Biomedical subjects

R Birkenhäger

Publications and source records attributed to R Birkenhäger.

15 recordsLinked to original sources

PubMed2006

[Evidence of a novel gene for the LAV-syndrome].

BACKGROUND: Both LAV- (large or enlarged vestibular aqueduct) and Pendred-syndrome are autosomal recessive diseases. In contrast to Pendred-syndrome, LAV-syndrome is characterised only by an enlarged vestibular aqueduct. Pendred-syndrome is a more complex disease. Classically it is characterised by sensorineural hearing loss and enlargement of the thyroid gland. Up to now, only mutations in SLC26A4 gene are known as being responsible for both syndromes. The gene for Pendred-syndrome (SLC26A4) has been localised by linkage analysis of chromosome 7q31. This protein is expressed in the inner ear, thyroid gland, kidney, and placenta. Functional analysis of the gene product (pendrin) in Xenopus laevis oocytes revealed that pendrin acts as an iodide/chloride and chloride/formate exchanger. METHOD: Each of the exons and flanking splice regions of the SLC26A4 gene were analysed by direct sequencing. Haplotype analysis was undertaken with microsatellite markers spanning a 5 Mbp area around the localisation of the SLC26A4 gene. RESULTS: In sequence analysis of 42 patients with bilateral enlargement of the vestibular aqueduct, no mutation could be identified in 30 % of cases. In some of these cases, a linkage to the gene localisation on chromosome 7q31 could not be detected. CONCLUSION: Our results indicate evidence for a second gene involved in the development of LAV-syndrome.

Adolescent↗

PubMed2006

[Pseudodominants of two recessive connexin mutations in non-syndromic sensorineural hearing loss?].

BACKGROUND: Hitherto more than hundred genes and gene loci for non-syndromic or syndromic deafness have been identified. Mutations in the connexin 26 gene (GJB2) account for up to 50 % of the cases of autosomal recessive hearing loss. The genes GJB2 (Connexin 26), GJB3 (connexin 31) and GJB6 (connexin 31) are located on chromosome 13q11-12. In the inner ear up to four different connexins are expressed. Connexins appertain to a group of gap junction proteins. These proteins can oligomerize to form single-membrane channels called connexons. Each connexon is composed of six subunits, that allow communication between adjacent cells by providing a channel for diffusion of ions, metabolites and second messengers. METHOD: Each of the exons and flanking splice regions of the connexin 26, 30, and 31 genes (GJB2, GJB3, and GJB6) have been analysed by direct sequencing. RESULTS: In the involved families three heterozygous mutations could be detected in the connexin 26 (GJB2) and connexin 30 (GJB6) genes. If a combination of two of those mutations occurs, 35DeltaG with 146/147DeltaC and 35DeltaG with GJB6-D13S1830 it results in hearing loss and deafness. CONCLUSION: By evidences of a familial background of hearing loss it is reasonable to analyse the connexin genes (GJB2, GJB3 and GJB6) for mutations, additionally to a specific hearing diagnostic, in order to enhance linguistic development through hearing aid or CI-implantation at an early stage.

Adult↗

PubMed2005

[The interesting case -- case no. 67].

BACKGROUND: Lipoma of the retropharyngeal space is a rare benign tumour often showing unspecific clinical symtoms. It can grow to an enormous extent causing total obstruction of the upper respiratory tract. Until now its etiology is unknown. With a variety of differential diagnoses, a diagnostic concept is necessary. CASE REPORT: A 41 year old male patient complained about a nondolent swelling of the neck. The radiological diagnostics showed a huge lipoma of the para- and retropharyngeal space with subtotal obstruction of the pharynx. The lipoma was removed completely via transcervical approach. CONCLUSION: Lipoma as differential diagnosis of retropharyngeal tumours always has to be considered. Surgical intervention is recommended. To prevent functional complications resulting from tumour and surgery and to get information about the extent of the lipoma, accurate radiological imaging is mandatory.

Adult↗

PubMed2004

[Identification of two heterozygous mutations in the SLC26A4/PDS gene in a family with Pendred-syndrome].

