The mammalian mitochondrial genome encodes 13 proteins, which are synthesized at the direction of nine monocistronic and two dicistronic mRNAs. These mRNAs lack both 59 and 39 untranslated regions. The mechanism by which the specialized mitochondrial translational apparatus locates start codons and initiates translation of these leaderless mRNAs is currently unknown. To better understand this mechanism, the secondary structures near the start codons of all 13 open reading frames have been analyzed using RNA SHAPE chemistry. The extent of structure in these mRNAs as assessed experimentally is distinctly lower than would be predicted by current algorithms based on free energy minimization alone. We find that the 59 ends of all mitochondrial mRNAs are highly unstructured. The first 35 nucleotides for all mitochondrial mRNAs form structures with free energies less favorable than À3 kcal/mol, equal to or less than a single typical base pair. The start codons, which lie at the very 59 ends of these mRNAs, are accessible within single stranded motifs in all cases, making them potentially poised for ribosome binding. These data are consistent with a model in which the specialized mitochondrial ribosome preferentially allows passage of unstructured 59 sequences into the mRNA entrance site to participate in translation initiation.

Objective Reverse cholesterol transport (RCT) involves the removal of cholesterol from peripheral tissue for excretion in the feces. Here, we determined whether red blood cells (RBCs) can contribute to RCT. Methods and Results We performed a series of studies in apoAI-deficient mice where the HDL-mediated pathway of RCT is greatly diminished. RBCs carried a higher fraction of whole blood cholesterol than plasma in apoAI-deficient mice, and as least as much of the labeled cholesterol derived from injected foam cells appeared in RBCs compared to plasma. To determine if RBCs mediate RCT to the fecal compartment, we measured RCT in anemic and control apoAI-deficient mice and found that anemia decreased RCT to the feces by over 35% after correcting for fecal mass. Transfusion of [3H]cholesterol labeled RBCs led to robust delivery of the labeled cholesterol to the feces in apoAI-deficient hosts. In wild type mice, the majority of the blood cholesterol mass, as well as [3H]cholesterol derived from the injected foam cells, was found in plasma, and anemia did not significantly alter RCT to the feces after correction for fecal mass. Conclusion The RBC cholesterol pool is dynamic and facilitates RCT of peripheral cholesterol to the feces, particularly in the low HDL state.

Drosophila N-cadherin (CadN) is an evolutionarily conserved classic cadherin which has a large, complex extracellular domain and a catenin-binding cytoplasmic domain. The CadN locus contains three modules of alternative exons (7a/b, 13a/b, and 18a/b) and undergoes alternative splicing to generate multiple isoforms. Using quantitative transcript analyses and green fluorescent protein-based cell sorting, we found that during development CadN alternative splicing is regulated in a temporal but not cell-type-specific fashion. In particular, exon 18b is predominantly expressed during early developmental stages, while exon 18a is prevalent at the late developmental and adult stages. All CadN isoforms share the same molecular architecture but have different sequences in their extracellular and transmembrane domains, suggesting functional diversity. In vitro quantitative cell aggregation assays revealed that all CadN isoforms mediate homophilic interactions, but the isoforms encoded by exon 18b have a higher adhesive activity than those by its alternative, 18a. Domainswapping experiments further revealed that the different sequences in the transmembrane domains of isoforms are responsible for their differential adhesive activities. CadN alternative splicing might provide a novel mechanism to fine-tune its adhesive activity at different developmental stages or to restrict the use of high-affinity 18b-type isoforms at the adult stage.

High levels of high density lipoprotein-cholesterol (HDL-C) are associated with lower risk for cardiovascular disease in epidemiological studies ( 1 ). Although several mechanisms may play a role in HDL's protective effect, HDL and its major protein constituent, apoA1, are critical components of the reverse cholesterol transport (RCT) pathway, in which cholesterol is removed from peripheral tissues and transferred to the liver for excretion. In the fi rst step of the RCT pathway, lipid-poor apoA1 acts as an acceptor for cell cholesterol and phospholipids via the cell membrane protein ABCA1, generating nascent HDL.Recently, "HDL is the good cholesterol" hypothesis has suffered several setbacks. For example, several trials of HDL-C-raising drugs, including cholesteryl ester transfer protein inhibitors and niacin, failed to demonstrate clinical benefi t ( 2-5 ). Furthermore, a Mendelian randomization study did not associate gene variants that solely alter HDL-C with coronary artery disease (CAD) incidence ( 6 ). That said, human and mouse models with defective RCT, via mutations in ABCA1, apoA1, or scavenger receptor class B type I (SR-BI), are more susceptible to atherosclerosis independent Abstract HDL functions are impaired by myeloperoxidase (MPO), which selectively targets and oxidizes human apoA1. We previously found that the 4WF isoform of human apoA1, in which the four tryptophan residues are substituted with phenylalanine, is resistant to MPO-mediated loss of function. The purpose of this study was to generate 4WF apoA1 transgenic mice and compare functional properties of the 4WF and wild-type human apoA1 isoforms in vivo . Male mice had signifi cantly higher plasma apoA1 levels than females for both isoforms of human apoA1, attributed to different production rates. With matched plasma apoA1 levels, 4WF transgenics had a trend for slightly less HDL-cholesterol versus human apoA1 transgenics. While 4WF transgenics had 31% less reverse cholesterol transport (RCT) to the plasma compartment, equivalent RCT to the liver and feces was observed. Plasma from both strains had similar ability to accept cholesterol and facilitate ex vivo cholesterol effl ux from macrophages. Furthermore, we observed that 4WF transgenic HDL was partially ( ‫ف‬ 50%) protected from MPOmediated loss of function while human apoA1 transgenic HDL lost all ABCA1-dependent cholesterol acceptor activity. In conclusion, the structure and function of HDL from 4WF transgenic mice was not different than HDL derived from human apoA1 transgenic mice. -Berisha, S.