Biomedical subjects

D Kaiser

Publications and source records attributed to D Kaiser.

At least 19 recordsLinked to original sources

PubMed1999

Myxococcus cells respond to elastic forces in their substrate.

Elasticotaxis describes the ability of Myxococcus xanthus cells to sense and to respond to elastic forces within an agar gel on which they rest. Within 5 min of the application of stress, each cell begins to reorient its long axis perpendicular to the stress force. The cells then glide in that direction, and the swarm becomes asymmetric. A quantifiable assay for the strength of elasticotaxis is based on the change in swarm shape from circular to elliptic. By using a collection of isogenic motility mutants, it has been found that the ability to respond to stress in agar depends totally on adventurous (A) motility, but not at all on social (S) motility or on the frz genes. In fact, S- mutants (which are moving only by means of A motility) respond to the applied stress more strongly than does the wild type, despite the fact that their spreading rates are slower than that of the wt strain. Based on the swarming and elasticotactic phenotypes of isogenic frizzy strains that were also defective either in A or S motility, frz behaves as if part of the S motility system.

Agar↗

PubMed1999

Cell fate and organogenesis in bacteria.

Intercellular signaling through the Notch receptor and its ligands leads to the spatial differentiation of cell fate in vertebrates and invertebrates. In Myxococcus xanthus, fruiting-body development requires the transmission of a cell-bound intercellular signal by the protein called C-factor, which is functionally equivalent to the eukaryotic Notch ligands. Functional parallels between these two signaling systems include strong positive and negative feedback, and a consequent role in spatial differentiation. Consideration of these parallels enables us to make testable experimental predictions about Notch and C-signaling.

Animals↗

PubMed1999

N-hydroxy peptides as substrates for alpha-chymotrypsin.

An N-hydroxylated peptide bond was found to be cleaved faster by an endopeptidase than the corresponding peptide bond. This preferred enzymatic cleavage was detected during proteolytic studies of the N-hydroxy peptide SIINFpsi[CO-N(OH)]GKL in the presence of the serine protease alpha-chymotrypsin in comparison with the natural SIINFEKL epitope and related analogs. For the first time, the replacement of the peptide bond by another motif afforded an oligomer which is degraded faster than the natural peptide. The N-hydroxy peptide is also more sensitive to the enzymatic degradation than the Gly-containing analog SIINFGKL. A tentative explanation for the unexpected higher cleavage rate of the CO-N(OH) bond is given on the basis of the N-OH intramolecular H-bonding capacity as indicated by NMR experiments. This property of the hydroxamate group may be of particular advantage for the introduction of a specific cleavage site within peptidomimetics or in prodrugs.

Amino Acid Sequence↗

PubMed1999

Type IV pili and cell motility.

Type IV pili (Tfp) mediate the movement of bacteria over surfaces without the use of flagella. These movements are known as social gliding in Myxococcus xanthus and twitching in organisms such as Pseudomonas aeruginosa and Neisseria gonorrhoeae. Tfp are localized polarly. Type IV pilins have a signature N-terminal domain, which forms a coiled-coil with other monomer units to polymerize a pilus fibre. At least 10 more proteins at the base of the fibre are conserved; they are related to the type II secretion system. Movements produced by Tfp range from short, jerky displacements to lengthy, smooth ones. Tfp also participate in cell-cell interactions, pathogenesis, biofilm formation, natural DNA uptake, auto-aggregation of cells and development. What is the means by which Tfp bring about the movement of cells?

Adenosine Triphosphatases↗

PubMed1999

[Bronchial carcinoma in young adults].

Lung cancer in young adults has been differently described in publications from various regions of the world, in respect of relative incidence, distribution of morphology and stages, and prognosis of the disease. We analysed retrospectively the data from the cancer registry of the Lungenklinik Heckeshorn in Berlin between 1986 and 1995 with regard to this topic. 106/4939 patients were 40 years old or younger (2.1%). The proportion of female patients was higher in the younger group (42 vs. 29%). Adenocarcinoma was the leading subtype in young patients (33 vs. 24%). The younger group showed a higher proportion of inoperable stages IIIB and IV for non-small cell lung cancer (71 vs. 46%). Younger patients with operable stages I to IIIA non-small cell lung cancer underwent surgery much more often than patients older than 40 years (90 vs. 49%), but for all patients with this morphology there was no difference in survival between younger and older patients (302 vs. 314 patients). We conclude that lung cancer presents a clearly different clinical picture in younger patients but not a better prognosis.

