Size Matters: The Influence of Isoform Size on the Intracellular Processing of Apolipoprotein(a)
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High plasma concentrations of Lipoprotein(a) (Lp(a)) have been identified as a risk factor for a variety of atherogenic disorders such as cerebrovascular disease, peripheral vascular disease, and coronary heart disease. Lp(a) consists of a lipoprotein moiety containing apolipoproteinB-100 (apoB-100), as well as apolipoprotein(a) (apo(a)), a unique glycoprotein to which the majority of Lp(a) functions are attributed. Variation in the number of identically repeated kringle IV type 2 (KIV2) motifs of apo(a) forms the molecular basis of Lp(a) isoform size heterogeneity, which is a hallmark of this lipoprotein. There is a general inverse correlation between apo(a) size and plasma Lp(a) concentrations, attributed in part to less efficient secretion of larger apo(a) isoforms from hepatic cells. The present study provides a preliminary investigation into processes involved in apo(a) secretion, with respect to isoform size, to understand this inverse correlation at a molecular level. Pulse-chase experiments were performed in human embryonic kidney (HEK 293) cells and human hepatoma (HepG2) cells, both stably expressing differently-sized recombinant apo(a) isoforms representing the range of apo(a) sizes observed in the population. The folding kinetics for the different apo(a) isoforms were determined by changes in the mobility of the non-reduced radiolabelled species on SDS-PAGE gels. In HEK 293 cells, the rate at which apo(a) is folded correlated well with isoform size. In HepG2 cells, however, folding times were comparable regardless of isoform size. Apo(a) secretion from both cell lines exhibited size-dependency. Preliminary experimentation on endoplasmic reticulum (ER)-resident protein modifications of apo(a) was performed, resulting in the identification of apo(a) interactions with PDI, Erp57, Calnexin, Grp78, Grp94, and EDEM. Preliminary experiments indicate a role for intracellular apo(a) degradation in the amount of apo(a) that is secreted from HepG2 cells, although an isoform size dependency of this degradation process cannot be established with current experimental data. Further experimentation is required to confirm enzyme interactions with differently-sized apo(a) isoforms, to identify other chaperones involved in apo(a) secretion, and to confirm the role of proteasomes in intracellular apo(a) degradation. This may, in turn, provide information regarding the mechanism of how apo(a) secretion from hepatic cells is regulated.