Worm Breeder's Gazette 13(4): 87 (October 1, 1994)

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.

Monoclonal Antibodies Against Caenorhabditis elegans Extracellular Matrix.

Myeongwoo Lee, Annemarie A. Vance, Anthony J. Otsuka, Hou Tak Cheung

Department of Biological Sciences, Illinois State University, Normal, Illinois, 61790-4120.

  Extracellular matrix (ECM) is a complex molecular meshwork of proteins and
carbohydrates that is found between cells. ECM gives tissues mechanical strength,
elasticity, and shape. The major classes of ECM components are collagens,
noncollagenous glycoproteins, and proteoglycans. Available evidence indicates that
ECM play an important role in the development of an organism. Until recently,
human or murine experimental models were used in most of the structural and
functional studies on the ECM components. Because of the complexity and long life
span of higher vertebrates, it is difficult to study the impact of ECM components on
development. We chose C. elegans as an experimental model to study the role of ECM
components in development because of its simple body plan, well-known genetics, and
well-established cell lineages.
  To identify and characterize the components of the C. elegans ECM, a panel of
monoclonal antibodies was prepared against a C. elegans ECM extract by 3.4M NaCI
extraction and followed by 2M urea treatment. Three monoclonal antibodies, LS 22, LS
25, and LS 28 have been characterized by immunofluorescence and immunoblotting.
  In immunofluorescence, LS 22 stained throughout the hermaphrodite gonad, as
well as oocytes from the disrupted gonads. In immunoblotting experiments, LS 22
recognized a series of epitopes at 110, 85, 67, 52, and 45 kD under reducing conditions.
However, LS 22 purified a 67.5 kD species by the immuno-affinity chromatography.
Furthermore, LS 22 crossreacted with the mouse EHS tumor matrix, a basement
membrane analog from Engelbreth-Holm-Swarm sarcoma.
  In situ immunolocalization using a second antibody, LS 25, showed that it
strongly stained body wall muscle structures with a striated pattern resembling
A-bands, the egg-laying muscle near vulva, and the lumen of the intestine. In the
wild-type embryo, LS 25 stained two-cell wide arrays corresponding to the muscle
quadrants of the one-fold embryo. In immunoblotting, LS 25 stained series of protein
bands ranging in molecular weight from 135 kD to 35 kD with the most intense bands at
135, 50, and 40 kD under reducing conditions. Since LS 25 recognized body wall
muscles in immunofluorescence staining, we screened the C. elegans muscle
mutants such as unc-54 ( e190 )and unc-45 ( e286 )with LS 25. LS 25 stained muscle
filaments in these mutants with disorganized filament patterns and localized
intestinal lumen structures with spotted patterns.
  Another monoclonal antibody, designated LS 28, stained hermaphrodite gonad
structures in the immunofluorescence studies. The staining extended throughout the
gonad with no crossreaction to the intestine and other structures such as muscle and
pharynx. Furthermore, LS 28 showed no staining with embryos and fertilized eggs.
The antigenic epitope of LS 28 was detected as a single band of 55 kD in immunoblotting
with C. elegans ECM extracts. This antibody also crossreacts with the mouse EHS
matrix and a purified mouse ECM component, laminin, at 205 kD bands.
  The crossreactivity of the monoclonal antibodies with known C. elegans ECM
components has not been tested yet. It will be of interest to determine the nature of LS
22 and LS 28 crossreactivity with both the worm gonad and mouse ECM. We are in the
process of screening mutants defective in muscle cell attachment and ECM
components with these antibodies.