Adult↗

PubMed1999

Increased release of transforming growth factor (TGF)-beta1, TGF-beta2, and chemoattractant mediators in pneumonia.

Transforming growth factor-beta (TGF-beta), interleukin-8 (IL-8), and leukotrienes are potent neutrophil chemoattractants that are released in several lung diseases. There is limited information about the release of TGF-beta in bronchoalveolar lavage fluid (BALF) of patients with pneumonia. Furthermore, it is not clear if TGF-beta is differentially expressed in different lung diseases. The aim of our study was to compare the concentrations of TGF-beta1 and TGF-beta2 in the BALF of patients with pneumonia and other lung diseases. Furthermore, correlation of the TGF-beta levels with the concentration of chemoattractant mediators as well as with indicators of macrophage and granulocyte activation should be investigated. Patients with pneumonia, interstitial lung disease (ILD), or chronic obstructive pulmonary diseases (COPD) were included. Patients with ischemic heart disease without pulmonary involvement served as controls. The concentrations of TGF-beta1 and TGF-beta2, of the chemoattractant cytokine IL-8, of leukotriene B4, and of the leukotrienes C4, D4, and E4 were measured. Neutrophil elastase and granulocyte content (PMN) were used as markers for granulocyte activation, and neopterin was used as a marker for the activation of macrophages. Significantly elevated levels of TGF-beta1 (mean = 0.216 ng/ml, p < 0.01) were found in patients with microbiologically positive pneumonia but not in patients with ILD or COPD. A significant (p < 0.001) correlation was found between the TGF-beta1 concentrations and the IL-8 levels and the percentage of granulocytes (r = 0.76, and r = 0.44, respectively). Elevated TGF-beta2 concentrations were measured in the BALF of patients with pneumonia (mean = 1.4 ng/ml, p < 0.01) and with ILD. Pneumonia was also associated with increased concentrations of leukotrienes C4, D4, and E4 (mean = 91.61 pg/ml, p < 0.05) and leukotriene B4 (mean = 203.9 pg/ml, p < 0.01), significantly elevated levels of PMN elastase (mean = 2958.26 ng/ml, p < 0.01), and neopterin (mean = 0.42 nmol/L). Our results strongly suggest that different lung diseases do differ with regard to the released cytokines. TGF-beta1 probably plays a key role in regulation of pulmonary inflammation, particularly in pneumonia.

Bronchoalveolar Lavage Fluid↗

PubMed1999

The Myxococcus xanthus pilQ (sglA) gene encodes a secretin homolog required for type IV pilus biogenesis, social motility, and development.

The Myxococcus xanthus sglA1 spontaneous mutation was originally isolated because it allowed dispersed cell growth in liquid yet retained the ability to form fruiting bodies. Consequently, most of today's laboratory strains either contain the sglA1 mutation or were derived from strains that carry it. Subsequent work showed that sglA was a gene for social gliding motility, a process which is mediated by type IV pili. Here sglA is shown to map to the major pil cluster and to encode a 901-amino-acid open reading frame (ORF) that is homologous to the secretin superfamily of proteins. Secretins form a channel in the outer membrane for the transport of macromolecules. The closest homologs found were PilQ proteins from Pseudomonas aeruginosa and Neisseria gonorrhoeae, which are required for type IV pili biogenesis and twitching motility. To signify these molecular and functional similarities, we have changed the name of sglA to pilQ. The hypomorphic pilQ1 (sglA1) allele was sequenced and found to contain two missense mutations at residues 741 (G-->S) and 762 (N-->G). In addition, 19 independent social (S)-motility mutations are shown to map to the pilQ locus. In-frame deletions of pilQ and its downstream gene, orfL, were constructed. pilQ is shown to be essential for pilus biogenesis, S-motility, rippling, and fruiting body formation, while orfL is dispensable for these processes. The pilQ1 allele, but not the DeltapilQ allele, was found to render cells hypersensitive to vancomycin, suggesting that PilQ1 alters the permeability properties of the outer membrane. Many differences between pilQ1 and pilQ+ strains have been noted in the literature. We discuss some of these observations and how they may be rationalized in the context of our molecular and functional findings.

Amino Acid Sequence↗

PubMed1999

Gliding mutants of Myxococcus xanthus with high reversal frequencies and small displacements.

