Processing and trafficking of clotting factor X in the secretory pathway. Effects of warfarin

Clotting factor X undergoes several post-translational processing steps in the liver before the protein appears in blood as the mature two-chain zymogen. In this study we have followed the factor X precursor through the secretory pathway in rat liver in order to identify the site for proteolytic processing of the precursor into a two-chain form and the site for warfarin inhibition of precursor trafficking within the pathway. Isolated rat liver Golgi apparatus was shown to harbour two single-chains of factor X precursors of 70 and 74 kDa and the heavy (50 kDa) and light chains of factor X. It was demonstrated that the two-chain factor X form was produced from a late processing form of the factor X precursor, which indicated that the site for proteolytic conversion to a two-chain form was in the trans-Golgi compartment. The 70 and 74 kDa single-chain precursors and also the light chain of the two-chain form were shown to contain the factor X propeptide which is normally removed before the coagulation factor appears in blood. The data demonstrate that intra-chain cleavage of a single chain factor X precursor in the trans-Golgi compartment can precede release of the propeptide. Warfarin was shown to affect trafficking of the factor X precursor between the endoplasmic reticulum (ER) and the Golgi apparatus. The data suggest a link between vitamin K-dependent gamma-carboxylation of the precursor and its exit from the ER. Warfarin administration resulted in accumulation of factor X precursors associated with the ER membrane. These precursors appear to be stabilized from intracellular degradation while in the ER. In contrast to the large increase in the factor X precursor concentration in the ER membrane, there was no change in the prothrombin precursor concentration as a result of warfarin action on the liver. However, intracellular turnover of the microsomal prothrombin precursor pool in warfarin-treated rats resulted in a pool of less negatively charged proteins, indicating ongoing protein synthesis but inhibition of gamma-carboxylation. The data are consistent with previous findings [Wallin & Martin (1988) J. Biol. Chem. 263, 9994-10001] suggesting that prothrombin and factor X are processed differently by the vitamin K-dependent carboxylase in the ER membrane.