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Molecular cloning of CD68, a human macrophage marker related to lysosomal glycoproteins
1Imperial Cancer Research Fund, John Radcliffe Hospital, Headington, Oxford, UK.
Insights
Researchers isolated CD68 cDNA clones, revealing it as a type I integral membrane protein. This glycoprotein is highly expressed in monocytes and macrophages, suggesting roles in these immune cells.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- CD68 is a 110-kDa transmembrane glycoprotein highly expressed by human monocytes and tissue macrophages.
- Its precise function remains largely unknown.
- CD68 is a key marker for myeloid cells.
Purpose of the Study:
- To isolate and characterize cDNA clones encoding the CD68 protein.
- To investigate the expression patterns of CD68 in hematopoietic cell lines.
- To elucidate the structural features and potential functional domains of CD68.
Main Methods:
- cDNA library screening using anti-CD68 monoclonal antibodies (MoAbs).
- Transient expression in COS cells.
- Analysis of CD68 transcript levels in various hematopoietic cell lines (U937, KG1, CEM, K562) with and without phorbol myristic acid (PMA) stimulation.
- cDNA sequence analysis to predict protein structure.
Main Results:
- CD68 cDNA clones were successfully isolated.
- CD68 transcripts are constitutively present in U937 cells and upregulated by PMA.
- CD68 transcripts are inducible by PMA in cell lines with low basal expression.
- The predicted CD68 protein is a type I integral membrane protein with a heavily glycosylated extracellular domain, featuring mucin-like and lysosomal/plasma membrane shuttling protein homology domains.
- The extracellular domain contains numerous N-linked and O-linked glycosylation sites.
Conclusions:
- CD68 is a type I integral membrane glycoprotein with a complex extracellular structure.
- Its expression is regulated in myeloid cell lines, suggesting a role in monocyte/macrophage differentiation or function.
- CD68 belongs to a family of hematopoietic mucin-like molecules, potentially involved in cell-cell interactions or adhesion.
Abstract:
CD68 is a 110-Kd transmembrane glycoprotein of unknown function highly expressed by human monocytes and tissue macrophages. We have isolated cDNA clones encoding CD68 from a U937 cDNA library by transient expression in COS cells and panning with the anti-CD68 monoclonal antibodies (MoAbs) Y2/131, Y1/82A, EBM11, and Ki-M6. CD68 transcripts are constitutively present in the promonocyte cell line U937 and are upregulated by phorbol myristic acid (PMA). By contrast, CD68 transcripts are absent or present at very low levels in many hematopoietic lines including KG1, CEM, and K562, but can be induced by exposure to PMA. The cDNA sequence predicts a type I integral membrane protein of 354 residues with a heavily glycosylated extracellular domain of 298 residues containing nine potential N-linked glycosylation sites and numerous potential O-linked glycosylation sites. The extracellular domain consists of two distinct regions separated by an extended proline hinge: a membrane-distal mucin-like domain containing short peptide repeats and consisting of 54% serine and threonine residues; and a membrane proximal domain that has significant sequence homology to a family of lysosomal/plasma membrane shuttling proteins known as the lamp 1 group. CD68 is a member of a growing family of hematopoietic mucin-like molecules, including leukosialin/CD43, the stem cell antigen CD34, and the lymph node high endothelial ligand for L-selectin GlyCAM-1.
