Regulation, replication, and integration functions of the Vibrio cholerae CTXφ are encoded by region RS2

CTXφ is a filamentous phage that encodes cholera toxin, one of the principal virulence factors of Vibrio cholerae . CTXφ is unusual among filamentous phages because it can either replicate as a plasmid or integrate into the V. cholerae chromosome at a specific site. The CTXφ genome has two regions, the 'core' and RS2. Integrated CTXφ is frequently flanked by an element known as RS1 which is related to RS2. The nucleotide sequences of RS2 and RS1 were determined. These related elements contain three nearly identical open reading frames (ORFs), which in RS2 were designated rstR , rstA2 and rstB2 . RS1 contains an additional ORF designated rstC . Functional analyses indicate that rstA2 is required for CTXφ replication and rstB2 is required for CTXφ integration. The amino terminus of RstR is similar to the amino termini of other phage-encoded repressors, and RstR represses the expression of rstA2 . Although genes with related functions are clustered in the genome of CTXφ in a way similar to those for other filamentous phages, the CTXφ RS2-encoded gene products mediating replication, integration and repression appear to be novel.     

Population ecology of the white-nosed coati (Nasua narica) on Barro Colorado Island, Panama

The white-nosed coati, Nasua narica , is a common Neotropical carnivore with a social structure of band-living adult females and solitary adult males. A coati population on Barro Colorado Island, Panama, was studied over a four-year period by mark-recapture, radiotelemetry. and direct observation of habituated individuals. The population density was approximately 51.5 individuals/km² and the sex ratio was 1:1. Band size varied from six to 26 individuals (mean = 15.3) with extensive fluctuation within and between years. Mean foraging group size was smaller (7.2 individuals) than population group size, and fluctuated with food availability, synchronous parturition, and the emigration of mature males. Mean home-range size of six bands was 0.33 km², and ranges of adjacent bands overlapped from 0–66%. One band fissioned during the study; however, the resulting bands did not disperse from the original home range. Seven adult males had a mean home-range size of 0.37 km², each extensively overlapping the home ranges of several other males. Observations of 10 adult males whose natal bands were known indicate that when males disperse they do not simultaneously leave the band's home range. Rather, their home ranges remain within or broadly overlapping those of their natal bands. This dispersal pattern is unusual within the order Carnivora.     

Targeting Determinants and Proposed Evolutionary Basis for the Sec and the Delta pH Protein Transport Systems in Chloroplast Thylakoid Membranes

Transport of proteins to the thylakoid lumen is accomplished by two precursor-specific pathways, the Sec and the unique Delta pH transport systems. Pathway selection is specified by transient lumen-targeting domains (LTDs) on precursor proteins. Here, chimeric and mutant LTDs were used to identify elements responsible for targeting specificity. The results showed that: (a) minimal signal peptide motifs consisting of charged N, hydrophobic H, and cleavage C domains were both necessary and sufficient for pathway-specific targeting; (b) exclusive targeting to the Delta pH pathway requires a twin arginine in the N domain and an H domain that is incompatible with the Sec pathway; (c) exclusive targeting to the Sec pathway is achieved by an N domain that lacks the twin arginine, although the twin arginine was completely compatible with the Sec system. A dual-targeting signal peptide, constructed by combining Delta pH and Sec domains, was used to simultaneously compare the transport capability of both pathways when confronted with different passenger proteins. Whereas Sec passengers were efficiently transported by both pathways, Delta pH passengers were arrested in translocation on the Sec pathway. This finding suggests that the Delta pH mechanism evolved to accommodate transport of proteins incompatible with the thylakoid Sec machinery.     

The Evolution of MHC Diversity by Segmental Duplication and Transposition of Retroelements

