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Biomedical subjects

W Bottke

Publications and source records attributed to W Bottke.

11 recordsLinked to original sources
PubMed1996

Ferritin mRNAs in Schistosoma mansoni do not have iron-responsive elements for post-transcriptional regulation.

Schistosoma mansoni possesses two isoforms of ferritin, soma and yolk ferritin. The soma ferritin occurs at a low level in most cells of both genders, whereas the yolk ferritin is a female-specific gene product that is expressed at high level in the vitellarium. In higher animals, ferritin mRNA is regulated by iron via the interaction of cytoplasmic binding proteins (IRPs) with a specific sequence element in the 5' untranslated region (UTR) referred to as the iron-responsive element (IRE). Sequence studies of the 5' UTRs, gel retardation assays, and hybridization experiments show that neither ferritin mRNAs of S. mansoni is regulated by an IRE/IRP mechanism. It is suggested that ferritins in schistosomes are controlled only at the transcriptional level.

Animals↗
PubMed1995

An isoform of ferritin as a component of protein yolk platelets in Schistosoma mansoni.

Schistosoma mansoni possesses two isoforms of the iron storage protein ferritin, Fer1 and Fer2. At the mRNA level as well as at the protein level, Fer1 is much more abundant than Fer2; females contain an about 15-fold excess of Fer1 compared with males. In contrast, nearly equal amounts of Fer2 occur in both sexes. By electron microscopy we identified ferritin as a component of electron dense membrane-bound bodies in cells of the vitellarium. The mode of formation of these inclusions (as inferred from electron microscopy) and the abundance of phospholipid multilayered membranes suggest that these bodies are of a lysosomal nature. Here we interpret these ferritin-containing inclusions as protein yolk platelets. To date, most of the literature does not contain any hints of the existence of protein yolk in trematodes. The possible function of ferritin in embryonic development is discussed.

Animals↗
PubMed1994

cDNA cloning and deduced amino acid sequence of two ferritins: soma ferritin and yolk ferritin, from the snail Lymnaea stagnalis L.

Pulmonate freshwater snails contain two different ferritin types, soma ferritin and yolk ferritin. A cDNA library was constructed from midgut gland poly(A)-rich RNA of the snail Lymnaea stagnalis L. and recombinant clones encoding both ferritin types were obtained by immunoscreening. The longest cDNA inserts had a length of 859 bp (soma ferritin) and 1548 bp (yolk ferritin) and the specificity of these inserts was confirmed by immunoprecipitation of both ferritin types translated in vitro from hybrid-selected mRNAs. The 5' untranslated region (UTR) of the soma ferritin mRNA contains a 28-bp element which shows 64% sequence identity with the iron-responsive element (IRE) of vertebrate ferritin mRNAs. The soma ferritin mRNA is strongly translated in the wheat germ system but poorly translated in rabbit reticulocyte lysate. The yolk ferritin mRNA, which contains no IRE, is equally well translated in both in vitro translation systems. The deduced amino acid sequence of the soma ferritin subunit (174 amino acid residues, M(r) 20140) shows 50-70% sequence identity with subunits of vertebrate ferritins. After removal of an 18-amino-acid-residue signal sequence the deduced protein sequence of yolk ferritin contains 221 amino acids (M(r) 25438). Sequence identity of this chain with other eukaryotic ferritin chains is only 31-42%. Both snail ferritin sequences are more similar to the H-subunit type of vertebrate ferritins than to the L-type and both have the H-specific amino acid residues of the ferroxidase centre. The yolk ferritin sequence has a 42-amino-acid-residue insertion predicted to reside in the L loop of the subunit.

Amino Acid Sequence↗
PubMed1994

Expression in Escherichia coli of a secreted invertebrate ferritin.

The coding regions of the cDNAs for cytoplasmic soma ferritin and secreted yolk ferritin from the snail Lymnaea stagnalis were inserted into the prokaryotic expression vector pEMBLex2. The vector directed the synthesis in Escherichia coli of soma ferritin up to a concentration of 15% of soluble proteins. Soma ferritin was expressed as the multimeric protein (480 kDa). Its similarity with natural soma ferritin was confirmed by PAGE, immunostaining and electron microscopy. Yolk ferritin was expressed in the form of inclusion bodies. Attempts to refold and assemble the purified yolk ferritin subunit in vitro failed. The yolk ferritin coding sequence was therefore inserted into the expression vector pMAL-p2. At a growth temperature of the bacterial cells of 23 degrees C and at an isopropyl beta-D-thiogalactopyranoside concentration of 50 microM, about 5% of the induced MalE-yolk-ferritin fusion protein was secreted into the periplasmic space and could be purified by affinity chromatography on amylose; the rest occurred as insoluble cytoplasmic inclusion bodies. Soluble MalE-yolk-ferritin fusion protein was capable of assembly into ferritin-like particles. Fully assembled yolk apoferritin shells (610 kDa) were obtained by digestion of these particles with proteinase K (yield: 180 micrograms yolk ferritin/l bacterial culture). Recombinant yolk ferritin was capable of taking up iron in vitro. Yolk ferritin (610 kDa) and soma ferritin (480 kDa) were run to the pore limit of a non-denaturing 5-20% PAGE gradient gel. Under these conditions, yolk ferritin had a higher mobility than soma ferritin (480 kDa) and therefore the yolk ferritin may have a rather compact structure. A 41-amino-acid-residue stretch of the insertion, a distinctive feature of the yolk ferritin subunit, was deleted by site-directed mutagenesis. The MalE-yolk-ferritin variant thus obtained was readily degradable by proteinase K and could not be assembled into ferritin-like particles. Therefore residues in the deleted peptide must be important for the maintenance of the native structure.