BACKGROUND: Pendred-syndrome is an autosomal recessive disease that is classically characterised by sensorineural hearing loss and enlargement of the thyroid gland. The gene SLC26A4/PDS for the pendred-syndrome has been localised by linkage analysis on chromosome 7q31. This protein is expressed in the inner ear, thyroid gland, kidney and placenta. Functional analysis in Xenopus laevis oocytes revealed that it acts as an iodide/chloride and chloride/formate exchanger. METHOD: Each of the exons and flanking splice regions of the SLC26A4/PDS gene was analysed by direct sequencing. RESULTS: In the involved family two heterozygous mutations could be detected which results by combination in hearing loss and deafness. CONCLUSION: By evidences of familial background in hearing loss and thyroid disorder it is reasonable to analyse the PDS gene for mutation to have early the possibility for medical care of linguistic development through hearing aid or CI-implantation.

Child↗

PubMed2001

Barttin is a Cl- channel beta-subunit crucial for renal Cl- reabsorption and inner ear K+ secretion.

Renal salt loss in Bartter's syndrome is caused by impaired transepithelial transport in the loop of Henle. Sodium chloride is taken up apically by the combined activity of NKCC2 (Na+-K--2Cl- cotransporters) and ROMK potassium channels. Chloride ions exit from the cell through basolateral ClC-Kb chloride channels. Mutations in the three corresponding genes have been identified that correspond to Bartter's syndrome types 1-3. The gene encoding the integral membrane protein barttin is mutated in a form of Bartter's syndrome that is associated with congenital deafness and renal failure. Here we show that barttin acts as an essential beta-subunit for ClC-Ka and ClC-Kb chloride channels, with which it colocalizes in basolateral membranes of renal tubules and of potassium-secreting epithelia of the inner ear. Disease-causing mutations in either ClC-Kb or barttin compromise currents through heteromeric channels. Currents can be stimulated further by mutating a proline-tyrosine (PY) motif on barttin. This work describes the first known beta-subunit for CLC chloride channels and reveals that heteromers formed by ClC-K and barttin are crucial for renal salt reabsorption and potassium recycling in the inner ear.

Absorption↗

PubMed2001

Mutation of BSND causes Bartter syndrome with sensorineural deafness and kidney failure.

Antenatal Bartter syndrome (aBS) comprises a heterogeneous group of autosomal recessive salt-losing nephropathies. Identification of three genes that code for renal transporters and channels as responsible for aBS has resulted in new insights into renal salt handling, diuretic action and blood-pressure regulation. A gene locus of a fourth variant of aBS called BSND, which in contrast to the other forms is associated with sensorineural deafness (SND) and renal failure, has been mapped to chromosome 1p. We report here the identification by positional cloning, in a region not covered by the human genome sequencing projects, of a new gene, BSND, as the cause of BSND. We examined ten families with BSND and detected seven different mutations in BSND that probably result in loss of function. In accordance with the phenotype, BSND is expressed in the thin limb and the thick ascending limb of the loop of Henle in the kidney and in the dark cells of the inner ear. The gene encodes a hitherto unknown protein with two putative transmembrane alpha-helices and thus might function as a regulator for ion-transport proteins involved in aBS, or else as a new transporter or channel itself.

Animals↗

PubMed1999

F0 complex of the Escherichia coli ATP synthase. Not all monomers of the subunit c oligomer are involved in F1 interaction.

The antigenic determinants of mAbs against subunit c of the Escherichia coli ATP synthase were mapped by ELISA using overlapping synthetic heptapeptides. All epitopes recognized are located in the hydrophilic loop region and are as follows: 31-LGGKFLE-37, 35-FLEGAAR-41, 36-LEGAAR-41 and 36-LEGAARQ-42. Binding studies with membrane vesicles of different orientation revealed that all mAbs bind to everted membrane vesicles independent of the presence or absence of the F1 part. Although the hydrophilic region of subunit c and particularly the highly conserved residues A40, R41, Q42 and P43 are known to interact with subunits gamma and epsilon of the F1 part, the mAb molecules have no effect on the function of F0. Furthermore, it could be demonstrated that the F1 part and the mAb molecule(s) are bound simultaneously to the F0 complex suggesting that not all c subunits are involved in F1 interaction. From the results obtained, it can be concluded that this interaction is fixed, which means that subunits gamma and epsilon do not switch between the c subunits during catalysis and furthermore, a complete rotation of the subunit c oligomer modified with mAb(s) along the stator of the F1F0 complex, proposed to be composed of at least subunits b and delta, seems to be unlikely.