Sequence analysis of a 237 kb genomic fragment from the central region of the MHC has revealed that the HLA-B and HLA-C genes are contained within duplicated segments peri-B (53 kb) and peri-C (48 kb), respectively, and separated by an intervening sequence (IF) of 30 kb. The peri-B and peri-C segments share at least 90% sequence homology except when interrupted by insertions/deletions including Alu, L1, an endogenous retrovirus, and pseudogenes. The sequences of peri-B, IF, and peri-C were searched for the presence of Alu elements to use as markers of evolution, chromosomal rearrangements, and polymorphism. Of 29 Alu elements, 14 were identified in peri-B, 11 in peri-C, and 4 in IF. The Alu elements in peri-B and peri-C clustered phylogenetically into two clades which were classified as ``preduplication' and ``postduplication' clades. Four Alu J elements that are shared by peri-B and peri-C and are flanked by homologous sequences in their paralogous locations, respectively, clustered into a ``preduplication' clade. By contrast, the majority of Alu elements, which are unique to either peri-B or peri-C, clustered into a postduplication clade together with the Alu consensus subfamily members ranging from platyrrhine-specific (Spqxcg) to catarrhine-specific Alu sequences (Y). The insertion of platyrrhine-specific Alu elements in postduplication locations of peri-B and peri-C implies that these two segments are the products of a duplication which occurred in primates prior to the divergence of the New World primate from the human lineage (35–44 mya). Examination of the paralogous Alu integration sites revealed that 9 of 14 postduplication Alu sequences have produced microsatellites of different length and sequence within the Alu 3′-poly A tail. The present analysis supports the hypothesis that HLA-B and HLA-C genes are products of an extended segmental duplication between 44 and 81 million years ago (mya), and that subsequent diversification of both genomic segments occurred because of the mobility and mutation of retroelements such as Alu repeats. Received: 21 May 1997 / Accepted: 9 July 1997     

Anthropometrics Do Not Influence Dual X-ray Absorptiometry (DXA) Measurement of Fat in Normal to Obese Adults: A Comparison with In Vivo Neutron Activation Analysis (IVNA)

Dual-energy X-ray absorptiometry (DXA) is now a commonly used method for the determination of bone mineral status and body composition in humans. The purposes of this study were to compare fat mass by in vivo neutron activation analysis (FM_IVNA) with that by DXA (FM_DXA) in an anthropometrically heterogeneous sample of healthy adult men (n=33) and women (n=36) (19=≤BMI≤39), and to determine whether differences in fat mass estimates between the two methods (ΔFM) were attributable to subject anthropometry as defined by several circumference (waist, iliac crest, thigh) and skinfold thickness (umbilical, suprailiac, abdominal) measurements. No significant differences between FM_DXA and FM_IVNA were observed in men (p=0.46) or women (p=0.09). The two methods were very highly correlated in both sexes (women r²=0.97, p<0.001, men r²=0.91, p<0.001), although the regression line for men was significantly different from the line of identity (p=0.043). These results suggest modest trends toward underestimation of FM_DXA in men when FM_IVNA<18 kg, and overestimation in men when FM_IVNA>18 kg. ΔFM (IVNA-DXA) was not significantly related to any combination of skinfold thicknesses and circumferences in either gender. Age explained 27% of the variance in ΔFM for the men (p=0.008). Furthermore, ΔFM was not significantly related to inter-method disparity in total-body bone mineral measurements in men or women (p<0.05). The present study demonstrates strong correlation in fat measurements between IVNA and DXA in men and women ranging from normal to markedly obese. Correction for subject anthropometry does not significantly improve this relationship.     

Insulin activation of phosphatidylinositol 3-kinase in human skeletal muscle in vivo

Hickey, Matthew S., Charles J. Tanner, D. Sean O'Neill,Lydia J. Morgan, G. Lynis Dohm, and Joseph A. Houmard. Insulin activation of phosphatidylinositol 3-kinase in human skeletal muscle invivo. J. Appl. Physiol. 83(3):718-722, 1997.The purpose of this investigation was to determinewhether insulin-stimulated phosphatidylinositol 3-kinase (PI3-kinase)activity is detectable in needle biopsies of human skeletal muscle.Sixteen healthy nonobese males matched for age, percent fat, fastinginsulin, and fasting glucose participated in one of two experimentalprotocols. During an intravenous glucose tolerance test (IVGTT)protocol, insulin-stimulated PI3-kinase activity was determined frompercutaneous needle biopsies at 2, 5, and 15 min post-insulinadministration (0.025 U/kg). In the second group, a 2-h, 100 mU · m² · min¹euglycemic hyperinsulinemic clamp was performed, and biopsies wereobtained at 15, 60, and 120 min after insulin infusion was begun.Insulin stimulated PI3-kinase activity by 1.6 ± 0.2-, 2.2 ± 0.3-, and 2.2 ± 0.4-fold at 2, 5, and 15 min, respectively, duringthe IVGTT. During the clamp protocol, PI3-kinase was elevated by 5.3 ± 1.3-, 8.0 ± 2.6-, and 2.7 ± 1.4-fold abovebasal at 15, 60, and 120 min, respectively. Insulin-stimulatedPI3-kinase activity at 15 min post-insulin administration wassignificantly greater during the clamp protocol vs. the IVGTT(P < 0.05). These observations suggest that insulin-stimulated PI3-kinase activity is detectable inneedle biopsies of human skeletal muscle, and furthermore, that theeuglycemic, hyperinsulinemic clamp protocol may be a useful tool toassess insulin signaling in vivo.

