Tunicate Lectins as Acute Phase Reactants

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The overall goals of this research program are to characterize fully the molecular and gene structures and binding properties of lectins from selected model invertebrates in order to understand their evolution as recognition molecules and elucidate the molecular mechanisms of their biological functions. During the past three years, with NSF support, four distinct L-fucosyl-binding lectins have been isolated from the plasma of the protochordate, C.picta, and the biochemical properties and binding specificities, in addition to L-fucose, have been characterized. C. picta lectins bind phosphocholine, galactan, and sialic and muramic acids. Furthermore, by the use of monoclonal antibodies and conventional antisera developed against the purified proteins and synthetic peptides, it was shown that those lectins are cross- reactive with mammalian acute phase reactants, in particular with c-reactive protein (CRP), serum amyloid P (SAP) and coagulation factor VIII (F VIII). Partial amino acid sequences have been determined for two of the lectins, and a peptide form one of the lectins (CPL-I) has been characterized that exhibits high homology to human F VIII. A poly(A)+ containing RNA from hemocytes has been isolated, double stranded cDNA synthesized, a library constructed using the expression vector, lambda gt11, and twelve positive recombinant phage clones have been isolated. Goals for the next three years are: 1) to complete the amino acid and nucleotide sequences of both lectins to determine the extent of homology to the acute phase proteins, CRP, SAP, and F VIII; 2) to develop probes required to identify and isolate genomic clones in order to characterize the structure of their genes; and 3) to carry out studies to identify and characterize the interactions of C. picta lectins with their putative natural ligands (exogenous or endogenous), in particular the bacterial community associated with C. picta colonies and the tunic sulphated galactans. Prior research from this laboratory has shown that the protochordates, tunicates, have carbohydrate-binding proteins (lectins) in their blood that resemble certain proteins found in the blood of higher vertebrates, the so-called "acute phase proteins", that are thought to be components of a primitive defense response. The continuation of these molecular-level studies of tunicate lectins will increase our knowledge of the physiology of these interesting marine organisms and expand our understanding of the origins of host defense mechanisms.