Myxococcus xanthus cells move on a solid surface by gliding motility. Several genes required for gliding motility have been identified, including those of the A- and S-motility systems as well as the mgl and frz genes. However, the cellular defects in gliding movement in many of these mutants were unknown. We conducted quantitative, high-resolution single-cell motility assays and found that mutants defective in mglAB or in cglB, an A-motility gene, reversed the direction of gliding at frequencies which were more than 1 order of magnitude higher than that of wild type cells (2.9 min-1 for DeltamglAB mutants and 2.7 min-1 for cglB mutants, compared to 0.17 min-1 for wild-type cells). The average gliding speed of DeltamglAB mutant cells was 40% of that of wild-type cells (on average 1.9 micrometers/min for DeltamglAB mutants, compared to 4.4 micrometers/min for wild-type cells). The mglA-dependent reversals and gliding speeds were dependent on the level of intracellular MglA protein: mglB mutant cells, which contain only 15 to 20% of the wild-type level of MglA protein, glided with an average reversal frequency of about 1.8 min-1 and an average speed of 2.6 micrometers/min. These values range between those exhibited by wild-type cells and by DeltamglAB mutant cells. Epistasis analysis of frz mutants, which are defective in aggregation and in single-cell reversals, showed that a frzD mutation, but not a frzE mutation, partially suppressed the mglA phenotype. In contrast to mgl mutants, cglB mutant cells were able to move with wild-type speeds only when in close proximity to each other. However, under those conditions, these mutant cells were found to glide less often with those speeds. By analyzing double mutants, the high reversing movements and gliding speeds of cglB cells were found to be strictly dependent on type IV pili, encoded by S-motility genes, whereas the high-reversal pattern of mglAB cells was only partially reduced by a pilR mutation. These results suggest that the MglA protein is required for both control of reversal frequency and gliding speed and that in the absence of A motility, type IV pilus-dependent cell movement includes reversals at high frequency. Furthermore, mglAB mutants behave as if they were severely defective in A motility but only partially defective in S motility.

Bacterial Proteins↗

PubMed1999

Apoptosis induced by lack of hemodynamic forces is a general endothelial feature even occuring in immortalized cell lines.

Confluent monolayers of primary endothelial cells display a high viability and an apparently constant cell density. However upon prolonged cultivation the monolayer degenerates with increasing numbers of senescent cells finally representing the whole culture. Recently we showed that lack of hemodynamic forces induces apoptosis in organ cultures as well as in confluent monolayers of human umbilical cord vein endothelial cells (HUVEC). The apoptosis started at a low level and was counteracted by a continuous proliferation of the remaining cells. Here we show that the induction of apoptosis by lack of hemodynamic forces is a general characteristic of vascular endothelial cells, valid for endothelial cells from various organs and species: human umbilical cord vein endothelial cells (HUVEC), human microvascular placental endothelial cells (HPEC) and bovine aorta endothelial cells (BAEC). Furthermore apoptosis due to the lack of hemodynamic forces can also be induced in various endothelial cell lines: EA.hy 926 derived from HUVEC and PBMEC-A1 derived from PBMEC. However degeneration of confluent monolayers does not occur with these cell lines even in monolayers kept for several weeks. This indicates that the degeneration of normal endothelial cell monolayers is caused by depletion of the proliferation potential of the endothelial cells.

Animals↗

PubMed1998

The guanosine nucleotide (p)ppGpp initiates development and A-factor production in myxococcus xanthus.

Guanosine 3'-di-5'-(tri)di-phosphate nucleotides [(p)ppGpp], synthesized in response to amino acid limitation, induce early gene expression leading to multicellular fruiting body formation in Myxococcus xanthus. A mutant (DK527) that fails to accumulate (p)ppGpp in response to starvation was found to be blocked in development prior to aggregation. By use of a series of developmentally regulated Tn5lac transcriptional fusion reporters, the time of developmental arrest in DK527 was narrowed to within the few hours of development, the period of starvation recognition. The mutant is also defective in the production of A-factor, an early extracellular cell-density signal. The relA gene from Escherichia coli, which encodes a ribosome-dependent (p)ppGpp synthetase, rescues this mutant. We also demonstrate that inactivation of the M. xanthus relA homolog blocks development and the accumulation of (p)ppGpp. Moreover, the wild-type allele of Myxococcus relA rescues DK527. These observations support a model in which accumulation of (p)ppGpp, in response to starvation, initiates the program of fruiting body development, including the production of A-factor.

4-Butyrolactone↗

PubMed1998

Alignment enhances the cell-to-cell transfer of pilus phenotype.

