Biomedical subjects

J Stolz

Publications and source records attributed to J Stolz.

At least 19 recordsLinked to original sources

PubMed2005

[Reconstruction surgical procedures for iatrogenic injuries to the ureter. Examples of uncommon, however successful approaches to their management].

The authors present several less usual reconstructions of ureters after the primary iatrogenic lesions. The appropriate ureteric capacity to drain the urine after previous injury is quite demanding and hence before the definitive decision is made the wholesome health status of the patient is to be considered and all possible solutions should be weighted. The efficient primary urinary diversion above the lesion is unavoidable. The genuine reconstruction could be deferred by several weeks or months in the view that in some cases the least invasive (endourologic) methods can completely avoid the need of open surgery in difficult adhesions after previous major surgery.

Abdomen↗

PubMed2005

[Expression of PAX2 and PAX8 genes in conventional type of renal carcinoma and their role in the tumor prognosis].

BACKGROUND: Sporadic renal cell carcinoma is one of the most common kidney malignancies in adults (85%). According to the IARC (The International Agency of Research on Cancer) Czech Republic has the first world position in the incidence and mortality for RCC. The prognosis of RCC is very poor because of high mortality around 70 to 50% and unpredictable progression after tumor removal. More precise molecular prognostic markers are required. Genes PAX2 and PAX8 control cell division during embryonic development and plays crucial role in tumor development because of stimulation of cell proliferation and/or inhibition of apoptotic program. METHODS AND RESULTS: Our RCC sample collection contains 64 tumor samples and 10 "normal" renal samples extracted from the affected kidney. mRNA was isolated from all samples and converted into cDNA. Expression of PAX genes was analyzed by using relative quantification real-time PCR with TaqMan labelled probe and GAPDH gene as an endogenous control. CONCLUSIONS: Expression of PAX2 gene was found in 97% and expression of PAX8 gene was found in 89% of analyzed tumor samples. The expression of both target genes was found in all "normal" renal samples. The level of expression of both PAX genes was very variable with the range from hundred times lower to forty times higher in comparison with the expression of chosen endogenous control. There were found no correlations between the expression of target genes and clinical-histological markers. These results do not have prognostic value yet because of short duration of patient observation. Follow-up clinical data are essential for completion of this research.

Biomarkers, Tumor↗

PubMed2003

[Benefits and risks of urologic laparoscopic surgery in adult patients].

The paper describes the advantages and disadvantages of the laparoscopic operations, the number of which steadily rises in urology. The laparoscopic surgery is considered to be a benefit regarding the short postoperative hospital stay, painless postoperative course, and virtually non-existing postoperative paralytic ileus. As disadvantage are deemed the long learning curve for the operating personal, and high economical costs, which could be cut down only if short off-work period in productive population is included. In the paper, the pathophysiological guidelines are outlined and emphasized during the laparoscopic operation, which the surgical and anesthesiological teams have to have in mind. On the own cohort of patients, the numbers and types of operations are described, which have been done at our department.

Adult↗

PubMed2002

[Adjuvant therapy of renal carcinoma].

A treatment approach in a group of 1498 patients with renal cell carcinoma (RCC) was studied. Out of these, 86 patients had the primarily generalised form. For the therapy of primary advanced metastasis, the treatment strategy was changed from conservative to an active approach. Such approach is based on the removal of 75% of the tumor mass, the patient should be in good conditions with no metastases in CNS, bone and liver, and the primary tumor should not be a sarcoma. The new treatment strategy of patients with advanced RCC combines surgery and chemoimmunotherapy. 47 patients from the studied group were treated with combined INF alpha + IL-2 + 5FU + isotretinoin using the Atzpodien scheme. The toxicity was minimal and only in six patients the treatment was interrupted because of the disease progression. Results of this treatment strategy are encouraging, however, further randomised trials are recommended.

Adult↗

PubMed2000

Plant sucrose-H+ symporters mediate the transport of vitamin H.

