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Biomedical subjects

M Mittag

Publications and source records attributed to M Mittag.

At least 19 recordsLinked to original sources
PubMed2002

[ANCA-positive vasculitis of the skin and kidneys associated with acne conglobata].

A 19 year old man with severe acne conglobata and ulcerated pyoderma gangraenosum-like skin lesions on the face was first treated with isotretinoin (Roaccutan((R))), then immunosuppressively with prednisolone, diaminodiphenylsulfone (Dapson-Fatol((R))) and mycophenolate mofetil (Cellcept((R))). Under a daily maintenance dose of immunosuppressive treatment with 2.5 mg prednisolone and 1 g mycophenolate mofetil, weakness, muscle and joint aches appeared. Due to proteinuria, haematuria and an elevated antineutrophil cytoplasmic antibody (cANCA) and the histological detection of a leukocytoclastic vasculitis, the diagnosis of cANCA positive vasculitis of the skin and kidneys was established. Therapy with cyclophosphamide (Endoxan((R))) along with prednisolone was effective. An exact classification of this disease process was not possible.

Acne Vulgaris↗
PubMed2001

Identification of target mRNAs for the clock-controlled RNA-binding protein Chlamy 1 from Chlamydomonas reinhardtii.

An endogenous clock regulates the temporal expression of genes/mRNAs that are involved in the circadian output pathway. In the green alga Chlamydomonas reinhardtii, a clock-controlled RNA-binding protein (Chlamy 1) was identified recently, which represents an analog of the circadian trans-acting factor CCTR from the phylogenetically diverse alga Gonyaulax polyedra. In order to identify in C. reinhardtii target mRNAs that can be recognized by Chlamy 1, gel mobility-shift assays and UV-crosslinking experiments were carried out, and revealed that this protein interacts specifically with the 3' untranslated regions of several mRNAs and recognizes them all via a common cis-acting element, composed of at least seven UG repeats. By using competition assays, it was found that the affinity of Chlamy 1 is highest for mRNAs whose products are key components of nitrogen and CO2 metabolism. Since the activities of enzymes involved in nitrogen metabolism vary in a temporal pattern that is opposite in phase to that of Chlamy 1 binding activity, the protein may repress the translation of the cognate mRNAs.

3' Untranslated Regions↗
PubMed2001

Circadian rhythms in microalgae.

Circadian rhythms have been described in a variety of microalgae. In each group, some model organisms arose and most detailed studies have been done with them. They include the cyanobacterium ("blue-green alga") Synechococcus and eukaryotic microalgae Gonyaulax polyedra (Dinophyta), Chlamydomonas reinhardtii (Chlorophyta), and Euglena gracilis (Euglenophyta). This review focuses on recent approaches to depict molecular components of the circadian system and the mechanisms of regulation in these organisms. In Synechococcus, the identification of the kailocus, which represents a central part of its oscillatory system, is discussed, as well as diverse approaches based on a luminescent reporter gene, which is driven by a clock-controlled cyanobacterial promoter. In eukaryotic microalgae, the diversity of genes/proteins that are controlled by the circadian clock is described and the kind of regulation (transcriptional and translational control) is emphasized. The role and function of conserved clock-controlled RNA-binding proteins such as CCTR from Gonyaulaxor Chlamy 1 from Chlamydomonas are discussed.

Animals↗
PubMed2000

Fibroblasts surrounding melanoma express elevated levels of matrix metalloproteinase-1 (MMP-1) and intercellular adhesion molecule-1 (ICAM-1) in vitro.