Social gliding motility of Myxococcus xanthus requires polar type IV pili. Tgl mutants lack pili and lack social motility. However, both defects can be rescued phenotypically, but not genotypically, when tgl+ donor and tgl- recipient cells make physical contact with each other. What is the cellular and molecular basis of this transfer of phenotype, which is called stimulation? Stimulation does not occur in liquid nor in soft (0.5%) agar; however, on a more firm surface (1.0% agar) cells stimulate each other efficiently. Microscopy revealed that cells placed on 1.0% agar readily became aligned whereas they remained poorly aligned on 0.5% agar. It was observed, moreover, that stimulation is greatly reduced when donor and recipient cells lack the ability to move because of mutations in any of several different motility genes. Microscopy showed that motile cells became highly aligned, and can even align nonmotile cells. Using a stimulation assay based on the assembly of pili, a strong correlation was found between conditions that promote cell alignment and the speed or extent of stimulation. Because pili are assembled only at the end(s) of a cell, and a parallel alignment of cells in contact brings them end-to-end, we would suggest that end-to-end contacts are important for stimulation of pilus assembly.

Bacterial Proteins↗

PubMed1998

Correlation of femoral and lumbar DXA and calcaneal ultrasound, measured in situ with intact soft tissues, with the in vitro failure loads of the proximal femur.

The objective of this study was to determine experimentally the sex-specific correlation of femoral and lumbar DXA and calcaneal ultrasound, measured in situ, with the in vitro failure loads of the proximal femur. Fifty-eight cadavers with intact skin and soft tissues (34 male, aged 81.2 +/- 8.7 years; 24 female, aged 83.7 +/- 10.6 years) were examined. The bone mass of the proximal femur and the lumbar spine were determined using dual-energy X-ray absorptiometry and the ultrasonic properties of the calcaneus with quantitative ultrasound. Afterwards, the right femora were excised 18 cm distal to the minor trochanter, and their load to failure determined with a material testing machine. Femoral fracture loads were significantly higher in males than in females, both before and after correcting for body height and weight. Femoral neck bone mineral density (BMD) was significantly correlated with femoral failure loads (r = 0.65 all specimens, 0.57 males (0.64 after excluding trochanteric fractures) and 0.77 females; p < 0.001). The correlations with the ultrasonic Stiffness Index of the calcaneus were in a similar range (r = 0.67 all specimens, 0.48 males (0.64 after excluding trochanteric fractures) and 0.65 females; p < 0.001). The correlations between femoral failure loads and the spinal BMD were lower (r = 0.40, p < 0.01), particularly in males (r = 0.30, not significant). In contrast to previous experimental investigations on excised bones, our results are consistent with clinical studies that have reported that ultrasound and femoral DXA have a similar ability to predict the risk of hip fracture.

Absorptiometry, Photon↗

PubMed1998

How and why myxobacteria talk to each other.

In response to starvation, myxobacteria build multicellular fruiting bodies that contain many thousands of cells and that have particular species-specific shapes. To coordinate fruiting body development, the myxobacterial cells signal to each other. The timing of and cellular responses to these signals help to give form to the fruiting body. Following identification of several signal molecules, important transcriptional regulators and other signals have recently been identified. Steps on signal transduction pathways have also been defined.

Bacterial Proteins↗

PubMed1998

Prediction of vertebral failure loads from spinal and femoral dual-energy X-ray absorptiometry, and calcaneal ultrasound: an in situ analysis with intact soft tissues.

The objective of the current study was to determine the correlation of spinal and femoral dual-energy X-ray absorptiometry (DXA) and calcaneal ultrasound, measured in situ with intact soft tissues, with the in vitro failure loads of lumbar vertebral bodies. Forty-nine cadavers with intact skin and soft tissues (32 men aged 82.1 +/- 9.0 years, 17 women aged 83.1 +/- 10.1 years) were examined. The bone mineral content (BMC), the projectional area, and the bone mineral density (BMD) of the lumbar spine and proximal femur were determined with DXA, and the ultrasonic properties of the calcaneus with quantitative calcaneal ultrasound. The fourth lumbar vertebra was then excised with adjacent intervertebral disks and its mechanical failure load determined, using a materials testing machine. Absolute fracture loads were significantly higher in men than in women, but they were similar after adjusting for body weight and height. Spinal DXA was significantly associated with vertebral failure load (r = 0.62 combined; r = 0.54 men; r = 0.58 women). Femoral DXA (r = 0.46) and calcaneal ultrasound (r = 0.48) showed somewhat lower correlation coefficients, with the speed of sound (SOS) being able to add predictive information in a stepwise regression model. Normalizing the vertebral failure loads to body weight and height reduced the correlations, with only spinal DXA yielding a significant relationship. Our data suggest that previous in vitro studies may have overestimated the association between spinal DXA and vertebral failure loads, presumably because measurements were performed on excised bones, but not in situ in the presence of soft tissue inhomogeneity. The results indicate that, even in a population of old age and under in situ conditions, spinal DXA may still be somewhat better than femoral DXA and calcaneal ultrasound in predicting vertebral failure loads.

