👤 Vassilis Theodoropoulos

APOC3 is produced mainly by the liver and intestine and approximately half of plasma APOC3 associates with HDL. Though it was believed that APOC3 associates with HDL by simple binding to preexisting particles, recent data support that biogenesis of APOC3-containing HDL (APOC3-HDL) requires Abca1. Moreover, APOC3-HDL contributes to plasma triglyceride homeostasis by preventing APOC3 association with triglyceride-rich lipoproteins. Interestingly, APOC3-HDL also shows positive correlation with the morbidly obese phenotype. However, the roles of APOC3 in HDL functionality and adipose tissue metabolic activity remain unknown. Therefore, here we investigated the direct effects of APOC3 expression on HDL structure and function, as well as white adipose tissue (WAT) and brown adipose tissue (BAT) metabolic activity. C57BL/6 mice were infected with an adenovirus expressing human APOC3 or a recombinant attenuated control adenovirus expressing green fluorescent protein and blood and tissue samples were collected at 5 days postinfection. HDL was then analyzed for its apolipoprotein and lipid composition and particle functionality. Additionally, purified mitochondria from BAT and WAT were analyzed for uncoupling protein 1, cytochrome c (Cytc), and Cytc oxidase subunit 4 protein levels as an indirect measure of their metabolic activity. Serum metabolomic analysis was performed by NMR. Combined, our data show that APOC3 modulates HDL structure and function, while it selectively promotes BAT metabolic activation. Show less

We have examined HP1beta-chromatin interactions in different molecular contexts in vitro and in vivo. Employing purified components we show that HP1beta exhibits selective, stoichiometric, and salt-resistant binding to recombinant histone H3, associating primarily with the helical "histone fold" domain. Furthermore, using "bulk" nucleosomes released by MNase digestion, S-phase extracts, and fragments of peripheral heterochromatin, we demonstrate that HP1beta associates more tightly with destabilized or disrupted nucleosomes (H3/H4 subcomplexes) than with intact particles. Western blotting and mass spectrometry data indicate that HP1beta-selected H3/H4 particles and subparticles possess a complex pattern of posttranslational modifications but are not particularly enriched in me3K9-H3. Consistent with these results, mapping of HP1beta and me3K9-H3 sites in vivo reveals overlapping, yet spatially distinct patterns, while transient transfection assays with synchronized cells show that stable incorporation of HP1beta-gfp into heterochromatin requires passage through the S-phase. The data amassed challenge the dogma that me3K9H3 is necessary and sufficient for HP1 binding and unveil a new mode of HP1-chromatin interactions. Show less

We have previously shown that the mouse heterochromatin protein 1 homologue M31 interacts dynamically with the nuclear envelope. Using quantitative in vitro assays, we now demonstrate that this interaction is potently inhibited by soluble factors present in mitotic and interphase cytosol. As indicated by depletion and order-of-addition experiments, the inhibitory activity co-isolates with a 55-kDa protein, which binds avidly to the nuclear envelope and presumably blocks M31-binding sites. Purification of this protein and microsequencing of tryptic peptides identify it as alpha2/6:beta2-tubulin. Consistent with this observation, bona fide tubulin, isolated from rat brain and maintained in a nonpolymerized state, abolishes binding of M31 to the nuclear envelope and aborts M31-mediated nuclear envelope reassembly in an in vitro system. These observations provide a new example of "moonlighting," a process whereby multimeric proteins switch function when their aggregation state or localization is altered. Show less