SUT2, a putative sucrose sensor in sieve elements

In leaves, sucrose uptake kinetics involve high- and low-affinity components. A family of low- and high-affinity sucrose transporters (SUT) was identified. SUT1 serves as a high-affinity transporter essential for phloem loading and long-distance transport in solanaceous species. SUT4 is a low-affinity transporter with an expression pattern overlapping that of SUT1. Both SUT1 and SUT4 localize to enucleate sieve elements of tomato. New sucrose transporter-like proteins, named SUT2, from tomato and Arabidopsis contain extended cytoplasmic domains, thus structurally resembling the yeast sugar sensors SNF3 and RGT2. Features common to these sensors are low codon bias, environment of the start codon, low expression, and lack of detectable transport activity. In contrast to LeSUT1, which is induced during the sink-to-source transition of leaves, SUT2 is more highly expressed in sink than in source leaves and is inducible by sucrose. LeSUT2 protein colocalizes with the low- and high-affinity sucrose transporters in sieve elements of tomato petioles, indicating that multiple SUT mRNAs or proteins travel from companion cells to enucleate sieve elements. The SUT2 gene maps on chromosome V of potato and is linked to a major quantitative trait locus for tuber starch content and yield. Thus, the putative sugar sensor identified colocalizes with two other sucrose transporters, differs from them in kinetic properties, and potentially regulates the relative activity of low- and high-affinity sucrose transport into sieve elements.     

Molecular examination of a chromosome region that controls resistance to potato Y and A potyviruses in potato

 The gene Ry _adg that confers resistance to potato Y potyvirus (PVY) in the cultivated potato [Solanum tuberosum subsp. andigena, line 2x(v-2)7] is located on chromosome XI in a segment that contains three other known resistance genes in other syntenic solanaceous species. One of them is the gene N that controls resistance to tobacco mosaic tobamovirus in tobacco and has previously been isolated and sequenced. Three sequence-related, resistance gene-like (RGL) DNA fragments (354–369 bp) highly homologous to the gene N were PCR-amplified from the potato line 2x(v-2)7. Two RGL fragments (79 and 81% homologous to the N gene) co-segregated with Ry _adg among the 77 F1 progeny tested. These RGLs may originate from a resistance gene family on chromosome XI. The potato line 2x(v-2)7 also expressed resistance to potato A potyvirus (PVA), which was controlled by another locus on chromosome XI mapped ca. 6.8 cM distal to Ry _adg . Received: 18 December 1997 / Accepted: 30 December 1997     

High error rates for avian molecular sex identification primer sets applied to molted feathers

ABSTRACT Molted feathers are becoming increasingly important as a source of DNA for identifying the sex of individuals, and accurate methods for molecular sex identification are needed. Three molecular sex identification primer sets have been developed for use in nearly all nonratite birds, but performance of these primer sets has not been evaluated for molted feathers. For two species of birds, the Ring‐necked Pheasant (Phasianus colchicus) and the Scarlet Macaw (Ara macao), we evaluated success and error rates among primer sets using DNA from molted feathers and assessed the percentage of times an incorrect sex would be assigned when analyses are completed in duplicate. Amplification success rates differed among the primer sets for both species, ranging from 67.5% to 89.2% (P= 0.0002 and 0.009), and error rates were high, ranging from 1.9% to 24.2%. Success rates and error rates were not consistent between species and among primer sets. To improve the accuracy of molecular sex identification tests when using molted feathers, we suggest determining acceptable confidence levels in the accuracy of sex assignment, conducting pilot tests to evaluate the performance of different primer sets, and using high‐resolution electrophoresis systems to increase detection of errors.     

Development of a sterile cultivation system for rooting of shoot tip cultures (red raspberries) in duroplast foam

In order to improve the availability of oxygen for shoot tip cultures during the course of rooting and to overcome some further agar-related problems a sterile foam cultivation system was developed. Three commercially available foam substrates have been tested, but only a wide pore phenol resin foam proved to be useful in the rooting of red raspberry shoots (Rubus idaeus cv. ‘Gigant’). In comparison to root growth in agarous medium, the newly developed foam cultivation system improved the ramification of the root system, increased the number of root hairs significantly and reduced the friability (lignification) of roots. Also shoot length increased significantly. From the growth experiments it can be concluded that the chemical rather than physical properties of the foam substrate are growth limiting. Still some efforts must be made to improve and standardize the properties of the foam substrate for routine use in mass propagation of organ cultures in vitro.     

Post-Prandial Protein Handling: You Are What You Just Ate

Background

Protein turnover in skeletal muscle tissue is highly responsive to nutrient intake in healthy adults.

Objective

To provide a comprehensive overview of post-prandial protein handling, ranging from dietary protein digestion and amino acid absorption, the uptake of dietary protein derived amino acids over the leg, the post-prandial stimulation of muscle protein synthesis rates, to the incorporation of dietary protein derived amino acids in de novo muscle protein.