A cDNA coding for a vitamin H (biotin) transport protein from Arabidopsis was identified by genetic complementation of a biotin uptake-deficient yeast mutant. Vitamin H transport by this protein was sensitive to the SH-group inhibitor p-chloromercuribenzene sulfonic acid (PCMBS) and to the uncoupler carbonyl cyanide-m-chlorophenylhydrazone (CCCP), suggesting an energy-dependent biotin-H+ symport mechanism. The transport activity could contribute to the so-far uncharacterized plant sucrose-H+ symporter AtSUC5 which mediates the energy-dependent transport of biotin and sucrose, and restores growth of the biotin transport-deficient yeast mutant on medium with low biotin concentrations. Functional comparison of the AtSUC5 transporter with previously characterized plant sucrose or monosaccharide transporters revealed that biotin transport may be a general and specific property of all plant sucrose transporters (sucrose/biotin-H+ symporters). This first report on a transporter with dual substrate specificity for two structurally unrelated molecules has a major impact on general thinking concerning the specificity of membrane transporters. The physiological relevance of this finding is discussed.

Arabidopsis↗

PubMed1999

Structural analysis of a plant sucrose carrier using monoclonal antibodies and bacteriophage lambda surface display.

Monoclonal antibodies were raised and selected against recombinant Plantago major PmSUC2 sucrose carrier protein. Epitopes of two monoclonal antibodies (PS2-1A2 and PS2-4D4) were mapped using N-terminally truncated PmSUC2 proteins and a lambda library displaying random PmSUC2 peptides. PS2-1A2 recognizes an octapeptide close to the N-terminus of PmSUC2, PS2-4D4 binds to a decapeptide at the very C-terminus. Analyses of antibody binding to yeast protoplasts with functionally active, tagged PmSUC2 protein revealed that both epitopes are located in cytoplasmic domains of PmSUC2. These results support a model for plant sucrose transporters containing 12 transmembrane helices with the N-terminus and the C-terminus on the cytoplasmic side of the plasma membrane.

Amino Acid Sequence↗

PubMed1999

Identification of the plasma membrane H+-biotin symporter of Saccharomyces cerevisiae by rescue of a fatty acid-auxotrophic mutant.

Bakers' yeast is auxotrophic for biotin (vitamin H) and depends on the efficient uptake of this compound from the environment. A mutant strain with strongly reduced biotin uptake and with reduced levels of protein biotinylation was identified. The strain was auxotrophic for long-chain fatty acids, and this auxotrophy could be suppressed with high levels of biotin in the medium. After transformation of this mutant with a yeast genomic library, the unassigned open reading frame YGR065C was identified to complement this mutation. This gene codes for a protein with 593 amino acids and 12 putative transmembrane helices. Northern blot analysis revealed that, in wild-type cells, the corresponding mRNA levels were increased at low biotin concentrations. Likewise, cellular biotin uptake was increased with decreasing biotin availability. Expression of YGR065C under the control of the constitutive ADH1 promoter resulted in very high biotin transport rates across the plasma membrane that were no longer regulated by the biotin concentration in the growth medium. We conclude that YGR065C encodes the first biotin transporter identified for a non-mammalian organism and designate this gene VHT1 for vitamin H transporter 1.

Amino Acid Sequence↗

PubMed1999

The fenpropimorph resistance gene FEN2 from Saccharomyces cerevisiae encodes a plasma membrane H+-pantothenate symporter.

The product of the FEN2 gene of Saccharomyces cerevisiae has previously been described as a protein conferring sensitivity to the antifungal agent fenpropimorph. Fen2p was postulated to act as a common regulator of carbon and nitrogen catabolite repression and of amino acid and ergosterol biosynthesis. In this paper, we present experimental evidence characterizing Fen2p as a plasma membrane-localized transporter for the vitamin pantothenate. The high affinity transport system (Km = 3.5 microM) is sensitive to uncouplers, suggesting a H+-pantothenate cotransport. Pantothenate transport rates in yeast are modulated by extracellular pantothenate, being maximal at low pantothenate concentrations. It is demonstrated that beta-alanine can suppress the growth defect of FEN2 wild-type and fen2 mutant cells on pantothenate-free medium. Evidence is presented that beta-alanine is transported by the general amino acid permease Gap1p. The relation among pantothenate transport, nitrogen catabolite repression, and sensitivity to the antifungal agent fenpropimorph is discussed.

