BIOSYNTHESIS OF HUMAN FICOLIN, AN ESCHERICHIA COLI-BINDING PROTEIN, BY MONOCYTES - COMPARISON WITH THE SYNTHESIS OF 2 MACROPHAGE-SPECIFIC PROTEINS, C1Q AND THE MANNOSE RECEPTOR

Ficolin is characterized by the presence of both collagen-like and fib rinogen-like sequences, and potentially has a similar overall structur e as the complement protein Clq and the collectins. Previous studies h ave reported the presence of human ficolin mRNA predominantly in perip heral blood leucocytes. In the present study, the cellular origin of h uman ficolin was investigated in further detail. Preliminary studies u sing reverse transcriptase-polymerase chain reaction (RT-PCR) showed t hat ficolin mRNA was synthesized by U937 cells, a human monocyte cell line. This finding suggested that blood monocytes also normally synthe size human ficolin. Peripheral blood monocytes from adult human donors were harvested at serial time-points (0-20 hr) after adhesion to tiss ue culture plates, and total RNA was isolated and assayed for ficolin mRNA by RT-PCR. Ficolin mRNA was highly expressed in monocytes through out the first 20 hr of adhesion. In contrast. Clq and mannose receptor mRNA were not delectable during the first 8 hr of adhesion, but were highly expressed by 20 hr. Cells were harvested at longer time interva ls (1, 2, 4, 6 and 8 days) to determine whether ficolin expression was temporally regulated at later stages of monocyte differentiation. Fic olin mRNA levels decreased sharply from day 1 to day 6. In contrast, t he levels of both Clq and mannose receptor mRNA showed no changing tre nd. These results are consistent with the absence of ficolin expressio n in many macrophage-rich tissues previously reported. The origin of f icolin from monocytes, together with its structural similarity to Clq and the collectins, raises the possibility that ficolin may be another plasma protein capable of binding to surface structures of micro-orga nisms. Escherichia coil was therefore incubated with human serum, and bound proteins, after elution with sugars, were analysed by Western bl otting using an antiserum raised against a synthetic ficolin peptide. The antiserum identified a polypeptide of approximately 42 000 MW, whi ch is similar in size to that of ficolin as predicted from its cDNA-de rived sequence.