Worm Breeder's Gazette 12(5): 30 (February 1, 1993)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
We have identified a new genetic locus, epi-1 ,that affects the structural integrity of basement membrane. Seven alleles were obtained following EMS mutagenesis. These mutations cause a wide spectrum of abnormalities that seem to result from a defect of basement membrane: (1) The basement membrane surrounding the epidermis, gonad, and intestine are disintegrated at various sites or absent entirely. The epidermis and intestine, normally separated by the basement membrane, adhere together at sites of cell contact. The basement membrane of the pharynx, in contrast, appears normal. (2) Migration of neuroblasts and growth of the excretory canals on the epidermis are abnormal (arrested or mispositioned). (3) Myofilaments of skeletal muscle are disorganized while pharyngeal muscles are normal. (4) Uterus and spermathecae fail to epithelialize. The gonadal lineages are normal, however, and these cells form highly differentiated, nonpolarized aggregates. (5) Freed of the gonadal basement membrane, proliferating germline cells spread in the body cavity.
Based on their genetic map position, we had hypothesized epi-1 alleles could be mutations of the laminin A subunit gene, whose partial clone has been isolated and physically mapped (clone GS#Y45 ,John Yochem and Iva Greenwald, personal communication). This clone carries 12 kb of genomic DNA that lacks the 5' part of the gene but contains the 3' end and the 3' flanking region. We have found that the epi-1 mutation can be rescued by microinjection of cosmids, E03C3 or K08C7 ,that cover the laminin A clone GS#Y45 .The genomic DNA in E03C3 is somewhat larger than that in K08C7 . K08C7 carries genomic DNA of 40 kb and its physical map for some restriction enzymes has been constructed. The location of the laminin A gene has been mapped in K08C7 and its sequencing is in progress.
We have screened Bob Barstead's cDNA library with the DNA of clone GS#Y45 as a probe and obtained 32 positive clones. Among the 32 clones, 22 clones are consistent with each other in their restriction maps and should be derived from the laminin A mRNA. Their sequencing is also in progress. Eight clones seem to be cloning artifacts. The identities of the remaining two clones are not known.