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R Baumeister

Publications and source records attributed to R Baumeister.

At least 37 records · Page 2Linked to original sources
PubMed1998

Caspase-mediated cleavage is not required for the activity of presenilins in amyloidogenesis and NOTCH signaling.

The Alzheimer's disease (AD) associated presenilin (PS) proteins are proteolytically processed. One of the processing pathways involves cleavage by caspases. Pharmacological inhibition of caspases is currently being discussed as a treatment for a variety of neurodegenerative diseases, including AD. We therefore inhibited caspase mediated processing of PS-1 and PS-2 in cells transfected with wt and mutant PS by mutagenizing the substrate recognition site or by using specific peptide aldehydes known to block caspases. We found that the inhibition of caspase mediated processing of PS proteins does not decrease its amyloidogenic activity. PS cDNA constructs with mutations in the caspase cleavage site are biologically active in Caenorhabditis elegans such as the wt human PS proteins, demonstrating that caspase-mediated cleavage is not required for the physiological PS function in NOTCH signaling.

Alzheimer Disease↗
PubMed1998

Proteolytic processing of Alzheimer's disease associated proteins.

Amyloid beta-peptide (A beta), the major component of senile plaques, is generated by proteolytic processing from the beta-amyloid precursor protein (beta APP). Mutations within the beta APP gene cause early onset familial AD (FAD) by affecting A beta generation. Interestingly, the much more abundant mutations within the presenilin (PS) genes also result in the abnormal generation of a 42 residue A beta (A beta 42), thus clearly supporting a pivotal role of A beta for the pathology of AD. PS proteins are proteolytically processed into stable 30 kDa N-terminal fragments (NTF) and 20 kDa C-terminal fragments (CTF). Beside the conventional proteolytic pathway. PS proteins can also be cleaved further C-terminal by proteases of the caspase superfamily. PS proteins were localized within the endoplasmic reticulum (ER) and early Golgi, compartments which we have demonstrated to be involved in A beta 42 generation and intracellular accumulation. Using Caenorhabditis elegans as a simple animal model, we demonstrate that PS proteins are involved in NOTCH signaling FAD causing mutations interfere with the biological function of PS proteins in NOTCH signaling.

Alzheimer Disease↗
PubMed1998

What do we learn from a few familial Alzheimer's disease cases?

Alzheimer's disease is the most common form of dementia. About 90% of the cases occur sporadically whereas in 10% of the cases mutations were found within three different genes. Mutations in the gene encoding the beta-Amyloid precursor protein (beta APP) are located in the ultimate neighborhood of the three proteases (secretases) involved in proteolytic processing of beta APP. These mutations cause an increased production of the long form of Amyloid beta-peptide (A beta) the major component of Amyloid plaques. In contrast to the 40 amino acid form (A beta 40), the 42 amino acid form (A beta 42) aggregates more rapidly, kills cultured neurons more efficiently, and precipitates preferentially in amyloid plaques. Interestingly, mutations in the Presenilin genes which are responsible for more then 40% of all familial AD cases also cause enhanced production of the elongated form of A beta. Therefore mutations in three different genes directly effect A beta production in a pathological manner, which strongly supports the amyloid cascade hypothesis.

Alzheimer Disease↗
PubMed1998

Group A streptococcal toxic shock syndrome with severe necrotizing fasciitis following hysterectomy--a case report.

In the last 10 years an increasing number of cases of group A streptococcal toxic shock syndrome have appeared in various clinical settings. The manifestation of this syndrome includes rapidly progressive multiorgan failure and soft-tissue necrosis. This report presents a case of streptococcal toxic shock syndrome caused by Streptococcus pyogenes with severe necrotizing fasciitis of the abdominal wall following hysterectomy. Aggressive surgical intervention with debridement of all necrotic tissue necessitated resection of the complete abdominal wall (skin, subcutaneous tissue, muscle and peritoneum). The abdominal wall defect was covered with free myocutaneous flaps and split-skin grafts. Optimal treatment, including adequate antibiotic therapy and radical surgical intervention, is an indispensable prerequisite of successful outcome.

Fasciitis, Necrotizing↗
PubMed1997

Human presenilin-1, but not familial Alzheimer's disease (FAD) mutants, facilitate Caenorhabditis elegans Notch signalling independently of proteolytic processing.