Amino Acid Sequence↗

PubMed1999

Context dependency in Stroop's paradigm: when are words treated as nonlinguistic objects?

The "memory" literature emphasizes the notion of encoding variability, whereas the "word recognition and attention" literature typically asserts that it is impossible to prevent immediate lexical and semantic activation when single words are presented to skilled readers. In the four experiments reported here, the presence/absence of a Stroop effect was associated with the nature of the between-trial context. These data can be understood as an expression of contextually driven encoding variability, but are problematic for the decontextualized "automatic" processing account that has prevailed in the attention and performance literature for the last six decades.

Field Dependence-Independence↗

PubMed1998

Bacteriophage lambda surface display of a bacterial biotin acceptor domain reveals the minimal peptide size required for biotinylation.

Phage display is a powerful technique for identifying specific ligands to a given target. In this work random peptides derived from the biotin accepting domain of the Klebsiella pneumoniae oxaloacetate decarboxylase were displayed on bacteriophage lambda heads to determine the minimal sequence length that is necessary to effect biotinylation in vivo. Phages with a functional biotinylation domain were identified after affinity purification with immobilised avidin. All biotinylated phages isolated this way were found to have a sequence of 66 amino acids from the parental protein in common. This minimal biotinylation domain is fully functional as a biotin acceptor and more resistant to proteolytic attack compared to domains of larger size derived from the same protein. The data present the first example of a posttranslational protein modification analysed in a phage display system. Moreover, a biotin domain of reduced size and improved stability was identified, that should be superior to the larger parental protein as a tag to generate biotinylated fusion proteins.

Amino Acid Sequence↗

PubMed1998

[Kidney sparing surgery].

From 1988 to 1998 107 nephron sparing surgery have been done for localised renal cell tumor. The age was between 21 to 75. Nephron sparing surgery means removal of tumor consumed part of renal tissue with oncology clean border in healthy renal tissue. Elective indication for nephron sparing surgery is renal cell carcinoma on single kidney or on bilateral function disabled kidneys. In the last years indication for nephron sparing surgery have spread also to patients with both kidneys, who have single extrarenal growing renal cell carcinoma to the diameter 4.5 cm. In our group of follow up patients, within the interval 3 to 96 months from operation we have 96.26% survival without evidence of recurrent renal cell carcinoma.

Adult↗

PubMed1996

Expression of the PmSUC1 sucrose carrier gene from Plantago major L. is induced during seed development.

A cDNA clone of the plasma membrane sucrose-H+ symporter PmSUC1 from Plantago major L. has been isolated and expressed in Saccharomyces cerevisiae. The PmSUC1 protein was characterized in transgenic yeast and in proteoliposomes with an artificial proton-motive-force (pmf) generator. PmSUC1 catalyzes the active uptake of sucrose or maltose in the presence of pmf and is sensitive to uncouplers. Unlike the extremely pH-dependent PmSUC2 sucrose-H+ symporter, PmSUC1 is relatively insensitive to changes of the extracellular pH. In leaves and petioles of P. major, expression of PmSUC1 mRNA is restricted to the vascular system. The important new feature about PmSUC1 is that the highest mRNA levels are found in non-vascular tissue of P. major flowers where the gene is transiently expressed during the early stages of seed development. In situ hybridization experiments show that PmSUC1 is expressed only in young ovules; the putative physiological role of PmSUC1 is discussed.

Amino Acid Sequence↗

PubMed1996

Purification of the Chlorella HUP1 hexose-proton symporter to homogeneity and its reconstitution in vitro.