ATP Synthetase Complexes↗

PubMed1998

Cooperative modulation by eIF4G of eIF4E-binding to the mRNA 5' cap in yeast involves a site partially shared by p20.

Interaction between the mRNA 5'-cap-binding protein eIF4E and the multiadaptor protein eIF4G has been demonstrated in all eukaryotic translation assemblies examined so far. This study uses immunological, genetic and biochemical methods to map the surface amino acids on eIF4E that contribute to eIF4G binding. Cap-analogue chromatography and surface plasmon resonance (SPR) analyses demonstrate that one class of mutations in these surface regions disrupts eIF4E-eIF4G association, and thereby polysome formation and growth. The residues at these positions in wild-type eIF4E mediate positive cooperativity between the binding of eIF4G to eIF4E and the latter's cap-affinity. Moreover, two of the mutations confer temperature sensitivity in eIF4G binding to eIF4E which correlates with the formation of large numbers of inactive ribosome 80S couples in vivo and the loss of cellular protein synthesis activity. The yeast 4E-binding protein p20 is estimated by SPR to have a ten times lower binding affinity than eIF4G for eIF4E. Investigation of a second class of eIF4E mutations reveals that p20 shares only part of eIF4G's binding site on the cap-binding protein. The results presented provide a basis for understanding how cycling of eIF4E and eIF4G occurs in yeast translation and explains how p20 can act as a fine, but not as a coarse, regulator of protein synthesis.

Amino Acid Sequence↗

PubMed1998

Topology of subunit a of the Escherichia coli ATP synthase.

The antigenic determinants of mAbs against subunit a of the Escherichia coli ATP synthase were mapped by ELISA using overlapping synthetic decapeptides. For two of the mAbs the epitopes are E4NMTPQD10 (GDH 14-5C6) and V29DPQ32 (GDH 8-8B3). Binding of these mAbs to membrane vesicles of different orientation revealed that both epitopes are accessible in vesicles with inside-out orientation. These results demonstrate that at least the N-terminal amino acids 1-32 of subunit a are located at the cytoplasmic side of the membrane. A further determination of the topology of subunit a was performed by inserting the reporter epitope DYKDDDDK (FLAG epitope) at different positions of the polypeptide chain. 10 of 13 insertions led to a functional F0F1 ATP synthase and allowed specific detection of the modified subunit a by immunoblotting using an mAb against the FLAG epitope. In addition, polyclonal anti-FLAG IgG was applied for the recognition of the mutant FLAG epitope DYKDDVDK. Cells carrying this mutant FLAG epitope at the C terminus of subunit a were able to grow on succinate as sole carbon and energy source, revealing a functional ATP synthase, in contrast to those carrying the original FLAG epitope at the same position. Binding studies with membrane vesicles of different orientation and anti-FLAG Ig demonstrated that both termini of the protein are located at the cytoplasmic side of the membrane, indicating that an even number of membrane-spanning segments is present in subunit a. In addition, insertion of two FLAG epitopes in tandem after K66, or one epitope after H95, and Q181 revealed that the polypeptide regions including these residues are accessible from the cytoplasmic surface of the membrane. These results support the view that the polypeptide chain of subunit a traverses the membrane six times.

Amino Acid Sequence↗

PubMed1997

VEGF and VEGF-C: specific induction of angiogenesis and lymphangiogenesis in the differentiated avian chorioallantoic membrane.

