Characterization of the activation-associated isoform of CD43 on murine T lymphocytes.

A rat mAb termed 1B11 recognizes a 130-kDa cell surface glycoprotein expressed on T lymphocytes. Transfection studies using the Cd43 gene transfected into murine L cells, and immunoblots using anti-peptide Abs specific for the CD43 polypeptide identified the 1B11 Ag as the 130-kDa isoform of murine CD43. mAb 1B11 fails to recognize the other major CD43 isoform, 115-kDa CD43, either by Western blotting or by FACS analysis, thus differing from the previously characterized anti-CD43 mAb S7 that recognizes only the CD43 115-kDa isoform and not the CD43 130-kDa isoform. CD43 130-kDa recognized by mAb 1B11 is differentially expressed on T lymphocytes. Whereas most CD4-8-, CD4+8+, and CD4-8+ thymocytes express 130-kDa CD43 constitutively, the Ag is expressed by less than 20% of CD4+ T cells in immature and mature populations. On activation, expression of 130-kDa CD43 is up-regulated dramatically on CD4+ T lymphocytes, and to a lesser extent on CD8+ T lymphocytes. In contrast, T cell activation resulted in only minor up-regulation of 115-kDa CD43. CD43 130-kDa contains sialylated O-linked carbohydrate; however, recognition by mAb 1B11 is not dependent on the presence of sialic acid. Interestingly, removal of sialic acid by neuraminidase treatment of 1B11-negative CD4+ T lymphocytes or 1B11-negative EL4 cells confers 1B11 reactivity, suggesting that the 1B11 epitope is masked by sialic acid residues on the CD43 115-kDa isoform. The isoelectric point (pl) of 130-kDa CD43 was determined to be 6.0, which is higher than the pl reported for 115-kDa CD43. Different molecular properties of 115-kDa and 130-kDa CD43 and their differential expression in T cell subsets may indicate specific roles for these CD43 isoforms in T cell ontogeny and/or T cell function.

Structure and neutrophil-activating properties of a novel inflammatory peptide (ENA-78) with homology to interleukin 8.

A new neutrophil-activating peptide, termed ENA-78, was identified in the conditioned media of stimulated human type II epithelial cell line A549. In response to stimulation with either interleukin 1 beta (IL-1 beta) or tumor necrosis factor alpha (TNF-alpha), ENA-78 was produced and secreted concomitantly with IL-8, GRO alpha, and GRO gamma. ENA-78 consists of 78 amino acids [sequence; see text] and has a molecular weight of 8,357. It has four cysteines positioned identically to those of IL-8 and analogues, and thus belongs to the CXC family of peptides. ENA-78 is related to neutrophil-activating peptide 2 (NAP-2) and GRO alpha (sequence identity, 53% and 52%, respectively) and IL-8 (22% identity). Like NAP-2 and GRO alpha, ENA-78 stimulates neutrophils, inducing chemotaxis, a rise in intracellular free calcium and exocytosis. Cross-desensitization experiments indicate that ENA-78 acts through the same type of receptors as IL-8, NAP-2, and GRO alpha.

Purification and characterization of a major phosphatidylserine-binding phosphoprotein from human platelets.

We describe the isolation, lipid-binding properties and partial amino acid sequence of PS-p68, a novel 68 kDa phosphatidylserine-binding protein from human platelets. PS-p68 is an abundant constituent of platelets, accounting for 0.5-0.75% of total cell protein. It was purified from platelet cytosol by affinity chromatography. Amino acid sequence analysis yielded no similarity to identified proteins. In contrast with most known phospholipid-binding proteins, PS-p68 does not bind Ca2+ and does not require Ca2+ for its binding of phosphatidylserine. Phosphatidylserine binding to PS-p68 was inhibited by phosphatidic acid and by alkylphospholipids. PS-p68 was isolated as a major phosphoprotein from 32P-labelled platelets and was found to function as a protein kinase C substrate in vitro. However, treatment of intact platelets with phorbol 12-myristate 13-acetate, thrombin or carbacyclin did not increase PS-p68 phosphorylation. Platelets appear to be the only blood cells containing PS-p68, which was not detected in neutrophils, monocytes and lymphocytes.