Tumour growth and metastasis involve the degradation of extracellular matrix components by matrix degrading enzymes produced by tumour cells and stromal fibroblasts. In this study, fibroblasts were obtained from biopsies on the border (TB) and 1 cm distant from the melanoma (TD) and cultured separately. Similar studies were performed with fibroblasts surrounding melanocytic nevi as control. The expression of matrix metalloproteinase-1 (MMP-1) mRNA and tissue matrix metalloproteinase inhibitor 1 (TIMP-1) were studied by Northern blot analysis. The activation antigen intercellular adhesion molecule-1 (ICAM-1) in TB-and TD-fibroblasts was investigated by flow cytometry. In melanoma, TB-fibroblasts showed an increased expression of MMP-1 mRNA mainly in fibroblasts obtained from tumours with extended invasive growth demonstrated by Clark level whereas the expression of the major specific inhibitor TIMP-1 was unaltered. In contrast, fibroblasts surrounding benign melanocytic nevi did not express elevated levels of MMP-1. The upregulation of MMP-1 in TB-fibroblasts compared to TD-fibroblasts was maintained during cultivation. Furthermore, MMP-1 mRNA expression and MMP-1 total protein amount in normal fibroblasts were increased by melanoma cell conditioned medium. We demonstrated an increased expression of ICAM-1 in TB-fibroblasts compared to TD-fibroblasts in vitro depending on the amount of inflammatory infiltrate in situ. The differences of ICAM expression disappeared during continued cell culture. These results support the idea that fibroblasts surrounding melanoma are activated and are possibly involved in the degradation of matrix proteins surrounding the tumour.

Culture Media, Conditioned↗
PubMed2000

A novel function of yeast fatty acid synthase. Subunit alpha is capable of self-pantetheinylation.

The prosthetic group of yeast fatty acid synthase (FAS), 4'-phosphopantetheine, is covalently linked to Ser180 of subunit alpha. It originates from coenzyme A and is transferred to the enzyme by a specific phosphopantetheine:protein transferase (PPTase). The present study demonstrates that the FAS-activating PPTase of yeast represents a distinct catalytic domain of the FAS complex and resides within the C-terminal portion of subunit alpha. The autoactivation capacity of yeast FAS became evident from in vitro pantetheinylation studies using purified apo-FAS preparations. These were readily converted to pantetheinylated holo-FAS simply upon addition of free coenzyme A. Pantetheinylation-competent apo-FAS was prepared in vitro by constructing hybrid oligomers containing alpha-subunits from two different pantetheine-less FAS-mutants. The respective mutants were selected according to their ability to complement each other, in vivo. In vitro formation of hybrid apo-FAS complexes was achieved by dimethylmaleic anhydride (DMMA) -induced reversible dissociation of mixtures of the two constituent mutant enzymes. This treatment was both necessary and sufficient to produce pantetheinylation-competent apo-FAS. Specific FAS activities were comparable independent of whether the apo-enzymes were pantetheinylated in vivo or in vitro. Apart from the induction of overall FAS activity, incorporation of phosphopantetheine into apo-FAS was also demonstrated by the use of 3H-labelled coenzyme A, leading to the formation of radioactively labelled FAS. It is concluded that pantetheinylation of yeast FAS is performed by an intrinsic catalytic activity of the apo-enzyme proper. The endogenous PPTase acts in trans between different subunits alpha in the alpha6beta6 oligomer. The self-pantetheinylation of yeast FAS represents the first example of an apo-enzyme being capable of post-translational autoactivitation.

Alleles↗
PubMed1998

[Grönblad-Strandberg syndrome from the angiological viewpoint].

HISTORY AND CLINICAL FINDINGS: A 42-year-old man was admitted for treatment of peripheral vascular disease in the left leg (stage III of Fontaine). A year before he had undergone a right aortofemoral bypass operation. On admission there was stenosis of the left pelvic axis and bilateral femoral artery occlusion. In addition there were changes in the skin with abnormal folds, loss of elasticity and yellowish spots over the sides of the neck and the flexor surfaces of all large joints. In addition vision in the left eye was impaired. These findings suggested connective tissue disease involving the skin, eye and arterial system. INVESTIGATIONS: Routine haematological tests were normal as were clotting parameters. Serum concentration of GOT, GPT, gamma-GT were slightly increased. There was a dysproteinaemia with raised HDL and LDL levels. Resting electrocardiogram was normal, showing sinus rhythm and left axis deviation. The crurobrachial pressure index was clearly abnormal: 0.6 on the right and 0.5 on the left. Angiography of the pelvic and left arteries revealed long-segment femoral and partial lower-leg occlusions bilaterally. Abdominal sonography indicated diffuse parenchymal calcifications in both kidneys and angioid streaks on bilateral fundoscopy. Skin biopsy showed defects of elastic fibres and perivascular inflammatory infiltration, while capillary microscopy revealed twisting of the capillaries, most of them with normal lumen. These findings taken together indicated pseudoxanthoma elasticum (PXE) or Grönblad-Strandberg syndrome. TREATMENT AND COURSE: A thrombendarterectomy was performed on the left superficial femoral artery, after which the left popliteal artery became palpable, the pressure indices for the left leg were slightly better, and the patient was discharged home without further complications and improved leg perfusion. CONCLUSION: Possible cardiovascular involvement had to be taken into account in patients with PXE, and long-term angiological monitoring is indicated.

