Background: Elevated serum LDL cholesterol is a major risk factor for atherosclerosis. Mechanisms that regulate LDL homeostasis are not well understood. Results: LRP6 forms a complex with LDLR and other endocytic proteins, and its knockdown or mutation impairs LDLR endocytosis. Conclusion: LRP6 regulates LDLR-dependent LDL uptake. Significance: LRP6 is a potential target for development of novel lipid-lowering drugs.

Vascular smooth muscle cell (VSMC) proliferation is an important event in atherosclerosis and other vasculopathies. PDGF signaling is a key mediator of SMC proliferation, but the mechanisms that control its activity remain unclear. We previously identified a mutation in LDL receptor-related protein 6 (LRP6), LRP6 R611C , that causes early atherosclerosis. Examination of human atherosclerotic coronary arteries showed markedly increased expression of LRP6 and colocalization with PDGF receptor β (PDGFR-β). Further investigation showed that wild-type LRP6 inhibits but LRP6 R611C promotes VSMC proliferation in response to PDGF. We found that wild-type LRP6 forms a complex with PDGFR-β and enhances its lysosomal degradation, functions that are severely impaired in LRP6 R611C . Further, we observed that wild-type and mutant LRP6 regulate cell-cycle activity by triggering differential effects on PDGF-dependent pathways. These findings implicate LRP6 as a critical modulator of PDGF-dependent regulation of cell cycle in smooth muscle and indicate that loss of this function contributes to development of early atherosclerosis in humans.V ascular smooth muscle cell (VSMC) proliferation is a major pathological component of atherosclerosis and many other vasculopathies. Activation of PDGF signaling is a key driver of VSMC proliferation, and both PDGF and PDGF receptor β (PDGFR-β) show increased expression in atherosclerotic lesions (1). Activation of PDGFR-β in response to endothelial injury induces VSMC proliferation and migration (2, 3). Conversely, inhibition of PDGF signaling reduces neointimal smooth muscle cell accumulation (4-8) and diminishes atherosclerotic burden (9, 10). Recent genetic evidence supports a causal role for increased VSMC proliferation in atherosclerosis (11).Upon activation, PDGFR-β is phosphorylated, leading to phosphorylation of recombinant activated factor (RAF) (12), ERK1/2 (13), and JAK1/STAT1 (14), which then activate nuclear transcription factors leading to increased expression of cell-cycle regulators including cyclin D1 (15). The detailed mechanisms regulating expression and activation of PDGF and PDGFR-β are incompletely understood.We recently identified a mutation in LDL receptor-related protein 6 (LRP6), LRP6 R611C , that underlies a Mendelian form of early atherosclerosis and metabolic syndrome. Mutation carriers have diffuse coronary artery and cerebrovascular disease and most often die of cardiovascular disease before age 50 y (16); they also show rapid disease progression after percutaneous coronary artery intervention. Although the genetic link between the identified LRP6 mutation and premature atherosclerosis is clear, the mechanisms by which the mutation imparts its effects have not yet been defined.LRP6 is a member of the LDL receptor family and is known for its function as a membrane coreceptor for canonical WinglessInt (Wnt) signaling. This protein is expressed abundantly in smooth muscle cells and has been shown to play a role in cell-cycle activity in response to Wnt stimulati...

The biochemistry of early stages of hematopoietic differentiation is difficult to study because only relatively small numbers of precursor cells are available. The murine EML cell line is a multipotential cell line that can be used to model some of these steps. We found that the lineage ؊ EML precursor cells can be separated into two populations based on cell surface markers including CD34. Both populations contain similar levels of stem cell factor (SCF) receptor (c-Kit) but only the CD34 ؉ population shows a growth response when treated with SCF. Conversely, the CD34 ؊ population will grow in the presence of the cytokine IL-3. The human ␤-globin locus control region hypersensitive site 2 plays different roles on ␤-globin transcription in the CD34 ؉ and CD34 ؊ populations. The two populations are present in about equal amounts in culture, and the CD34 ؉ population rapidly regenerates the mixed population when grown in the presence of SCF. We suggest that this system may mimic a normal developmental transition in hematopoiesis.c-Kit ͉ CD34 ͉ EML cells ͉ hypersensitive site 2

Mature B cells, upon activation, progressively differentiate through centroblasts into centrocytes and finally to plasmacytes that express large amounts of selected immunoglobulins. A significant part of this maturation is thought to involve induction of the unfolded protein response (UPR). We have compared gene expression in normal germinal center centroblasts, centrocytes, lymphoblastoid cells undergoing induced UPR, and the CCL155 plasmacytoma cell line. In the centroblast to centrocyte transition there is a change in the expression of a relatively small number of genes. These include a limited subset of the genes upregulated by a fully activated UPR as well as a small number of other transcription factors, some disulphide isomerases, and other genes. This is consistent with a model in which this transition is mediated by changes in the levels of expression of transcription factor B-lymphocyte-induced maturation protein 1 (Blimp1) (PRDM1), BACH2, X-box binding protein 1 (XBP1), interferon regulatory factor 4 (IRF4), and possibly vitamin D receptor (VDR) expression, together with post-transcriptional changes and a limited induction of aspects of the UPR.

Erythroid myeloid lymphoid (EML) cells are an established multipotent hematopoietic precursor cell line that can be maintained in medium including stem cell factor (SCF). EML cultures contain a heterogeneous mixture of cells, including a lineage-negative, CD34 + subset of cells that propagate rapidly in SCF and can clonally regenerate the mixed population. A second major subset of EML cells consists of lineage-negative. CD34 − cells that can be propagated in IL-3 but grow slowly, if at all, in SCF, although they express the SCF receptor (c-kit). The response of these cells to IL-3 is stimulated synergistically by SCF, and we present evidence that both the synergy and the inhibition of c-kit responses may be mediated by direct interaction with IL-3 receptor. Further, the relative level of tyrosine phosphorylation of various substrates by either cytokine alone differs from that produced by the combination of the two cytokines, suggesting that cell signaling by the combination of the two cytokines differs from that produced by either alone.