Chloroplast DNA variation and the recent radiation of the giant senecios (Asteraceae) on the tall mountains of eastern Africa.

Chloroplast DNA restriction-site variation was surveyed among 40 accessions representing all 11 species of giant senecios (Dendrosenecio, Asteraceae) at all but one known location, plus three outgroup species. Remarkably little variation (only 9 variable sites out of roughly 1000 sites examined) was found among the 40 giant senecio accessions, yet as a group they differ significantly (at 18 sites) from Cineraria deltoidea, the closest known relative. This pattern indicates that the giant senecios underwent a recent dramatic radiation in eastern Africa and evolved from a relatively isolated lineage within the Senecioneae. Biogeographic interpretation of the molecular phylogeny suggests that the giant senecios originated high on Mt. Kilimanjaro, with subsequent dispersion to the Aberdares, Mt. Kenya, and the Cherangani Hills, followed by dispersion westward to the Ruwenzori Mountains, and then south to the Virunga Mountains, Mt. Kahuzi, and Mt. Muhi, but with dispersion back to Mt. Elgon. Geographic radiation was an important antecedent to the diversification in eastern Africa, which primarily involved repeated altitudinal radiation, both up and down the mountains, leading to morphological parallelism in both directions. In general, the plants on a given mountain are more closely related to each other than they are to plants on other mountains, and plants on nearby mountains are more closely related to each other than they are to plants on more distant mountains. The individual steps of the geographic radiation have occurred at various altitudes, some clearly the result of intermountain dispersal. The molecular evidence suggests that two species are extant ancestors to other species on the same or nearby mountains.

Targeted mutation reveals a central role for SR-BI in hepatic selective uptake of high density lipoprotein cholesterol

Scavenger receptor BI (SR-BI) is a cell surface receptor that binds high density lipoproteins (HDL) and mediates selective uptake of HDL cholesteryl esters (CE) in transfected cells. To address the physiological role of SR-BI in HDL cholesterol homeostasis, mice were generated bearing an SR-BI promoter mutation that resulted in decreased expression of the receptor in homozygous mutant (designated SR-BI att) mice. Hepatic expression of the receptor was reduced by 53% with a corresponding increase in total plasma cholesterol levels of 50–70% in SR-BI att mice, attributable almost exclusively to elevated plasma HDL. In addition to increased HDL-CE, HDL phospholipids and apo A-1 levels were elevated, and there was an increase in HDL particle size in mutant mice. Metabolic studies using HDL bearing nondegradable radiolabels in both the protein and lipid components demonstrated that reducing hepatic SR-BI expression by half was associated with a decrease of 47% in selective uptake of CE by the liver, and a corresponding reduction of 53% in selective removal of HDL-CE from plasma. Taken together, these findings strongly support a pivotal role for hepatic SR-BI expression in regulating plasma HDL levels and indicate that SR-BI is the major molecule mediating selective CE uptake by the liver. The inverse correlation between plasma HDL levels and atherosclerosis further suggests that SR-BI may influence the development of coronary artery disease.

A test of alternative models of diversification in tropical rainforests: Ecological gradients vs. rainforest refugia

Comparison of mitochondrial and morphological divergence in eight populations of a widespread leaf-litter skink is used to determine the relative importance of geographic isolation and natural selection in generating phenotypic diversity in the Wet Tropics Rainforest region of Australia. The populations occur in two geographically isolated regions, and within each region, in two different habitats (closed rainforest and tall open forest) that span a well characterized ecological gradient. Morphological differences among ancient geographic isolates (separated for several million years, judging by their mitochondrial DNA sequence divergence) were slight, but morphological and life history differences among habitats were large and occurred despite moderate to high levels of mitochondrial gene flow. A field experiment identified avian predation as one potential agent of natural selection. These results indicate that natural selection operating across ecological gradients can be more important than geographic isolation in similar habitats in generating phenotypic diversity. In addition, our results indicate that selection is sufficiently strong to overcome the homogenizing effects of gene flow, a necessary first step toward speciation in continuously distributed populations. Because ecological gradients may be a source of evolutionary novelty, and perhaps new species, their conservation warrants greater attention. This is particularly true in tropical regions, where most reserves do not include ecological gradients and transitional habitats.

The Phosphatidylinositol 3-Phosphate Binding Protein Vac1p Interacts with a Rab GTPase and a Sec1p Homologue to Facilitate Vesicle-mediated Vacuolar Protein Sorting