Design

12 healthy young males ingested 20 g intrinsically [1-¹³C]-phenylalanine labeled protein. In addition, primed continuous L-[ring-²H₅]-phenylalanine, L-[ring-²H₂]-tyrosine, and L-[1-¹³C]-leucine infusions were applied, with frequent collection of arterial and venous blood samples, and muscle biopsies throughout a 5 h post-prandial period. Dietary protein digestion, amino acid absorption, splanchnic amino acid extraction, amino acid uptake over the leg, and subsequent muscle protein synthesis were measured within a single in vivo human experiment.

Results

55.3±2.7% of the protein-derived phenylalanine was released in the circulation during the 5 h post-prandial period. The post-prandial rise in plasma essential amino acid availability improved leg muscle protein balance (from -291±72 to 103±66 μM·min^-1·100 mL leg volume^-1; P<0.001). Muscle protein synthesis rates increased significantly following protein ingestion (0.029±0.002 vs 0.044±0.004%·h^-1 based upon the muscle protein bound L-[ring-²H₅]-phenylalanine enrichments (P<0.01)), with substantial incorporation of dietary protein derived L-[1-¹³C]-phenylalanine into de novo muscle protein (from 0 to 0.0201±0.0025 MPE).

Conclusion

Ingestion of a single meal-like amount of protein allows ~55% of the protein derived amino acids to become available in the circulation, thereby improving whole-body and leg protein balance. About 20% of the dietary protein derived amino acids released in the circulation are taken up in skeletal muscle tissue following protein ingestion, thereby stimulating muscle protein synthesis rates and providing precursors for de novo muscle protein synthesis.

Trial Registration

trialregister.nl 3638     

Criteria Used in Clinical Practice to Guide Immunosuppressive Treatment in Patients with Primary Sclerosing Cholangitis

Background & Aims

Current guidelines recommend immunosuppressive treatment (IT) in patients with primary sclerosing cholangitis (PSC) and elevated aminotransferase levels more than five times the upper limit of normal and elevated serum IgG-levels above twice the upper limit of normal. Since there is no evidence to support this recommendation, we aimed to assess the criteria that guided clinicians in clinical practice to initiate IT in patients with previously diagnosed PSC.

Methods

This is a retrospective analysis of 196 PSC patients from seven German hepatology centers, of whom 36 patients had received IT solely for their liver disease during the course of PSC. Analyses were carried out using methods for competing risks.

Results

A simplified autoimmune hepatitis (AIH) score >5 (HR of 36, p<0.0001) and a modified histological activity index (mHAI) greater than 3/18 points (HR 3.6, p = 0.0274) were associated with the initiation of IT during the course of PSC. Of note, PSC patients who subsequently received IT differed already at the time of PSC diagnosis from those patients, who did not receive IT during follow-up: they presented with increased levels of IgG (p = 0.004) and more frequently had clinical signs of cirrhosis (p = 0.0002).

Conclusions

This is the first study which investigates the parameters associated with IT in patients with PSC in clinical practice. A simplified AIH score >5 and a mHAI score >3, suggesting concomitant features of AIH, influenced the decision to introduce IT during the course of PSC. In German clinical practice, the cutoffs used to guide IT may be lower than recommended by current guidelines.     

Shedding of dipeptidyl peptidase 4 is mediated by metalloproteases and up-regulated by hypoxia in human adipocytes and smooth muscle cells

Dipeptidyl peptidase 4 is an important drug target for diabetes and a novel adipokine. However, it is unknown how soluble DPP4 (sDPP4) is cleaved from the cell membrane and released into the circulation. We show here that MMP1, MMP2 and MMP14 are involved in DPP4 shedding from human vascular smooth muscle cells (SMC) and MMP9 from adipocytes. Hypoxia increased DPP4 shedding from SMC which is associated with increased mRNA expression of MMP1. Our data suggest that constitutive as well as hypoxia-induced DPP4 shedding occurs due to a complex interplay between different MMPs in cell type-specific manner.     

Operons Are a Conserved Feature of Nematode Genomes

The organization of genes into operons, clusters of genes that are co-transcribed to produce polycistronic pre-mRNAs, is a trait found in a wide range of eukaryotic groups, including multiple animal phyla. Operons are present in the class Chromadorea, one of the two main nematode classes, but their distribution in the other class, the Enoplea, is not known. We have surveyed the genomes of Trichinella spiralis, Trichuris muris, and Romanomermis culicivorax and identified the first putative operons in members of the Enoplea. Consistent with the mechanism of polycistronic RNA resolution in other nematodes, the mRNAs produced by genes downstream of the first gene in the T. spiralis and T. muris operons are trans-spliced to spliced leader RNAs, and we are able to detect polycistronic RNAs derived from these operons. Importantly, a putative intercistronic region from one of these potential enoplean operons confers polycistronic processing activity when expressed as part of a chimeric operon in Caenorhabditis elegans. We find that T. spiralis genes located in operons have an increased likelihood of having operonic C. elegans homologs. However, operon structure in terms of synteny and gene content is not tightly conserved between the two taxa, consistent with models of operon evolution. We have nevertheless identified putative operons conserved between Enoplea and Chromadorea. Our data suggest that operons and “spliced leader” (SL) trans-splicing predate the radiation of the nematode phylum, an inference which is supported by the phylogenetic profile of proteins known to be involved in nematode SL trans-splicing.