Use of a heterologous monoclonal antibody for cloning and detection of glial fibrillary acidic protein in the bovine ventricular ependyma.
From protozoans to vertebrates, ciliated cells are characterized by well-developed cytoskeletal structures. An outstanding example is the epiplasm, a thick, submembranous skeleton that serves to anchor basal bodies and other cell surface-related organelles in ciliated protozoans. An epiplasm-like cytoskeleton has not yet been observed in metazoan ciliated cells. In a previous study, we reported on MAb E501, a monoclonal antibody raised against epiplasmin-C, the major membrane skeletal protein in the ciliate Tetrahymena pyriformis. It was shown that MAb E501 cross-reacts with glial fibrillary acidic protein (GFAP), the class III intermediate filament protein found in astrocytes and other related glial elements. Here we used a post-embedding immunogold-staining method to localize MAb E501 cross-reactive antigens in ciliated cells from the ventricular ependyma in bovine embryos. When ependymocytes were treated with MAb E501, the ciliated region of the cell cortex was devoid of significant labeling. Instead, a gold particle deposit was evident around the nucleus, with only conventional ependymocytes being immunostained. Similar results were obtained by utilizing a rabbit antiserum against GFAP, revealing glial filaments and indicating an astroglial lineage of conventional bovine ependymocytes. In contrast, secretory ependymocytes of the subcommissural organ (SCO) were not stained by either of the two antibodies. Using MAb E501 as a heterologous probe, we cloned bovine GFAP cDNA. In situ hybridization experiments failed to detect GFAP transcripts in SCO ependymocytes, confirming the abscence of immunoreactivity in these cells.
Bouchard
P
P
Laboratoire de Biologie Comparée des Protistes, UPRES A 6023 CNRS, Université Blaise Pascal, Aubière, France.
Ravet
V
V
Meiniel
R
R
Creveaux
I
I
Meiniel
A
A
Vellet
A
A
Vigues
B
B
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis; D001253:Astrocytes / Q000737:chemistry; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002417:Cattle; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D004805:Ependyma / Q000737:chemistry* / Q000196:embryology / Q000648:ultrastructure; D015723:Gene Library; D005904:Glial Fibrillary Acidic Protein / Q000032:analysis* / Q000235:genetics* / Q000648:ultrastructure; D017403:In Situ Hybridization; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D005904:Glial Fibrillary Acidic Protein; D012333:RNA, Messenger