ATP-Binding Cassette Transporters↗
PubMed1982

Isolation and properties of vitellogenic ferritin from snails.

The iron storage protein ferritin is the principal yolk protein in oocytes of the snails Planorbarius corneus L. and Lymnaea stagnalis L. This report gives an account of the isolation procedure and of some properties of snail ferritins. The isolation procedure includes a heat-denaturation step, gel filtration on Sepharose 6B and two ultracentrifugation steps followed by electrophoresis. Ferritins from both snails are highly reminiscent of vertebrate and plant ferritins in terms of heat stability, absorption spectrum and ultrastructure, and both share common antigen determinants with horse spleen ferritin. In different electrophoresis systems snail ferritins display considerable heterogeneity and microheterogeneity. Electrophoresis in the presence of sodium dodecyl sulphate (SDS) yields two major polypeptides with molecular weights of 19 000 and 24 000, which are interpreted to be authentic subunits of the ferritin molecule. Different organs and tissues of the snails differ in subunit composition. Midgut gland ferritin consists predominantly of the 19 000 Mr polypeptide, while in embryos only the 24 000 Mr band was found. No carbohydrates or lipids could be detected by staining acrylamide gels. Results from SDS/acrylamide electrophoresis, electrophoresis under non-denaturing conditions on gradient gels and from isoelectric focusing indicate that the ferritins of both snails are composed of at least two different types of ferritin that are tissue-specific. One ferritin is typical of somatic tissue (midgut gland) and is most probably a homopolymer of the 19 000 Mr subunit. The other ferritin is typical of oocytes, but since it is an exogenous protein it is also encountered in the midgut gland (the presumed site of yolk synthesis) and the haemolymph. Vitellogenic ferritin is either a homopolymer of the 24 000 Mr subunit or is predominantly composed of it. So far, there is no evidence for a precursor-product relationship between the two subunits.

Animals↗
PubMed1976

Chromosome-associated paracrystalline nuclear inclusions in the spermatocytes of a pulmonate snail, Planorbarius corneus L.

Chromosome-associated paracrystalloids are regularly found in the spermatocytes of snails which were reared in the laboratory. They seem to be largely specific for the male gametocytes as they have been observed only in few cases in the oocytes. It is likely that paracrystalloids are formed during pachytene at the site of large heterochromatic knobs which originate by fusion of heterochromatic terminal segments of some bivalents. During diplotene they are always connected with the telomeres of three or four bivalents, thus forming a large trefoil-like structure. During metaphase I paracrystalloids are shed off from the chromosomes and transferred to the cytoplasm. In early spermatids they are found again in the nuclei, where they "fade away" during spermiogenesis. Histochemically they consist of basic proteins, which are probably crystallized in the cubic system. Radioactive labeling of the structure could not be achieved, neither by 3H-uridine or thymidine, nor by amino acids. The functional significance of this peculiar structure in unknown. Certain features justify a comparison with synaptonemal polycomplexes.

Animals↗
PubMedDate not supplied

Structure, function, and evolution of ferritins.

The ferritins of animals and plants and the bacterioferritins (BFRs) have a common iron-storage function in spite of differences in cytological location and biosynthetic regulation. The plant ferritins and BFRs are more similar to the H chains of mammals than to mammalian L chains, with respect to primary structure and conservation of ferroxidase center residues. Hence they probably arose from a common H-type ancestor. The recent discovery in E. coli of a second type of iron-storage protein (FTN) resembling ferritin H chains raises the question of what the relative roles of these two proteins are in this organism. Mammalian L ferritins lack ferroxidase centers and form a distinct group. Comparison of the three-dimensional structures of mammalian and invertebrate ferritins, as well as computer modeling of plant ferritins and of BFR, indicate a well conserved molecular framework. The characterisation of numerous ferritin homopolymer variants has allowed the identification of some of the residues involved in iron uptake and an investigation of some of the functional differences between mammalian H and L chains.

Amino Acid Sequence↗