Adult↗
PubMed1998

[Progressive systemic scleroderma--prognosis determining involvement of internal organ systems].

Prognosis of systemic sclerosis (scleroderma, Ssc) is largely depending on involvement of internal organs. Abnormalities of the gastrointestinal tract are found most frequently (85%), especially decreased motility of the oesophagus, which has little impact on the longterm clinical course of Ssc. Pulmonary manifestations can be demonstrated in 40-90% of patients; one must distinguish between pulmonary hypertension or fibrotic lung disease. The heart is affected in 50% of cases. Patchy or diffuse myocardial fibrosis, as well as pericarditis and pericardial effusions can induce symptoms of arrhythmia or congestive heart failure. Renal involvement is associated with increased mortality and occurs in 45% of Ssc, producing proteinuria, hypertension, scleroderma renal crisis and renal failure. In conclusion, involvement of the lungs, heart and kidneys are determining factors for the longterm course of systemic sclerosis.

Heart Diseases↗
PubMed1998

Serum levels of soluble Fas/APO-1 receptor are increased in systemic sclerosis.

It has been suggested that rheumatic diseases may result from a deficit in Fas-mediated T-cell apoptosis. Recent studies have demonstrated increased soluble Fas in sera from lupus erythematosus patients. We were interested to determine whether elevated soluble Fas levels are associated with systemic sclerosis. Soluble Fas levels were retrospectively assayed using a sandwich enzyme-linked immunosorbent assay in serum from 30 patients with systemic sclerosis and 15 normal controls. Hospital medical records were retrospectively reviewed for clinical and laboratory characteristics of the patients. Soluble Fas levels were analysed in subsets of patients with limited (lcSSc) versus diffuse cutaneous systemic sclerosis (dcSSc) and correlated with inflammatory activity. In systemic sclerosis soluble Fas serum levels (lcSSc, 2.19 +/- 0.71 ng/ml, dcSSc 2.53 +/- 1.37 ng/ml) were significantly higher than in normal controls (1.26 +/- 0.36 ng/ml). However, there were no significant differences in soluble Fas levels between lcSSc and dcSSc and poor correlation between soluble Fas levels and inflammatory activity status. Detection of elevated soluble Fas might serve as a clinical marker for immunological dysregulation in systemic sclerosis, but not for inflammatory disease activity.

Antibodies, Antinuclear↗
PubMed1998

The mRNA level of the circadian regulated Gonyaulax luciferase remains constant over the cycle.

The expression of luciferin-binding protein (LBP) and luciferase (LCF), two proteins that are involved in bioluminescence in Gonyaulax polyedra, is controlled by a cellular circadian clock. In the case of LBP, its temporal expression is reported to be regulated at the translational level, involving both 5' and 3' untranslated regions (UTRs) of lbp mRNA. Here, we show that the amounts of lcf mRNA are constant throughout the day-night cycle, indicating that the circadian expression of LCF is also regulated at the translational level.

Animals↗
PubMed1997

Differential translational initiation of lbp mRNA is caused by a 5' upstream open reading frame.

Expression of the luciferin-binding protein (LBP) from Gonyaulax polyedra is regulated by the circadian clock at the translational level. Here we report that in vitro translation of lbp mRNA results in the synthesis of two LBP variants of different sizes, which is shown to be due to translational initiation at different in-frame AUG codons on lbp mRNA. Differential initiation is caused by a small open reading frame (ORF, situated in the 5' untranslated region of lbp mRNA), which gives rise to a leaky scanning mechanism. In Gonyaulax, only one of these variants, which is produced by initiation from the first AUG of the lbp ORF, exhibits a circadian rhythm and is far more abundant during night phase.

Amino Acid Sequence↗
PubMed1997

In vitro mutagenesis of binding site elements for the clock-controlled proteins CCTR and Chlamy 1.