Activated GTP-bound Rab proteins are thought to interact with effectors to elicit vesicle targeting and fusion events. Vesicle-associated v-SNARE and target membrane t-SNARE proteins are also involved in vesicular transport. Little is known about the functional relationship between Rabs and SNARE protein complexes. We have constructed an activated allele of VPS21, a yeast Rab protein involved in vacuolar protein sorting, and demonstrated an allele-specific interaction between Vps21p and Vac1p. Vac1p was found to bind the Sec1p homologue Vps45p. Although no association between Vps21p and Vps45p was seen, a genetic interaction between VPS21 and VPS45 was observed. Vac1p contains a zinc-binding FYVE finger that may bind phosphatidylinositol 3-phosphate [PtdIns(3)P]. In other FYVE domain proteins, this motif and PtdIns(3)P are necessary for membrane association. Vac1 proteins with mutant FYVE fingers still associated with membranes but showed vacuolar protein sorting defects and reduced interactions with Vps45p and activated Vps21p. Vac1p membrane association was not dependent on PtdIns(3)P, Pep12p, Vps21p, Vps45p, or the PtdIns 3-kinase, Vps34p. Vac1p FYVE finger mutant missorting phenotypes were suppressed by a defective allele of VPS34. These data indicate that PtdIns(3)P may perform a regulatory role, possibly involved in mediating Vac1p protein–protein interactions. We propose that activated-Vps21p interacts with its effector, Vac1p, which interacts with Vps45p to regulate the Golgi to endosome SNARE complex.

Localization of atherosclerosis susceptibility loci to chromosomes 4 and 6 using the Ldlr knockout mouse model

Atherosclerosis is a complex disease resulting from the interaction of multiple genes. We have used the Ldlr knockout mouse model in an interspecific genetic cross to map atherosclerosis susceptibility loci. A total of 174 (MOLF/Ei × B6.129S7-Ldlrtm1Her) × C57BL/6J-Ldlrtm1Her backcross mice, homozygous for the Ldlr null allele, were fed a Western-type diet for 3 months and then killed for quantification of aortic lesions. A genome scan was carried out by using DNA pools and microsatellite markers spaced at ≈18-centimorgan intervals. Quantitative trait locus analysis of individual backcross mice confirmed linkages to chromosomes 4 (Athsq1, logarithm of odds = 6.2) and 6 (Athsq2, logarithm of odds = 6.7). Athsq1 affected lesions in females only whereas Athsq2 affected both sexes. Among females, the loci accounted for ≈50% of the total variance of lesion area. The susceptible allele at Athsq1 was derived from the MOLF/Ei genome whereas the susceptible allele at Athsq2 was derived from C57BL/6J. Inheritance of susceptible alleles at both loci conferred a 2-fold difference in lesion area, suggesting an additive effect of Athsq1 and Athsq2. No associations were observed between the quantitative trait loci and levels of plasma total cholesterol, high density lipoprotein cholesterol, non-high density lipoprotein cholesterol, insulin, or body weight. We provide strong evidence for complex inheritance of atherosclerosis in mice with elevated plasma low density lipoprotein cholesterol and show a major influence of nonlipoprotein-related factors on disease susceptibility. Athsq1 and Athsq2 represent candidate susceptibility loci for human atherosclerosis, most likely residing on chromosomes 1p36–32 and 12p13–12, respectively.

Expression of a delta 9 14:0-acyl carrier protein fatty acid desaturase gene is necessary for the production of omega 5 anacardic acids found in pest-resistant geranium (Pelargonium xhortorum).

Anacardic acids, a class of secondary compounds derived from fatty acids, are found in a variety of dicotyledonous families. Pest resistance (e.g., spider mites and aphids) in Pelargonium xhortorum (geranium) is associated with high levels (approximately 81%) of unsaturated 22:1 omega 5 and 24:1 omega 5 anacardic acids in the glandular trichome exudate. A single dominant locus controls the production of these omega 5 anacardic acids, which arise from novel 16:1 delta 11 and 18:1 delta 13 fatty acids. We describe the isolation and characterization of a cDNA encoding a unique delta 9 14:0-acyl carrier protein fatty acid desaturase. Several lines of evidence indicated that expression of this desaturase leads to the production of the omega 5 anacardic acids involved in pest resistance. First, its expression was found in pest-resistant, but not suspectible, plants and its expression followed the production of the omega 5 anacardic acids in segregating populations. Second, its expression and the occurrence of the novel 16:1 delta 11 and 18:1 delta 13 fatty acids and the omega 5 anacardic acids were specific to tall glandular trichomes. Third, assays of the recombinant protein demonstrated that this desaturase produced the 14:1 delta 9 fatty acid precursor to the novel 16:1 delta 11 and 18:1 delta 13 fatty acids. Based on our genetic and biochemical studies, we conclude that expression of this delta 9 14:0-ACP desaturase gene is required for the production of omega 5 anacardic acids that have been shown to be necessary for pest resistance in geranium.

Rho protein regulates tight junctions and perijunctional actin organization in polarized epithelia.

The rho family of GTP-binding proteins regulates actin filament organization. In unpolarized mammalian cells, rho proteins regulate the assembly of actin-containing stress fibers at the cell-matrix interface. Polarized epithelial cells, in contrast, are tall and cylindrical with well developed intercellular tight junctions that permit them to behave as biologic barriers. We report that rho regulates filamentous actin organization preferentially in the apical pole of polarized intestinal epithelial cells and, in so doing, influences the organization and permeability of the associated apical tight junctions. Thus, barrier function, which is an essential characteristic of columnar epithelia, is regulated by rho.