Evolution of the fibrinogen γ′ chain: implications for the binding of factor XIII, thrombin and platelets

Abstract

The c¢ chain of human fibrin(ogen) has received much attention of late because of its apparent regulatory roles, both with regard to the binding of factor (F)XIII by fibrinogen [1,2] and the binding of thrombin by fibrin [3,4]. The c¢ chain is mostly a longer version of the c chain, the result of alternative splicing [5,6], or in this case non-splicing. The result in humans is a c chain 20 residues longer than the more usual spliced form. The extension is markedly anionic, in part because of two sulfated tyrosines [7] that have been implicated in binding to thrombin.Interestingly, mouse fibrin does not bind thrombin with the same avidity as human fibrin, an observation attributed to its shorter c¢ extension. In a recent genetic engineering study, the mouse c¢ sequence was replaced by the human version, after which tight thrombin binding was observed [8]. The experiment raises the question, are humans (or primates) unique in their ability to regulate coagulation by tight binding of c¢ to thrombin? In an effort to answer this question, we conducted a bioinformatics survey in which we examined the genomes of 16 vertebrates in search of putative c¢ sequences. The sine qua non for having two versions of the c chain is an intron near the 3¢ terminus of the coding region. It has long been known that the lamprey c-chain gene lacks the intron [9], as does the puffer fish [10].Publicly accessible data bases were searched for lamprey, puffer fish, zebra fish, frog, chicken, platypus, opossum, dog, cat, mouse, rat, ox, horse, macaque, chimpanzee and human genes. These databases can be accessed through various portals, including http://www.ncbi.nlm.gov, http://genome.wustl.edu/, http://genome.ucsc.edu/cgi-bin/hgGateway/ and http://www. sanger.ac.uk/. Various kinds of BLAST searches [11] were conducted, TBLASTN in particular being used to search translated amino acid sequences against raw DNA data. The highly conserved region composed of c-chain residues 380-410, which is present in both c and c¢ chains, was mostly used as the probe, high scoring hits being examined for introns in the region and, when found, downstream DNA sequences scrutinized for splicing candidates.We confirmed that the intron allowing alternative splicing leading to two forms of fibrinogen c chains does not occur in any fish or frog. It exists in the green lizard gene, but the most probable candidate sequence for a spliced form was a solitary isoleucine residue; direct experiment will be needed to determine whether splicing really occurs. Of more interest, the unspliced form of the lizard gene has an RGD sequence at the same position as occurs in frog and lamprey (Fig. 1A,C).The chicken gene also has the intron, but an almost immediate terminator codon leads to an unspliced form shorter than the spliced alternative (Fig. 1). Suspiciously, an RGD sequence occurs three residues after the terminator codon, followed by another terminator that coincides with lizard and frog (Fig. 1C).The situation in the platypus (a monotreme) is similar. There is an intron at...

Comparative genomic study of arachnid immune systems indicates loss of beta‐1,3‐glucanase‐related proteins and the immune deficiency pathway

“…The extrinsic pathway is activated by trauma, which leads to the activation of the serine protease thrombin. Thrombin cleaves fibrinogen to fibrin, which is the main component of the clot in mammals (Doolittle et al, 2009). Fibrinogen is functionally equivalent to coagulogen in T. tridentatus but not homologous (Theopold et al, 2004).…”

Step-by-Step Evolution of Vertebrate Blood Coagulation

“…Not all species have active γ ′ sequences exhibiting both functions; some species such as the mouse do not bind thrombin at this site at all (Mosesson et al 2009). WGS studies now reveal that the intron involved in this particular γ-chain splicing did not make its appearance until the evolution of reptiles and birds (Doolittle et al 2009). …”

The pleiotropic role of the fibrinogen γ′ chain in hemostasis

“…8 A recent paper on the evolution of the fibrinogen ␥Ј chain shows evidence that the ␥Ј sequence is directly descended from a primitive unspliced version of the ␥ chain gene. 9 Because of the overall negative charge of the ␥Ј carboxy-terminal 20 amino acids, plasma fibrinogen is separable by anion-exchange chromatography into 2 major peaks. 10 Peak 1 fibrinogen is homodimeric with respect to its ␥ chains (␥A/␥A), whereas peak 2 fibrinogen molecules are heterodimeric (␥A/␥Ј).…”