Transmembrane domain of CD44 is required for its detergent insolubility in fibroblasts.
The hyaluronan receptor CD44 is an abundant glycoprotein expressed on a variety of different cell types. In fibroblasts a significant portion of receptor molecules remain in the detergent-insoluble fraction after Triton X-100 extraction. Detergent insolubility of these CD44 molecules has been interpreted to reflect their association with the cytoskeleton. In this study we examined the structural features of CD44 required for its Triton X-100 insolubility in murine fibroblasts. We expressed in L cells the wild-type hematopoietic form of CD44, a mutant CD44 lacking the cytoplasmic domain, and two mutant CD44 molecules with substituted transmembrane domains. Immunofluorescence and cell surface iodination were performed and the detergent extraction profile of the transfected CD44 molecules was determined. No difference in detergent solubility was observed between wild-type and tailless mutant-transfected molecules. However, both CD44 mutants with a heterologous transmembrane domain, derived from either the CD3 zeta chain or CD45, were completely soluble in Triton X-100. These results demonstrate that the transmembrane region but not the cytoplasmic domain of CD44 is required for the detergent-insolubility in these cells. No obvious colocalization of CD44 and actin stress fibers was observed before or after treatment with cytochalasin D, and no change in the detergent extraction profile of wild-type and mutant CD44 molecules was effected by cytochalasin D. In equilibrium density sucrose gradients the Triton-insoluble CD44 component was found in the low density fractions, indicating an association with Triton X-100-insoluble lipids.(ABSTRACT TRUNCATED AT 250 WORDS)
Perschl
A
A
Department of Cancer Biology, Salk Institute, San Diego, California 92186-5800, USA.
Lesley
J
J
English
N
N
Hyman
R
R
Trowbridge
I S
IS
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism*; D018960:Hyaluronan Receptors; D007739:L Cells; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017830:Octoxynol; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016201:Receptors, Lymphocyte Homing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D012995:Solubility; D014162:Transfection
D002352:Carrier Proteins; D018960:Hyaluronan Receptors; D011956:Receptors, Cell Surface; D016201:Receptors, Lymphocyte Homing; D017830:Octoxynol