Genotyping over 100,000 SNPs on a pair of oligonucleotide arrays

We present a genotyping method for simultaneously scoring 116,204 SNPs using oligonucleotide arrays. At call rates >99%, reproducibility is >99.97% and accuracy, as measured by inheritance in trios and concordance with the HapMap Project, is >99.7%. Average intermarker distance is 23.6 kb, and 92% of the genome is within 100 kb of a SNP marker. Average heterozygosity is 0.30, with 105,511 SNPs having minor allele frequencies >5%.     

Targeting of the Arf-like GTPase Arl3p to the Golgi requires N-terminal acetylation and the membrane protein Sys1p

The GTPase Arl3p is required to recruit a second GTPase, Arl1p, to the Golgi in Saccharomyces cerevisiae. Arl1p binds to the GRIP domain, which is present in a number of long coiled-coil proteins or 'golgins'. Here we show that Arl3p is not myristoylated like most members of the Arf family, but is instead amino-terminally acetylated by the NatC complex. Targeting of Arl3p also requires a Golgi membrane protein Sys1p. The human homologues of Arl3p (Arf-related protein 1 (ARFRP1)) and Sys1p (hSys1) can be isolated in a complex after chemical cross-linking. This suggests that the targeting of ARFRP1/Arl3p to the Golgi is mediated by a direct interaction between its acetylated N terminus and Sys1p/hSys1.     

Kin or self‐recognition? Colonial fusibility of the bryozoan Celleporella hyalina

We estimated fusion frequency with respect to coancestry in the bryozoan Celleporella hyalina, whose briefly planktonic sexually produced larvae settle on algal substrata and proceed to form encrusting colonies by iterative budding. Frequency of fusion between paired colonies growing on an artificial substratum was positively correlated with coefficient of relatedness, with allorecognition ability increasing during the early stages of colonial growth after larval settlement. Parents repressed the growth of F1 progeny with which they had fused. The results are concordant with the Feldgarden-Yund model of selection for self-recognition, which regards fusion with kin as an inevitable source of error whose cost diminishes with increasing relatedness. Contrary to fusion compatibility, gametic compatibility is negatively correlated with coancestry, indicating a need for further research on the possibility of common or linked genetic control that has opposite effect at somatic and gametic levels.     

Structure-function analysis of the presumptive Arabidopsis auxin permease AUX1

We have investigated the subcellular localization, the domain topology, and the amino acid residues that are critical for the function of the presumptive Arabidopsis thaliana auxin influx carrier AUX1. Biochemical fractionation experiments and confocal studies using an N-terminal yellow fluorescent protein (YFP) fusion observed that AUX1 colocalized with plasma membrane (PM) markers. Because of its PM localization, we were able to take advantage of the steep pH gradient that exists across the plant cell PM to investigate AUX1 topology using YFP as a pH-sensitive probe. The YFP-coding sequence was inserted in selected AUX1 hydrophilic loops to orient surface domains on either apoplastic or cytoplasmic faces of the PM based on the absence or presence of YFP fluorescence, respectively. We were able to demonstrate in conjunction with helix prediction programs that AUX1 represents a polytopic membrane protein composed of 11 transmembrane spanning domains. In parallel, a large aux1 allelic series containing null, partial-loss-of-function, and conditional mutations was characterized to identify the functionally important domains and amino acid residues within the AUX1 polypeptide. Whereas almost all partial-loss-of-function and null alleles cluster in the core permease region, the sole conditional allele aux1-7 modifies the function of the external C-terminal domain.     

Force-velocity relationship and stretch-shortening cycle function in sprint and endurance athletes

This study examined the torque-velocity and power-velocity relationships of quadriceps muscle function, stretch shortening cycle function, and leg-spring stiffness in sprint and endurance athletes. Isokinetic maximal knee extension torque was obtained from 7 sprinters and 7 endurance athletes using a Con-trex isokinetic dynamometer. Torque and power measures were corrected for lean-thigh cross-sectional area and lean-thigh volume, respectively. Stretch-shortening cycle function and muscle stiffness measurements were obtained while subjects performed single-legged squat, countermovement, and drop-rebound jumps on an inclined sledge and force-plate apparatus. The results indicated that sprinters generated, on average, 0.15 +/- 0.05 N.m.cm(-2) more torque across all velocities compared with endurance athletes. Significant differences were also found in the power-velocity relationships between the 2 groups. The sprinters performed significantly better than the endurance athletes on all jumps, but there were no differences in prestretch augmentation between the groups. The average vertical leg stiffness during drop jumps was significantly higher for sprinters (5.86 N.m(-1)) compared with endurance runners (3.38 N.m(-1)). The findings reinforce the need for power training to be carried out at fast contraction speeds but also show that SSC function remains important in endurance running.     

CD1d function is regulated by microsomal triglyceride transfer protein

CD1d is a major histocompatibility complex (MHC) class I-related molecule that functions in glycolipid antigen presentation to distinct subsets of T cells that express natural killer receptors and an invariant T-cell receptor-alpha chain (invariant NKT cells). The acquisition of glycolipid antigens by CD1d occurs, in part, in endosomes through the function of resident lipid transfer proteins, namely saposins. Here we show that microsomal triglyceride transfer protein (MTP), a protein that resides in the endoplasmic reticulum of hepatocytes and intestinal epithelial cells (IECs) and is essential for lipidation of apolipoprotein B, associates with CD1d in hepatocytes. Hepatocytes from animals in which Mttp (the gene encoding MTP) has been conditionally deleted, and IECs in which Mttp gene products have been silenced, are unable to activate invariant NKT cells. Conditional deletion of the Mttp gene in hepatocytes is associated with a redistribution of CD1d expression, and Mttp-deleted mice are resistant to immunopathologies associated with invariant NKT cell-mediated hepatitis and colitis. These studies indicate that the CD1d-regulating function of MTP in the endoplasmic reticulum is complementary to that of the saposins in endosomes in vivo.     

Elevated dietary salt suppresses renin secretion but not thirst evoked by arterial hypotension in rats

Increased dietary salt intake was used as a nonpharmacological tool to blunt hypotension-induced increases in plasma renin activity (PRA) in order to evaluate the contribution of the renin-angiotensin system (RAS) to hypotension-induced thirst. Rats were maintained on 8% NaCl (high) or 1% NaCl (standard) diet for at least 2 wk, and then arterial hypotension was produced by administration of the arteriolar vasodilator diazoxide. Despite marked reductions in PRA, rats maintained on the high-salt diet drank similar amounts of water, displayed similar latencies to drink, and had similar degrees of hypotension compared with rats maintained on the standard diet. Furthermore, blockade of ANG II production by an intravenous infusion of the angiotensin-converting enzyme inhibitor captopril attenuated the hypotension-induced water intake similarly in rats fed standard and high-salt diet. Additional experiments showed that increases in dietary salt did not alter thirst stimulated by the acetylcholine agonist carbachol administered into the lateral ventricle; however, increases in dietary salt did enhance thirst evoked by central ANG II. Collectively, the present findings suggest that hypotension-evoked thirst in rats fed a high-salt diet is dependent on the peripheral RAS despite marked reductions in PRA.