IntroductionThe LDL receptor plays a critical role in the regulation of plasma LDL levels by mediating approximately two thirds of LDL clearance (1-3). Loss of LDL receptor function leads to decreased LDL catabolism and elevated LDL levels (4). LDL receptor levels are affected by diet, hormones, and most dramatically, by mutations in the LDL receptor locus that lead to familial hypercholesterolemia (FH).Early studies of LDL metabolism in patients with FH revealed that in addition to the LDL clearance defect, they overproduce LDL (5, 6) and relatively small VLDL particles (7). VLDL is the metabolic precursor of LDL and is converted to LDL through the action of lipoprotein lipase, a triacylglycerol lipase that acts upon VLDL while it circulates in the bloodstream (8). Increased production of VLDL can lead to increased LDL simply by providing more precursor. In addition, impaired clearance of VLDL remnants can lead to LDL overproduction (9).A long-standing paradox in the lipoprotein field is posed by the cholesterol-lowering drugs known as statins. These drugs inhibit 3-hydroxy-3-methyl-glutaryl coenzyme A reductase, a tightly regulated step in the cholesterol biosynthetic pathway (10). Cells respond to the dearth of cholesterol by upregulating transcription of cholesterol-regulated genes, including the LDL receptor (11). Statins are ineffective in patients homozygous for null alleles of the LDL receptor (12). It has therefore been inferred that statins act by increasing LDL catabolism via upregulation of the LDL receptor. Paradoxically, statins do not always affect the LDL clearance rate. Rather, in many clinical studies (13)(14)(15) and animal studies (16,17), statins decrease VLDL and/or LDL production (reviewed in ref. 18).The post-translational fate of apoB, the major protein component of VLDL, can be explained by multiple mechanisms. In human and rat hepatoma cell lines, a large proportion of newly synthesized apoB is degraded within the secretory pathway (19). Thus, the rate of apoB secretion, and hence, VLDL secretion, from the liver is determined by the proportion of apoB that escapes coor post-translational degradation (20,21). In addition, reuptake of newly secreted lipoproteins has also been proposed to regulate the net output of apoB (22).How can the presence or absence of a functional LDL receptor affect the production of lipoproteins? We addressed this question by studying apoB secretion in cultured hepatocytes isolated from wild-type mice and mice lacking a functional LDL receptor (Ldlr -/-). Similar to FH, previous studies with Ldlr -/-mice revealed a decrease in LDL clearance (23) and a marked increase in plasma apoB levels (23-26). Our results with primary hepatocytes from these animals indicate that the LDL receptor is involved Jaap Twisk and Donald L. Gillian-Daniel contributed equally to this work.Received for publication October 6, 1999, and accepted in revised form December 28, 1999. Familial hypercholesterolemia is caused by mutations in the LDL receptor gene (Ldlr). Elevated plasma LDL ...

This chapter describes programs created to improve the preparation of future STEM faculty for their roles as educators.

The translation of specific maternal mRNAs is regulated during early development. For some mRNAs, an increase in translational activity is correlated with cytoplasmic extension of their poly(A) tails; for others, translational inactivation is correlated with removal of their poly(A) tails. Recent results in several systems suggest that events at the 3 end of the mRNA can affect the state of the 5 cap structure, m 7 G(5)ppp(5)G. We focus here on the potential role of cap modifications on translation during early development and on the question of whether any such modifications are dependent on cytoplasmic poly(A) addition or removal. To do so, we injected synthetic RNAs into Xenopus oocytes and examined their cap structures and translational activities during meiotic maturation. We draw four main conclusions. First, the activity of a cytoplasmic guanine-7-methyltransferase increases during oocyte maturation and stimulates translation of an injected mRNA bearing a nonmethylated GpppG cap. The importance of the cap for translation in oocytes is corroborated by the sensitivity of protein synthesis to cap analogs and by the inefficient translation of mRNAs bearing nonphysiologically capped 5 termini. Second, deadenylation during oocyte maturation does not cause decapping, in contrast to deadenylation-triggered decapping in Saccharomyces cerevisiae. Third, the poly(A) tail and the N-7 methyl group of the cap stimulate translation synergistically during oocyte maturation. Fourth, cap ribose methylation of certain mRNAs is very inefficient and is not required for their translational recruitment by poly(A). These results demonstrate that polyadenylation can cause translational recruitment independent of ribose methylation. We propose that polyadenylation enhances translation through at least two mechanisms that are distinguished by their dependence on ribose modification.

Mutations in the low density lipoprotein (LDL) receptor (LDLR)cause hypercholesterolemia because of inefficient LDL clearance from the circulation. In addition, there is a paradoxical oversecretion of the metabolic precursor of LDL, very low density lipoprotein (VLDL). We recently demonstrated that the LDLR mediates presecretory degradation of the major VLDL protein, apolipoprotein B (apoB). Kinetic studies suggested that the degradation process is initiated in the secretory pathway. Here, we evaluated the ability of several LDLR variants that are stalled within the secretory pathway to regulate apoB secretion. Both a naturally occurring mutant LDLR and an LDLR consisting of only the ligand-binding domains and a C-terminal endoplasmic reticulum (ER) retention sequence were localized to the ER and not at the cell surface. In the presence of either of the ER-localized LDLRs, apoB secretion was essentially abolished. When the ligand-binding domain of the truncated receptor was mutated the receptor was unable to block apoB secretion, indicating that the inhibition of apoB secretion depends on the ability of the LDLR to bind to its ligand. These findings establish LDLR-mediated pre-secretory apoB degradation as a pathway distinct from reuptake of nascent lipoproteins at the cell surface. The LDLR provides an example of a receptor that modulates export of its ligand from the ER.M olecular defects in the low density lipoprotein (LDL) receptor (LDLR) cause Familial Hypercholesterolemia (FH), a condition associated with elevated plasma LDL cholesterol levels (1). Reduced expression, altered ligand binding, or defective transport to the cell surface all lead to a reduction in the functionally effective population of LDLRs at the cell surface.LDL is produced in the circulation from its precursor, very low density lipoprotein (VLDL). Apolipoprotein B (apoB) is the major protein component of VLDL and LDL. Two observations have suggested that the LDLR might be involved in apoB secretion. First, overproduction of apoB-containing lipoproteins occurs in some cases of FH (2-4). Second, drugs that lower LDL levels by increasing the expression of the LDLR (statins) in many instances have been shown to lower LDL without increasing LDL clearance; i.e., they lower LDL and͞or VLDL production (5).The proportion of apoB that escapes degradation within the secretory pathway primarily determines the rate of VLDL secretion. We recently demonstrated that the presence of the LDLR greatly increases the proportion of apoB subject to presecretory degradation. Our results directly link VLDL overproduction in FH with the loss of the LDLR (6).Several additional studies support a role for the LDLR in modulating apoB secretion. Increased secretion of VLDL is observed in vivo from both Ldlr Ϫ/Ϫ and transgenic Ldlr Ϫ/Ϫ mice that overexpress the nuclear form of sterol regulatory element binding protein-1a (SREBP-1a) and in vitro in hepatocytes from these animals (7). In contrast, transgenic SREBP-1a animals with a wild-type LDLR accumulate cholesterol a...

No abstract