Absorptiometry, Photon↗

PubMed1998

The pilH gene encodes an ABC transporter homologue required for type IV pilus biogenesis and social gliding motility in Myxococcus xanthus.

Type IV pilus genes have been shown to be required for social gliding motility in Myxococcus xanthus. We report the discovery of four additional pil genes: pilD, a homologue of type IV prepilin leader peptidases; and pilG, pilH and pilI, which have no known homologues in other type IV pilus systems. pilH encodes an ATP-binding cassette (ABC) transporter homologue, the first such homologue to be required for the biogenesis of any bacterial pilus type. pilG and pilI are co-transcribed with pilH and appear to be functionally related to pilH. Null mutants of pilG, pilH and pilI all lack social motility, are deficient in pilus production, have elevated sporulation efficiencies and display similar developmental abnormalities. In addition, all three mutations reduced the amount of PilA found in the supernatant after cells were sedimented from liquid culture. We suggest that the products of these three genes form a single ABC exporter complex, in which pilI is an integral membrane protein with membrane-spanning domains, and pilG is an accessory factor. The complex may participate in pilus assembly and/or the export of PilA pilin.

ATP-Binding Cassette Transporters↗

PubMed1998

Targeted mutagenesis of sigma54 activator proteins in Myxococcus xanthus.

Myxococcus xanthus DNA segments related to the highly conserved central sequence of sigma54 activator proteins have been investigated. A genetic technique designed to inactivate a gene that encodes such an activator by inserting a plasmid-borne internal fragment of the putative gene has been tested. When the internal fragment inserted by homologous recombination into the corresponding chromosomal locus, the expected duplication of the gene was observed by Southern hybridization. The single restriction fragment characteristic of each segment was replaced in the insertion strains by two hybridizing fragments, and one of these fragments hybridized with the kanamycin resistance gene of the plasmid vector. The combined molecular weights of the two fragments from the insertion strains were equal to the molecular weight of the original fragment plus the expected molecular weight contributed by the vector. In the duplication, one copy is expected to have an N-terminal deletion and the other copy is expected to have a C-terminal deletion. In most cases, the net result should be loss of activator function. If an activator is essential for vegetative growth, then it should not be possible to obtain the insertion strain by plasmid integration. Indeed, integrants for three of the segments were not obtained in repeated trials; however, a plausible explanation for these results other than lethality can be offered. Of the seven insertions validated by Southern hybridization, four strains exhibited defects in the development of fruiting bodies. One of these failed to develop in submerged culture, though it developed normally on agar. The other three showed arrested development of fruiting bodies, each at a morphologically different stage of aggregation. One of the mutants may be defective in the reception pathway of A-signal.

Bacterial Proteins↗

PubMed1998

Contact stimulation of Tgl and type IV pili in Myxococcus xanthus.

Myxococcus xanthus tgl mutants lack social motility and type IV pili but can be transiently stimulated to swarm and to make pili by contacting tgl+ cells. The absence of pili in tgl mutants is shown not to be due to the absence of pilin. The rate of pilus elongation after Tgl stimulation is shown to be similar to the rate of pilus elongation in wild-type cells, using a new more rapid assay for stimulation.

Bacterial Proteins↗

PubMed1997

sigma54, a vital protein for Myxococcus xanthus.

The rpoN gene encoding the transcription factor sigma54 in Myxococcus xanthus has been cloned using a heterologous rpoN probe. The sequence of the cross-hybridizing DNA confirmed the existence of an ORF 1518 bp long that encodes a well conserved member of the sigma54 family of sigma factors. Low- as well as high-stringency hybridizations detected only a single rpoN gene in the M. xanthus chromosome. In other bacteria, sigma54 is an alternative sigma, and null mutants are viable. However, all attempts to construct a strain containing a null mutation in the M. xanthus rpoN have been unsuccessful. Partial diploids of rpoN+/rpoN null are viable. Recombination experiments with such partial diploids showed the impossibility of constructing, either by segregation or by transduction, a viable null haploid under any of a wide range of growth conditions. The product of the rpoN gene, sigma54, therefore appears to be essential for growth in M. xanthus.

Amino Acid Sequence↗