The luciferin-binding protein (LBP) from the dinoflagellate, Gonyaulax polyedra, is regulated by a circadian clock at the translational level. A 22-nucleotide long interval in the lbp 3' untranslated region, which contains seven UG-repeats, was characterized as a circadian cis-acting element, to which a clock controlled factor (CCTR) binds. Recently we have found that the phylogenetically distant green alga, Chlamydomonas reinhardtii, contains a CCTR analog, called Chlamy 1. Here we show that the flanking nucleotides surrounding the UG-repeats are required for high binding activity of CCTR and Chlamy 1. The absence of three or more UG-repeats abolishes binding with both proteins.

Animals↗
PubMed1996

Conserved circadian elements in phylogenetically diverse algae.

Circadian expression of the luciferin-binding protein (LBP) from the dinoflagellate Gonyaulax polyedra is regulated at the translational level. A small interval in the lbp 3'-untranslated region, which contains seven UG-repeats, serves as a cis-acting element to which a trans-acting factor (CCTR) binds in a circadian manner. Its binding activity correlates negatively with the circadian expression of LBP. Here I report the identification of a protein in the green alga Chlamydomonas reinhardtii that represents a CCTR analog. It binds both specifically and under control of the circadian clock to the UG-repeat region. The data show for the first time that circadian cis-elements implicated in translational regulation have been conserved during evolution.

Animals↗
PubMed1995

Substrate and product binding sites of yeast fatty acid synthase. Stoichiometry and binding kinetics of wild-type and in vitro mutated enzymes.

The four known substrate binding sites of yeast fatty acid synthase (FAS), Ser819 (acetyltransferase, OHAC) and Ser5421 (malonyl/palmitoyl transferase, OHMa1) of subunit beta and Ser180 (pantetheine binding site, SHc) and Cys1305 (3-oxoacyl synthase, SHp) of subunit alpha were replaced, by targeted in vitro mutagenesis, by the non-acylatable amino acids glutamine, glycine or alanine. The four mutated FAS proteins together with two pairs of double mutants (OHAc/OHMa1 and SHc/SHp) were episomally expressed in appropriate delta fas1 or delta fas2 deletion strains. The purified enzymes isolated from these transformants were used for comparative acyl binding studies with the substrates [1-14C]acetyl-CoA and [2-14C]malonyl-CoA. Malonate was found to be transacylated to enzyme-bound pantetheine (SHc) exclusively by the Ser5421 hydroxyl group of malonyltransferase (OHMa1) while acetate could use both the acetyl (Ser819) and the malonyl (Ser5421) transferase active sites on its way to the SHc and SHp binding sites. Acylation of SHc with either substrate was unaffected by the absence of the 'peripheral' SH group (SHp) while binding of acetate to SHp was dependent on enzyme-bound pantetheine (SHc). These genetic data support a revised model regarding the intra-molecular channeling of acetate and malonate within yeast fatty acid synthase. Quantitative acyl binding studies revealed a maximum of 2-3 mol rather than the expected 12 mol of malonate and of 6-7 mol rather than 24 mol of acetate bound/mol hexameric yeast FAS. Only 20-30% of the malonyl-enzyme and 35-50% of the acetyl enzyme represented performic-acid-labile thioester bonds. The binding characteristics of both substrates, exhibiting Hill coefficients distinctly lower than 1, as well as their non-linear Lineweaver-Burk and Scatchard plots, point to a marked negative cooperativity among the 12 yeast FAS subunits. The observed sub-stoichiometric substrate binding characteristics of the enzyme are ascribed to this effect. An a priori asymmetry of the complex appears unlikely since the coenzyme-A:FAS transacylation equilibrium may be shifted towards the fully acetylated enzyme in the presence of N-ethylmaleimide. In contrast to the limited acylation capacity of the 'resting' enzyme, complete acylation of yeast FAS at all of its 12 SHc and SHp sites is observed under steady-state conditions of fatty acid biosynthesis. Under these conditions, the enzyme exhibits full-site reactivity at its SHp, SHc and OHAc sites, but a concomitant 18-fold increase in Km of the coenzyme-A:OHAc transacylation reaction keeps the acyl-O-ester content of the acylated enzyme at less than 5% of the total.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetates↗