     

A Modified Self-thinning Equation to Describe Size/Density Relationships for Defoliated Swards

Use of the self-thinning rule to describe size/density compensation(SDC) in defoliated swards is examined. It is shown that defoliationrelated variation in leaf area and associated morphogeneticchanges in plant structure necessitate slope corrections, designatedC_a and C_r , respectively. The theory predicts that reduced leafarea in more heavily defoliated swards will result in SDC atslopes more negative than -3/2 (variable leaf area SDC), andthat there will be a transition to -3/2 (constant leaf area)SDC at higher herbage mass. Empirical data from previous experiments with Lolium perenneL. and Medicago sativa L. are examined, and appear to confirmthe theoretical predictions, including the slope change at thepoint of transition from variable to constant leaf area SDC.This transition point, designated d_i , is subject to interspecificvariation and is related to the mature shoot size of a particularspecies. Some applications of this theory are discussed, andin particular a sward productivity index is proposed.Copyright1995, 1999 Academic Press Variable leaf area self-thinning, size/density compensation, Lolium perenne, Medicago sativa, sward productivity index     

Evidence that neutral spingomyelinase of cultured murine neuroblastoma cells is oriented externally on the plasma membrane

The activity of the neutral, Mg²⁺-stimulated sphingomyelinase of cultured neuroblastoma cells (N1E-115) is enriched in the plasma membrane fraction and is reduced following treatment of intact or broken cells with trypsin, α-chymotrypsin, papain, and protease. Two protease-sensitive enzymes of the cell interior (lactate dehydrogenase and NADPH-cytochrome c reductase) are not affected by protease treatment of intact cells. These results indicate that the neutral, Mg²⁺-stimulated sphingomyelinase is oriented externally on the plasma membrane of the cultured neuroblastoma cell.     

Serum indole-3-acetic acid in control subjects and newly abstinent alcoholics after an oral loading with L-tryptophan: a preliminary study using liquid chromatography with amperometric detection

Described in this paper is a rapid, isocratic assay for serum indole-3-acetic acid (IAA). The sample preparation involves only protein precipitation using sulfosalicylic acid, and the sensitivity of amperometric detection is in the picogram range. The chromatographic analysis time is approximately 4 min. The devised method was used for a longitudinal study of IAA levels in serum samples from control subjects and newly abstinent alcoholics. Dietary variations were eliminated by administering a 2.0-g loading dose of L-Trp to all subjects investigated. The results are presented in the form of cumulative frequency polygons. Preliminary data indicate no differences in IAA levels between newly abstinent alcoholics and control subjects.     

Albumin-induced plasma volume expansion: diurnal and temperature effects

To develop a reliable procedure for the acute expansion of plasma volume (PV), 26 male volunteers were randomly assigned to either a thermoneutral (25 degrees C and 40% relative humidity) or hot-dry (37 degrees C and 25% relative humidity) environment; subsequently each subject was seated for at least 1 h and then infused intravenously with either 100 or 200 ml of a 25% albumin solution or 0.9% saline. On the day before each infusion, PV was estimated by dye dilution using indocyanine green. Net percent change in PV (using hematocrit and hemoglobin values) was calculated at 1, 3, 6, 9, 12, and 24 h postinfusion. The PV of subjects residing in the heat after a 100-ml saline infusion increased significantly over 1-h values at 6, 9, and 12 h postinfusion but not at 24 h. The same trend, although not significant, was apparent at room temperature. The data suggest a slow isooncotic circadian pattern of PV expansion and contraction. The infusion of hyperoncotic albumin produced rapid expansion of plasma volume. With the low dose (25 g) at 1 h postinfusion, the expansion was 379 +/- 102 ml in the heat and 301 +/- 160 ml at room temperature. With the high dose (50 g) at 1 h postinfusion, the expansion was 479 +/- 84 ml in the heat and 427 +/- 147 ml at room temperature. The high dose produced an expansion that persisted for at least 9 h in subjects in either environment. The data suggest a mechanism for the retention of fluid during heat acclimatization and a useful procedure for plasma volume expansion in humans.