The majority of cases with familial Alzheimer's disease (FAD) are linked to mutations of the presenilin (PS) genes. These genes show considerable sequence similarity to the sel-12 gene of Caenorhabditis elegans, which has been postulated to function in the facilitated signalling by lin-12 and glp-1. In order to analyse the functional conservation of the presenilins, we introduced the human PS-1 cDNA, as well as clinical and deletion mutant proteins, into sel-12 mutant animals and tested their potential to rescue the egg-laying defect. Human PS-1 expressed from the sel-12 promoter fully rescued the sel-12 phenotype, whereas two missense mutations, C410Y and A246E, identified in pedigrees with FAD, exhibited a strongly decreased rescuing activity. The large hydrophilic loop and transmembrane domain 7 are required for the biological activity of PS-1. PS-1 protein was proteolytically cleaved in C. elegans as it is in human cells. A PS-1 splice variant (FAD mutation deltaexon9) that does not undergo proteolytic cleavage also substituted for sel-12. The conservation of function of human PS-1 and C. elegans sel-12 suggests that presenilin proteins are required, directly or indirectly, for the proper operation of the Notch signalling pathway. FAD-associated mutant proteins tested showed different rescuing activities, indicating that they might affect different functional or regulatory aspects of PS-1. Proteolytic processing is not a prerequisite for PS-1 function in C. elegans.

Alzheimer Disease↗
PubMed1996

Lineage-specific regulators couple cell lineage asymmetry to the transcription of the Caenorhabditis elegans POU gene unc-86 during neurogenesis.

The POU homeo box gene unc-86 specifies neuroblast and neural identities in the developing Caenorhabditis elegans nervous system. After an asymmetric neuroblast division, unc-86 is expressed in one of two daughter cells in 27 lineage classes that are not obviously related by function or position. We show here that unc-86 transcriptional regulatory regions detect cell lineage asymmetry to activate unc-86 expression in one of two neuroblast daughter cells. Distinct regulatory regions activate unc-86 expression in particular sets of sublineages. Therefore the unc-86 regulatory region integrates distinct cell lineage asymmetry cues to activate unc-86 expression in the many classes of neuroblast cell lineages. In agreement with such lineage-specific regulation of unc-86 asymmetric activation, mutations in lin-11 (LIM homeo box), ham-1, and lin-17 affect the asymmetry of unc-86 expression in particular cell lineages, and mutations in lin-32 (achaete/scute family), vab-3 (Pax-6 homolog) and egl-5 (Abd-B homolog) affect the establishment of unc-86 expression in other cell lineages. Homologs of unc-86 and many of these unc-86 regulators have been implicated in control of neurogenesis in vertebrates and invertebrates. These data suggest that unc-86 acts in a phylogenetically conserved pathway that couples neuroblast cell lineage asymmetry to the generation of diverse neural types.

Animals↗
PubMed1994

In vivo fluorescence microscopy for the assessment of microvascular reperfusion injury in small bowel transplants in rats.

With the use of in vivo fluorescence microscopy we have analyzed microvascular reperfusion injury of small bowel isograft transplants in rats. Following 1 hr cold storage in University of Wisconsin solution, the small bowel was transplanted heterotopically, and the intestinal microcirculation was quantitatively analyzed 20-60 min after onset of reperfusion. The intestinal grafts' capillary perfusion of both the mucosa and the circular and longitudinal muscles was not found altered when compared with the intestinal capillary perfusion of sham-operated controls. In contrast, leukocyte-endothelial cell interaction, including leukocyte rolling (40 +/- 5%) and sticking (280 +/- 100 mm-2) in submucosal postcapillary venules, was significantly increased when compared with nontransplanted controls (12 +/- 8% and 20 +/- 10 mm-2, P < 0.01 and P < 0.05, respectively). Leukocyte-endothelial cell interaction was associated with a marked alteration of lymphatic capillary drainage, as indicated by the low functional density of lymphatic microvessels of 10.2 +/- 6.1 cm-1 (P < 0.01 vs. sham-operated controls (39.2 +/- 6.1 cm-1)). From these results we propose that leukocyte-endothelial cell interaction, not capillary "no-reflow," is the primary step in the manifestation of microvascular reperfusion injury following a short period of cold ischemia in small bowel grafts.

Animals↗
PubMed1992

Contacts between Tet repressor and tet operator revealed by new recognition specificities of single amino acid replacement mutants.