A prokaryotic biotin acceptor domain was fused to the carboxy terminal end of the Chlorella hexose-proton symporter. The plant symporter is biotinylated in vivo when expressed in Schizosaccharomyces pombe. The extended biotinylated transport protein is fully active, catalyzes accumulation of D-glucose analogs and restores growth of a glucose-uptake-deficient yeast strain. Crude membranes were solubilized with octyl-beta-D-glucoside in the presence of Escherichia coli L-alpha-phosphatidylethanolamine. Biotinylated symporter was purified to homogeneity by biotinavidin affinity chromatography. The symporter protein was reconstituted together with cytochrome-c oxidase prepared from beef heart mitochondria into proteo-liposomes. Cytochrome-c oxidase is a redox-driven H(+)-pump generating a proton motive force (inside negative and alkaline) while transferring electrons from cytochrome-c to oxygen; this energy is used by the symporter to accumulate D-glucose at least 30-fold. In the absence of the driving force the transport protein facilitates diffusion of D-glucose until the concentration equilibrium is reached. It was shown that maximal transport activity depends highly on the amount of co-reconstituted cytochrome-c oxidase and that the symporter possesses 10% of its in vivo turnover number under optimized in vitro transport conditions.

Amino Acid Sequence↗

PubMed1995

Rapid purification of a functionally active plant sucrose carrier from transgenic yeast using a bacterial biotin acceptor domain.

A rapid and efficient method has been used for the purification of a Plantago major sucrose carrier from Saccharomyces cerevisiae. The C-terminal fusion of a bacterial biotin acceptor domain to the carrier protein did not interfere with the targeting to the yeast plasma membrane nor with the catalytic activity of the sucrose carrier. The chimeric construct is biotinylated by yeast cells in vivo and represents the only biotinylated protein in yeast membranes. Solubilized biotinylated carrier protein binds selectively to immobilized monomeric avidin and can be eluted as pure protein with free biotin. The purified protein is functionally active and catalyzes the energy-dependent transport of sucrose into proteoliposomes.

Amino Acid Sequence↗

PubMed1994

Sugar transport across the plasma membranes of higher plants.

The fluxes of carbohydrates across the plasma membranes of higher-plant cells are catalysed mainly by monosaccharide and disaccharide-H+ symporters. cDNAs encoding these different transporters have been cloned recently and the functions and properties of the encoded proteins have been studied extensively in heterologous expression systems. Several of the proteins have been identified biochemically in these expression systems and their location in plants has been shown immunohistochemically or with transgenic plants which were transformed with reporter genes, expressed under the control of the promoters of individual transporter genes. In this paper we summarize the current knowledge on the molecular biology and biochemistry of higher-plant sugar transport proteins.

Biological Transport↗

PubMed1994

SUC1 and SUC2: two sucrose transporters from Arabidopsis thaliana; expression and characterization in baker's yeast and identification of the histidine-tagged protein.

An important, most likely essential step for the long distance transport of sucrose in higher plants is the energy-dependent, uncoupler-sensitive loading into phloem cells via a sucrose-H+ symporter. This paper describes functional expression in Saccharomyces cerevisiae of two cDNAs encoding energy-dependent sucrose transporters from the plasma membrane of Arabidopsis thaliana, SUC1 and SUC2. Yeast cells transformed with vectors allowing expression of either SUC1 or SUC2 under the control of the promoter of the yeast plasma membrane ATPase gene (PMA1) transport sucrose, and to a lesser extent also maltose, across their plasma membranes in an energy-dependent manner. The KM-values for sucrose transport are 0.50 mM and 0.77 mM, respectively, and transport by both proteins is strongly inhibited by uncouplers such as carbonyl cyanide m-chlorophenylhydrazone (CCCP) and dinitrophenol (DNP), or SH-group inhibitors. The VMAX but not the KM-values of sucrose transport depend on the energy status of transgenic yeast cells. The two proteins exhibit different patterns of pH dependence with SUC1 being much more active at neutral and slightly acidic pH values than SUC2. The proteins share 78% identical amino acids, their apparent molecular weights are 54.9 kDa and 54.5 kDA, respectively, and both proteins contain 12 putative transmembrane helices. A modified SUC1-His6 cDNA encoding a histidine tag at the SUC1 C-terminus was also expressed in S. cerevisiae. The tagged protein is fully active and is shown to migrate at an apparent molecular weight of 45 kDa on 10% SDS-polyacrylamide gels.

