plane perpendicular the other cuticular structures. Freeze-fracture TEM indicates that the struts are anchored to the cortical and basal layers (2). The struts represent a small, contained system that could be useful in studying extracellular macromolecular assembly. The components of this system may be encoded by the bli genes. The six bli genes have not been not well studied. It is known that the Bli phenotype is adult-specific and produces animals with cuticles covered by fluid-filled swellings that seem to result from a separation of the cortical and basal layers. Studies of
bli-1 mutant animals via TEM has shown that the struts are absent from the medial layer, which is instead filled with a granular electron-dense material. This leads us to believe that the Bli genes may encode the set of proteins that are necessary to process and construct the struts. Only one of the six Bli genes,
bli-4, has been studied at a molecular level. This gene encodes
kex2/subtilisin-like pro-protein convertases that may process cuticle collagens, which are likely components of struts (3). Mutations in
bli-1 and
bli-2 show partially penetrant intergenic non-complementation, meaning that although mutations in both genes are recessive, 50% of animals with a genotype
bli-2 +/+
bli-1 are blistered (Kramer, unpublished). This implies that BLI-1 and BLI-2 interact or are involved in the same process. In addition,
bli-1 and
bli-2 phenotypes are can be suppressed by mutations in some cuticle collagens (4). These interactions may indicate the components to which the struts are anchored. Both genes have been mapped to linkage group II, in areas that have been completely sequenced by the genome project. Using genetic and molecular analyses, the
bli-1 gene has been placed in an interval defined by the left end of mnDf90 and the right ends of mnDf57 and mnDf58. This area corresponds to a size of approximately three cosmid lenghts. Using PCR analysis, the
col-39 gene has been mapped to lie within this interval as well. In addition to
col-39, sequence data indicates that this region contains four other collagen genes. The
bli-2 gene has been mapped to an interval defined by the Tc1 polymorphism
stP50 and the
rbl-3 gene. The
bli-2 gene is uncovered by the ccDf5 deficiency, while the
rbl-3 gene lies outside of its right breakpoint. These data limit the region in which
bli-2 lies to roughly three cosmid lengths. Sequencing of this genomic region has indicated that no collagen genes or other genes that encode obivious cuticle proteins lie in this area. We will be presenting work relevant to determining the genomic regions encoding each of these loci, as well as the results of ultrastructural analysis of the cuticles of
bli-1 and
bli-2 mutant animals. (1) Cox, G.N., Staprans, S. and Edgar, R.S. (1981). Dev. Biol. 86: 456-470. (2) Piexoto, C.A. and De Souza, W. (1995). Tissue & Cell: 27: 561-568. (3) Thacker, C., Peters, K., Srayko, M. and Rose, A. (1995). Genes & Dev. 9: 956-971. (4) Cox, G.N., Laufer, J.S., Kusch. M. and Edgar, R.S. (1980). Genetics 95: 317-339.