We have analyzed the DNA binding properties of Tet-repressor mutants with single amino acid residue replacements at eight positions within the alpha-helix-turn-alpha-helix DNA-binding motif. A saturation mutagenesis of Gln38, Pro39, Thr40, Tyr42, Trp43 and His44 in the second alpha-helix was performed; in addition, several substitutions of Thr27 and Arg28 in the first alpha-helix were constructed. The abilities of these mutant repressors to bind a set of 16 operator variants were determined and revealed 23 new binding specificities. All repressor mutants with DNA-binding activity were inducible by tetracycline, while mutants lacking binding activity were trans-dominant over the wild-type. All mutant proteins were present at the same intracellular steady-state concentrations as the wild-type. These results suggest the structural integrity of the mutant repressors. On the basis of the new recognition specificities, five contacts between a repressor monomer and each operator half-site and the chemical nature of these repressor-operator interactions are proposed. We suggest that Arg28 contacts guanine of the G.C base-pair at operator position 2 with two H-bonds, Gln38 binds adenine of the A.T base-pair at position 3 with two H-bonds, and the methyl group of Thr40 participates in a van der Waals' contact with cytosine of the G.C base-pair at position 6 of tet operator. A previously unrecognized type of interaction is proposed for Pro39, which inserts its side-chain between the methyl groups of the thymines of T.A and A.T base-pairs at positions 4 and 5. Computer modeling of these proposed contacts reveals that they are possible using the canonical structures of the helix-turn-helix motif and B-DNA. These contacts suggest an inverse orientation of the Tet repressor helix-turn-helix with respect to the operator center as compared with non-inducible repressor-operator complexes, and are supported by similar contacts of other repressor-operator complexes.

Amino Acid Sequence↗
PubMed1992

Functional roles of amino acid residues involved in forming the alpha-helix-turn-alpha-helix operator DNA binding motif of Tet repressor from Tn10.

The Tn10 derived Tet repressor contains an amino acid segment with high homology to the alpha-helix-turn-alpha-helix motif (HTH) of other DNA binding proteins. The five most conserved amino acids in HTH are probably involved in structural formation of the motif. Their functional role was probed by saturation mutagenesis yielding 95 single amino acid replacement mutants of Tet repressor. Their binding efficiencies to tet operator were quantitatively determined in vivo. All functional mutants contain amino acid substitutions consistent with their proposed role in a HTH. In particular, only the two smallest amino acids (serine, glycine) can substitute a conserved alanine in the proposed first alpha-helix without loss of activity. The last position of the first alpha-helix, the second position in the turn, and the fourth position in the second alpha-helix require mostly hydrophobic residues. The proposed C-terminus of the first alpha-helix is supported by a more active asparagine compared to glutamine replacement mutant of the wt leucine residue. The turn is located close to the protein surface as indicated by functional lysine and arginine replacements for valine. A glycine residue at the first position in the turn can be replaced by any amino acid yielding mutants with at least residual tet operator affinity. A structural model of the HTH of Tet repressor is presented.

Amino Acid Sequence↗
PubMed1992

The Tn10-encoded tetracycline resistance mRNA contains a translational silencer in the 5' nontranslated region.

We performed a mutational analysis of the left half of Tn10-encoded tet operator O2, located in the 5' nontranslated region of the mRNA for the resistance protein TetA, and determined the importance of that region for translation efficiency and mRNA stability. Transcriptional fusions of 17 mutants to lacZ expressed the same amounts of beta-galactosidase, while translational fusions varied 35-fold in expression efficiency. The mRNA half-lives varied 24-fold, with 9.6 min for the most highly expressed mRNA and 0.4 min for the least efficiently expressed mRNA. Toeprint experiments were performed to distinguish whether these mutations define a determinant of mRNA stability or influence translation initiation. The highly expressed mRNA was 24-fold more efficient in forming the initiation complex in vitro than the low-expression mutant. It was concluded that this sequence, albeit located upstream of the ribosome-binding sequence, is an important determinant for efficient initiation of translation. Secondary-structure calculations of the mRNAs revealed no correlation of the potential to form double strands masking the ribosome-binding sequence with expression efficiency.

Base Sequence↗
PubMed1991

Lack of a 5' non-coding region in Tn1721 encoded tetR mRNA is associated with a low efficiency of translation and a short half-life in Escherichia coli.

The repressor-encoding tetR gene from Tn1721 is expressed with a very low efficiency. Its mRNA lacks an untranslated leader sequence. We have constructed protein fusions with the lacZ gene which contain between 14 and 157 5' nucleotides from the tetR gene. Since they are all expressed with similar efficiencies we conclude that the sequence information for initiation of translation is contained within the first 14 bases of the tetR coding region. These fusion transcripts are about 20-fold less efficiently translated than the wild type lacZ transcript. A toeprint analysis confirms that the initiation complex is indistinguishable from those formed by regular transcripts with 5' untranslated regions but occurs in a very low amount in vitro. Thus, the absence of a 5' leader causes a poor rate of translation initiation. The half-lives of tetR and tetR-lacZ mRNAs are about 30 seconds, which is 3-times lower than that of the wt lacZ mRNA. Inactivation of the ams/rne locus in E. coli stabilizes the tetR transcript more than ten-fold. The influence of translation on the tetR half-life is discussed.

Base Sequence↗
PubMed1991

Amino acids determining operator binding specificity in the helix-turn-helix motif of Tn10 Tet repressor.