The lymphangiogenic potency of endothelial growth factors has not been studied to date. This is partially due to the lack of in vivo lymphangiogenesis assays. We have studied the lymphatics of differentiated avian chorioallantoic membrane (CAM) using microinjection of Mercox resin, semi- and ultrathin sectioning, immunohistochemical detection of fibronectin and alpha-smooth muscle actin, and in situ hybridization with VEGFR-2 and VEGFR-3 probes. CAM is drained by lymphatic vessels which are arranged in a regular pattern. Arterioles and arteries are accompanied by a pair of interconnected lymphatics and form a plexus around bigger arteries. Veins are also associated with lymphatics, particularly larger veins, which are surrounded by a lymphatic plexus. The lymphatics are characterized by an extremely thin endothelial lining, pores, and the absence of a basal lamina. Patches of the extracellular matrix can be stained with an antibody against fibronectin. Lymphatic endothelial cells of differentiated CAM show ultrastructural features of this cell type. CAM lymphatics do not possess mediae. In contrast, the lymphatic trunks of the umbilical stalk are invested by a single but discontinuous layer of smooth muscle cells. CAM lymphatics express VEGFR-2 and VEGFR-3. Both the regular pattern and the typical structure of these lymphatics suggest that CAM is a suitable site to study the in vivo effects of potential lymphangiogenic factors. We have studied the effects of VEGF homo- and heterodimers, VEGF/PlGF heterodimers, and PlGF and VEGF-C homodimers on Day 13 CAM. All the growth factors containing at least one VEGF chain are angiogenic but do not induce lymphangiogenesis. PlGF-1 and PlGF-2 are neither angiogenic nor lymphangiogenic. VEGF-C is the first lymphangiogenic factor and seems to be highly chemoattractive for lymphatic endothelial cells. It induces proliferation of lymphatic endothelial cells and development of new lymphatic sinuses which are directed immediately beneath the chorionic epithelium. Our studies show that VEGF and VEGF-C are specific angiogenic and lymphangiogenic growth factors, respectively.

Actins↗

PubMed1996

Schizosaccharomyces pombe has a novel eukaryotic initiation factor 4F complex containing a cap-binding protein with the human eIF4E C-terminal motif KSGST.

Genetic and biochemical analyses were performed on the cytoplasmic cap-binding complex (eukaryotic initiation factor (eIF) 4F) of Schizosaccharomyces pombe. Genomic and cDNA sequencing of the S. pombe gene (tif1) encoding the cap-binding component eIF4E revealed the presence of two introns in a reading frame of 219 codons. The encoded sequence of 218 amino acids shows a greater degree of identity to the mammalian eIF4E sequence than does its counterpart from Saccharomyces cerevisiae. In particular, unlike its S. cerevisiae counterpart, S.pombe eIF4E has a C-terminal Ser209 within the motif KSGST that is a site of phosphorylation in hamster and rabbit eIF4E. Of relevance to its potential regulatory role, eIF4E was found to be encoded by an mRNA with a six-nucleotide leader and to be of low abundance in vivo. Cross-linking experiments identified S. pombe eIF4E as the major cap-binding protein while a further protein, p36, also showed cap-dependent binding. eIF4A was not associated with the cap-binding complex. While S. pombe eIF4E was shown capable of binding S. cerevisiae p20, an equivalent protein was absent from the eIF4F complex isolated from S. pombe cells. S. pombe 4F therefore shows a remarkable combination of structural and functional properties, some of which it shares with its higher and its lower eukaryotic counterparts.

Amino Acid Sequence↗

PubMed1996

Synthesis and physiological activity of heterodimers comprising different splice forms of vascular endothelial growth factor and placenta growth factor.

Vascular endothelial growth factor (VEGF) and placenta growth factor (PIGF) are members of a dimeric-growth-factor family with angiogenic properties. VEGF is a highly potent and specific mitogen for endothelial cells, playing a vital role in angiogenesis in vivo. The role of PIGF is less clear. We expressed the monomeric splice forms VEGF-165, VEGF-121, PIGF-1 and PIGF-2 as unfused genes in Escherichia coli using the pCYTEXP expression system. In vitro dimerization experiments revealed that both homo- and hetero-dimers can be formed from these monomeric proteins. The dimers were tested for their ability to promote capillary growth in vivo and stimulate DNA synthesis in cultured human vascular endothelial cells. Heterodimers comprising different VEGF splice forms, or combinations of VEGF/PIGF splice forms, showed mitogenic activity. The results demonstrate that four different heterodimeric growth factors are likely to have as yet uncharacterized functions in vivo.

Alternative Splicing↗

PubMed1996

The promoter-proximal, unstable IB region of the atp mRNA of Escherichia coli: an independently degraded region that can act as a destabilizing element.

