The Ig-ITIM family member PECAM-1 is expressed in vascular and endothelial cells, and its functions include suppression of mitochondria-dependent apoptosis. Previous studies have identified distinct PECAM-1 cytoplasmic domain splice variants at the mRNA, but not protein, level. Several relatively abundant mRNA isoforms lack exon 15 (Δ15) and would theoretically encode a protein with a truncated cytoplasmic domain and a unique C-terminal sequence. Using a novel rabbit polyclonal antibody that specifically recognizes Δ15 PECAM-1, we found that the Δ15 PECAM-1 isoform was expressed in human tissues, including brain, testes and ovary. This isoform was also expressed on the cell surface of human platelets, human umbilical vein endothelial cells (HUVECs) and the Jurkat T-cell leukemia, human erythroleukemia (HEL) and U937 histiocytic lymphoma cell lines. Furthermore, murine platelets and lung lysates demonstrated abundant amounts of exon-15-deficient PECAM-1.
Functional studies revealed that Δ15 PECAM-1 retains both its homophilic binding capacity and its ability to signal by means of its immunoreceptor tyrosine-based inhibitory motif (ITIM) domains. Δ15 PECAM-1 was unable, however, to protect against apoptosis induced by overexpression of Bax or treatment with the chemotherapy agent etoposide. These studies suggest a novel role for the PECAM-1 C-terminus in cytoprotective signaling and highlight a need for further characterization of expression of PECAM-1 isoforms in normal and malignant tissues.
Platelet/endothelial cell adhesion molecule (hereafter referred to as PECAM-1; also known as PECA1 or CD31) is a 130-kDa cell-surface protein that is a member of the immunoglobulin immunoreceptor tyrosine-based inhibitory motif Gentaur Molecular (Ig-ITIM)-containing family (Newman, 1999).
PECAM-1 is expressed on endothelial cells and a variety of hematopoietic cells, including most human CD34+ hematopoietic progenitor cells.
PECAM-1 expression is maintained at relatively high levels on cells of the myeloid and megakaryocytic lineages but is selectively lost during erythroid and B-cell (Jackson et al., 2000) differentiation. Mature human platelets constitutively express 5000 to 10,000 PECAM-1 molecules per cell (Mazurov et al., 1991; Metzelaar et al., 1991; Newman, 1994), whereas monocytes and neutrophils display 50,000 to 100,000 molecules per cell under resting conditions. PECAM-1 is also expressed in certain T-cell subsets (Tanaka et al., 1992; Torimoto et al., 1992; Zehnder et al., 1992) and is a major constituent of the intercellular junction of endothelial cells (Muller et al., 1989; Albelda et al., 1990; Newman et al., 1990; Newman, 1994). Interestingly, PECAM-1 has also been shown to be expressed in human cytotrophoblasts (Zhou et al., 1997a; Zhou et al., 1997b), spermatozoa (Nixon et al., 2005) and brown adipose tissue (Rosso and Lucioni, 2006).
Previous studies in a number of laboratories have shown the existence of alternatively spliced PECAM1 mRNAs in human and murine hematopoietic cells, murine embryos and tissues, and human tissues and endothelial cells (reviewed in Newman and Newman, 2003). Such mRNA species are relatively abundant at certain developmental stages (Baldwin et al., 1994) and in certain tissues (Sheibani et al., 1997; Sheibani et al., 1999; Robson et al., 2001; Wang and Sheibani, 2002; Li et al., 2005) and have the potential to encode PECAM-1 isoforms that differ markedly in their biological properties; however, the lack of isoform-specific reagents has limited the detection of PECAM-1 isoforms at the protein level. Notably, all of the exons encoding the cytoplasmic domain of PECAM-1 are phase 1 exons – that is, they end with a nucleotide that becomes part of the first triplet in the codon encoded by the following exon– with the exception of exon 15, which is a phase 0 exon.
Splicing out of exon 15, therefore, results not only in loss of the amino acids normally encoded by exon 15 but also in a change in the reading frame of downstream exon 16 such that it now encodes a novel C-terminal sequence that ends in the amino acids ENGRLP. Because of the potential for variant PECAM-1 isoforms to confer distinct adhesive and signaling properties to the vascular cells in which they are expressed, we sought to determine whether a PECAM-1 isoform that is missing exon 15 (Δ15) is expressed as a protein in human and murine tissues and, if so, whether it functions differently in its ability to protect cells from apoptosis – a property previously shown to require the PECAM-1 cytoplasmic domain (Bergom et al., 2006). We report herein a potential role for the C-terminal region of PECAM-1 in cytoprotective signaling that might have implications for the expression of different PECAM-1 isoforms during embryogenesis, development and the progression of cancer.
CD31 is a glycoprotein expressed on endothelial cells and in platelets. It is known to be involved in cell signaling and cell adhesion.
Antibody to CD31 is of value in the study of benign and malignant vascular tumors. Staining for CD31 has also been used to measure angiogenesis, which reportedly predicts tumor recurrence.
Host Species: Rabbit
Species Reactivity: Human. Others not tested.
Immunogen: Synthetic peptide corresponding to C-terminus of mouse CD31 protein
Molecular Weight: 100 and 130 kDa
Positive Control: Tonsil, Placenta
Cellular Localization: Endothelial cells in blood vessel of tonsil
- Immunohistochemistry (Formalin/paraffin).
Cell adhesion molecule which is required for leukocyte transendothelial migration (TEM) under most inflammatory conditions. Tyr-679 plays a critical role in TEM and is required for efficient trafficking of PECAM1 to and from the lateral border recycling compartment (LBRC) and is also essential for the LBRC membrane to be targeted around migrating leukocytes. Prevents phagocyte ingestion of closely apposed viable cells by transmitting ‘detachment’ signals, and changes function on apoptosis, promoting tethering of dying cells to phagocytes (the encounter of a viable cell with a phagocyte via the homophilic interaction of PECAM1 on both cell surfaces leads to the viable cell’s active repulsion from the phagocyte.
During apoptosis, the inside-out signaling of PECAM1 is somehow disabled so that the apoptotic cell does not actively reject the phagocyte anymore. The lack of this repulsion signal together with the interaction of the eat-me signals and their respective receptors causes the attachment of the apoptotic cell to the phagocyte, thus triggering the process of engulfment). Modulates BDKRB2 activation (By similarity). Induces susceptibility to atherosclerosis.