Each of 22 amino acids in the proposed alpha-helix-turn-alpha-helix operator binding motif of the Tn10 encoded Tet repressor was replaced by alanine and one residue was replaced by valine to determine their role in tet operator recognition by a 'loss of contact' analysis with 16 operator variants. One class of amino acids consisting of T27 and R28 in the first alpha-helix and L41, Y42, W43 and H44 in the recognition alpha-helix are quantitatively most important for wild-type operator binding. These residues are probably involved in the structural architecture of the motif. A second class of residues is quantitatively less important for binding, but determines specificity by forming base pair specific contacts to three positions in tet operator. This property is most clearly demonstrated for Q38 and P39 and to a lesser extent for T40 at the N-terminus of the recognition alpha-helix. The contacted operator base pairs indicate that the N-terminus of the recognition alpha-helix is located towards the palindromic center in the repressor-operator complex. Although the orientation of the recognition alpha-helix in the Tet repressor-tet operator complex is inversed as compared with the lambda- and 434 repressor-operator complexes, the reduced operator binding of the TA27 mutation in the first alpha-helix suggests that the hydrogen bonding networks connecting the two alpha-helices may be similar in these proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1991

Selection for Tn10 tet repressor binding to tet operator in Escherichia coli: isolation of temperature-sensitive mutants and combinatorial mutagenesis in the DNA binding motif.

We have constructed a genetic assay which selects positively for a functional interaction between Tet repressor and its cognate operator in Escherichia coli. In this strain Tet repressor blocks expression of lacI and lacZ. This leads to derepression of a lacPO controlled galK gene. The strain can be selected by growth on galactose as the sole carbon source and screened for the beta-galactosidase phenotype. These features allow the identification of one candidate among 10(8) false clones on a single plate. The assay was applied to select mutants with a ts DNA binding phenotype and to screen oligonucleotide generated Tet repressor mutants. Analysis of these mutations revealed that they affect DNA and inducer binding and possibly the dimerization domains. These mutations are located at residues 21, 48, 49, 89 and at the C terminus of the protein (193), respectively.

Amino Acid Sequence↗
PubMed1988

A threonine to alanine exchange at position 40 of Tet repressor alters the recognition of the sixth base pair of tet operator from GC to AT.

The tet operators of two naturally evolved tetracycline resistance determinants differ by a G.C to A.T transition at the sixth base pair. This mutation prevents heterologous recognition of these tet operators by their respective two Tet repressor proteins. The amino acid side chains responsible for this sequence-specific distinction of operators were determined. For this purpose in vitro recombinants of the two tetR genes were constructed. Restriction sites were introduced by oligonucleotide-directed mutagenesis in both genes followed by the exchange of different coding segments between them. The encoded chimeric Tet repressor proteins were expressed and their operator recognition specificity was scored in vivo. Exchanging gradually smaller coding segments led finally to a single amino acid exchange in both genes at position 40 of the primary structures. Each Tet repressor containing Thr at this position recognizes the G.C operator while those with Ala recognize the A.T operator regardless of the rest of the sequences. This result demonstrates clearly that the amino acid 40 of Tet repressor contacts and recognizes base pair 6 of tet operator. Sterical interference of the large Thr side chain with the methyl group of A.T and a possible involvement of the hydroxyl in hydrogen bonding to the operator are discussed as the molecular basis of this differentiation between A.T and G.C base pairs.

Amino Acid Sequence↗
PubMed1987

[Microsurgical reconstruction of large 3-layer defects of the cheek].

For reconstruction of large penetrating defects of the cheek besides pedicled tissue transfer numerous free flaps are available to the head and neck surgeon. The inferior epigastric abdominal wall flap, in particular, has proved a versatile and reliable flap, just as the cutaneous flaps from the dorsum of the foot and from the forearm. While the abdominal wall flap offers both, a cutaneous and a peritoneal surface, the latter is used for replacement of intra-oral lining, the cutaneous flaps have to be folded to obtain two surfaces for reconstruction of skin and intra-oral lining. The surgical technique of the epigastric abdominal wall flap is presented and its use is discussed.

Carotid Artery, External↗
PubMed1984

[Postnatal development of the width and height of the palate and the palate foramina].

During the postnatal development the anterior width of the palatum durum increases between the 2nd and 5th year more than the posterior palatal width. Also increases the height of the palatum durum and large during the 2nd dentition as to the distance of the foramen palatinum majus from the dens caninus . The size of the foramina palatina enlarges inconstantly during postnatal growth. In children there are 2.37, in adult 2.42 (mean) foramina palatina minora on the right side. Side differences are not significant.

Adolescent↗