Human keratinocytes express a new CD44 core protein (CD44E) as a heparan-sulfate intrinsic membrane proteoglycan with additional exons.

Abstract

Abstract. We previously identified a 90-kD (GP90), collagen-binding, membrane glycoprotein, termed extracellular matrix receptor 1II (ECMR HI), that is homologous to the lymphocyte homing receptor and CD44 antigen (Gallatin, W. M., E. A. Wayner, P. A. Hoffman, T. St. John, E. C. Butcher, and W. G. Carter. 1989. Proc. Natl. Acad. Sci. USA. 86:4654-4658). CD44 is abundantly expressed in many epithelial tissues, and is localized predominantly to filopodia in cultured keratinocytes. Here we establish CD44 as a polymorphic family of related membrane proteoglycans and glycoproteins possessing extensive diversity in both glycosylation and core protein sequence. Human neonatal foreskin keratinocytes (HFKs) and QG56 lung squamous carcinoma cells express an alternatively spliced form of the CD44 core protein (termed CD44E) that contains an additional 132 amino acids in the carbohydrate attachment region of the extracellular domain. HFKs, HTI080 fibrosarcoma and QG56 cells, as well as many other human cells, contain varying ratios of GP90 and structurally related, higher molecular mass forms of CD44 that express the following characteristics: (a) each form reacted with anti-CD44 (mAbs) P1G12, P3H9, and P3H5. Each of these mAbs recognized a distinct, nonovedapping epitope present on each CD44 form. (b) Differences in mass were due primarily to variation in carbohydrate moieties, including sulfated aspargine-linked glycopeptides (GP), chondroitin sulfate (CS), and heparan sulfate (HS) glycosaminoglycans, as well as O-linked mucin and polylactosamine structure(s). The major polymorphic forms were designated HT1080 GP90 and CS180, QG56 GP230, and HFK HS/CS250, based on dominant carbohydrate moieties and relative mass. (c) The polymorphic forms use CD44 and CD44E core proteins, each containing a unique set of potential attachment sites for O-and N-glycosides and glycosaminoglycans. (d) Immunofluorescence microscopy, differential extraction with Triton-X-114 detergent, and incorporation into liposomes indicated that all the forms were membrane bound glycoconjugates. These results define CD44 as a structurally diverse, but immunologically related, set of intrinsic membrane macromolecules, and suggests that these structurally varied forms might be expected to manifest multiple functions.

Antibodies to CD44 enhance adhesion of normal CD34+ cells and acute myeloblastic but not lymphoblastic leukaemia cells to bone marrow stroma

“…The length of the protein backbone can vary with the insertion of up to 10 variable domains in the membrane proximal region (Screaton et al, 1992). The molecule is also heavily glycosylated and the extent and the nature of the attached carbohydrate creates even greater diversity (Brown et al, 1991;Carter & Wayner, 1988;Jalkanen et al, 1988). CD44H, the form responsible for lymphocyte homing and hyaluronic acid binding, is the smallest, with a molecular weight of 80-100 kD (Goldstein et al, 1989;Stamenkovic et al, 1989).…”

CD44: A Multitude of Isoforms with Diverse Functions

“…The so-called epithelial form of CD44, CD44E, contains exons 8v-9v-10v (#5 in Figure 3: Stamenkovic et al, 1991;Brown et al, 1991;Dougherty et al, 1991). Variations of these exons were detected in the murine colon carcinoma line CMT93 (#1 -#5 in Figure 3); exon 10v can even appear in a smaller version, due to an exon internal splice site (at position 390 of the murine CD44 sequence: #2 in Figure 3; unpublished results).…”

“…In the mouse there is evidence for four polyadenylation sites, which generate RNAs of 1.6; 3.2; 4.0 and 5.2 kb (Wolffe et al, 1990). Between exons 5 and 16 (see Figure 1), near the membrane proximal region of the standard or hematopoietic form of CD44 (CD44S or CD44H), additional sequences of varying lengths have been detected in rats, humans and mice Stamenkovic et al, 1991;Hofmann et al, 1991;Brown et al, 1991;Dougherty et al, 1991;Cooper et al, 1992;Jackson et al, 1992;He et al, 1992;Screaton et al, 1992). While the 5' region of the standard form (exons 1 to 4) and the 3' region (exons 17, 18, and 20; exon 19 encodes the 3' untranslated region of the short cytoplasmic form in humans) are highly conserved among the three species with 82% and 87% identity, the flanking regions (exons 5 and 16) are less well conserved with only 54% and 35% identity, respectively .…”

“…It seemed rather unlikely that all these functions were associated with one and the same molecule, though differences in the posttranslational modification may as well modulate the adhesive properties (Brown et al, 1991). Thus, the description of new extracellular regions led to the assumption that the multitude of functions may be attributed to the various isoforms Brown et al, 1991;Hofmann et al, 1991;He et al, 1992;Matsumura and Tarin, 1992).…”

“…Thus, the description of new extracellular regions led to the assumption that the multitude of functions may be attributed to the various isoforms Brown et al, 1991;Hofmann et al, 1991;He et al, 1992;Matsumura and Tarin, 1992).…”