Summary
Trocarin D is a prothrombin activator from the Tropidechis carinatus venom. It is a functional and structural homologue to mammalian blood coagulation
factor Xa.Trocarin D is hypothesised to have evolved from its factor X counterpart
(TrFX) through gene duplication and recruitment.The genes of trocarin D and TrFX have
significant sequence identities, except for insertions/deletions in their intron 1
and promoter regions. In trocarin D intron 1 region, there are three insertions and
two deletions. In trocarin D promoter region, there is a novel 264 bp insertion which
has potential cis-elements.This insertion is termed as Venom Recruitment/Switch Element (VERSE) and is hypothesised to account for switching the low-level constitutive expression
of factor X in the liver to the high-level inducible expression of trocarin D in the
venom gland. To understand the role of VERSE in the trocarin D expression,its cis-elements were characterised by luciferase assays in mammalian cell lines as well
as snake venom gland cells. The ability of VERSE to drive luciferase expression is comparable to that of the trocarin D promoter.
The predicted cis-elements are important in promoting expression as their mutagenesis resulted in lower
luciferase expression.VERSE minimal core promoter and three novel cis-elements (two up-regulatory and one suppressor elements) were identified using deletion/site-directed
mutagenesis studies. VERSE is primarily responsible for the increase of trocarin D expression. The insertions/deletions
within trocarin D intron 1 need to be characterised for their role in tissue-specific
and inducible expression of trocarin D.
Keywords
Factor X - promoter - prothrombin activator - recruitment - snake venom