Adenosine Triphosphatases↗

PubMed1994

Functional reconstitution of the solubilized Arabidopsis thaliana STP1 monosaccharide-H+ symporter in lipid vesicles and purification of the histidine tagged protein from transgenic Saccharomyces cerevisiae.

Complete DNA sequences encoding the Arabidopsis thaliana STP1 monosaccharide/H+ symporter or a histidine-tagged STP1-His6 protein were expressed in baker's yeast Saccharomyces cerevisiae. Both wild-type STP1 and the recombinant his-tagged protein were located in the plasma membranes of transformed yeast cells. The C-terminal modification caused no loss of transport activity compared with the wild-type protein. Anti-STP1-antibodies were used to confirm the identity of the protein in yeast and to compare the apparent molecular weights of STP1 proteins in membrane extracts from yeast or Arabidopsis thaliana. Purified yeast plasma membranes were fused with proteoliposomes consisting of Escherichia coli lipids and beef heart cytochrome-c oxidase. Addition of ascorbate/TMPD/cytochrome-c to these fused vesicles caused an immediate formation of membrane potential (inside negative; monitored with [3H]tetraphenylphosphonium cations) and a simultaneous, uncoupler-sensitive influx of D-glucose into the energized vesicles. STP1-His6 protein is functionally active after solubilization with octyl-beta-D-glucoside, which was shown by insertion of the protein into proteoliposomes by detergent dilution and determination of the resulting transport capacity. Detergent extracts from either total membranes or plasma membranes of transgenic yeast cells were used for one-step purification of the STP1-His6 protein on Ni(2+)-NTA columns. The identity of the purified protein was checked by immunoblotting and N-terminal sequencing.

Amino Acid Sequence↗

PubMed1994

A phloem-specific sucrose-H+ symporter from Plantago major L. supports the model of apoplastic phloem loading.

In this paper the cloning of a full-length cDNA clone encoding the PmSUC2 sucrose-H+ symporter from Plantago major is described. This plant allows the simple preparation of vascular bundles from the basal regions of fully developed source leaves and thus a separation of vascular and non-vascular tissue. A cDNA library was constructed from poly(A)+ RNA isolated from vascular bundles and used for the subsequent cloning of cDNAs. The respective mRNA is specifically expressed in the vascular bundles as shown on Northern blots of total RNA from vascular and non-vascular tissues. The PmSUC2 protein has 12 putative transmembrane helices and is highly homologous to other plant sucrose transporters. Substrate specificity and energy dependence of the transporter encoded by this cDNA were determined by expression in baker's yeast Saccharomyces cerevisiae. The PmSUC2 protein catalyses the transport of sucrose into transgenic yeast cells. Invertase null mutants of yeast expressing PmSUC2 accumulate sucrose more than 200-fold. This transport was sensitive to uncouplers or SH-group inhibitors. Plasma membranes from yeast cells expressing the PmSUC2 protein were purified and fused to proteoliposomes containing cytochrome-c-oxidase. In this system sucrose is accumulated only when proton motive force is generated, indicating that PmSUC2 is a sucrose-H+ symporter. The apparent molecular weight of the PmSUC2 protein is 35 kDa on 10% SDS-polyacrylamide gels. The presented data strongly support the theory of phloem loading from the apoplastic space by a sucrose-H+ symporter.

Amino Acid Sequence↗