Differential expression of the genes in the Escherichia coli atp (unc) operon is achieved via control of the translational initiation, translational coupling and mRNA stability of the respective genes. The atpIB region of the polycistronic mRNA is less stable than the remaining seven genes. We have investigated the functional half-lives of the atp genes in reconstructed versions of the operon. In order to be able to do this reliably, we have readdressed the interpretation of the complex functional inactivation data obtained by means of transcriptional inhibition using rifampicin. Our results indicate the usable information to be gleaned from this commonly applied technique, while identifying the potential errors in their quantitative interpretation. We estimate that the functional half-life of atpB is slightly over one-half that of atpE and the other atp genes, while atpI is at least two times less stable than atpB. The instability of the atpI mRNA was also demonstrated by its rapid fragmentation. Relocation of atpIB to a position in the promoter-distal region of the operon between atpG and atpD did not change the inactivation rate of atpB. However, it did destabilize the atpG mRNA. Examination of the physical degradation of atpI mRNA shows particularly rapid cleavage in this gene, thus explaining the destabilization effect. The atpIB segment is therefore an autonomously unstable region that can act as a destabilizing element for upstream-located genes in a polycistronic environment.

Escherichia coli↗

PubMed1996

VEGF121 induces proliferation of vascular endothelial cells and expression of flk-1 without affecting lymphatic vessels of chorioallantoic membrane.

We have studied the effect of VEGF(121) homodimer and VEGF(121/165) heterodimer on the chorioallantoic membrane (CAM) of 13-day-old chick embryos. The factors were applied in doses of 2-4 micrograms and the effects were evaluated macroscopically after 2 and 3 days. Histological studies were performed on semi- and ultrathin sections. Proliferation was studied according to the BrdU-anti-BrdU method on whole mounts and sections. The labeling density was quantified in whole mounts. The fractal dimension, D, of the vascular tree was assessed as a value for vascular bifurcation density. Both forms of VEGF induce brush-like vessel formation in the precapillary region. New capillaries are found in the stroma of the CAM, which normally does not contain capillaries. Our results show that VEGF(121) is a specific endothelial cell mitogen. A fourfold increase of BrdU-labeled endothelial cells is found after VEGF(121) application. The fractal dimension of the vascular tree increases from 1.26 in the controls to 1.44 (VEGF(121)) and 1.41 (VEGF(121/165)). The endothelial cells of the newly formed capillaries possess many mitochondria and micropinocytotic vesicles, but no fenestrations. These capillaries are obviously formed by intussusceptive microvascular growth. Signs of sprouting are almost absent. An effect on the lymphatic vessels of the CAM is not detectable. Compared to VEGF(165) and VEGF(121/165), VEGF(121) diffuses over a slightly greater distance. Using in situ hybridization, VEGF receptor-2 (flk-1/Quek1) and the homologous flt-4 (Quek2) receptor were studied in the CAM of normal quail embryos and after VEGF(121) application on the CAM of 11-day-old quail embryos. During normal development, flk-1 expression becomes restricted to vascular endothelial cells of large vessels in the stroma of the CAM. VEGF(121) application induces expression of flk-1 in capillaries that normally do not express the receptor. In the normal development of the CAM, flt-4 becomes restricted to endothelial cells of vessels that appear to be lymphatic vessels. Application of VEGF(121) does not alter flt-4 expression.

Allantois↗

PubMed1995

The F0 complex of the Escherichia coli ATP synthase. Investigation by electron spectroscopic imaging and immunoelectron microscopy.

Cholate-solubilized F0 complexes of the ATP synthase (F0F1) from Escherichia coli were studied by application of conventional transmission electron microscopy and electron spectroscopic imaging (ESI) of negatively stained samples. Using the ESI mode, the structural organization of the F0 complex (diameter of 7.5 +/- 0.5 nm) could be observed in more detail and defined projections could be distinguished. Projection A appears as a deltoid-like structure with bilateral symmetry. Projection B has an overall trapezoidal shape with some similarity in shape to the letter W. Applying the ESI mode to the ac complex dissolved in cholate-containing buffer, an elongated structure consisting of two intensity maxima could be observed. Simulations with models of the F0 and the ac complex revealed that the projections observed can be obtained by tilting and rotating a model in which subunit a and the two copies of subunit b are located outside the subunit c oligomer. This view of structural organization was supported by results obtained with F0 complexes decorated with monoclonal antibodies against subunits a, b or c.

Antibodies, Monoclonal↗