ABSTRACT
Bmf is a proapoptotic member of the BH3-only subgroup of Bcl-2 family proteins, which is associated to myosin V motors by binding to the dynein light chain 2 (DLC2). It acts as a sentinel detecting intracellular damages on the main cytoskeletal structures. The cloning and characterization of the chicken (Gallus gallus) Bmf cDNA and splicing variant is described in this report. The Bmf cDNA was amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) using oligonucleotide primers derived from in silico sequences. The chicken Bmf cDNA encodes a protein of 193 amino acids, showing homology to mammalian Bmf proteins. A splicing variant of the chicken Bmf (Bmf(S), short isoform of Bmf) coding a protein of 118 amino acids was also identified. This is the first Bmf isoform identified so far which lacks the DLC2-binding domain although retaining the BH3 domain. Both chicken Bmf isoforms induced apoptosis 24 h after transfection in MCF7 and HeLa cell lines, but chicken Bmf(S) exhibits a higher proapoptotic activity. In addition, mRNA expression analysis showed that chicken Bmf transcription is ubiquitous in all embryo developmental stages, suggesting a role for this protein in the control of the development process.
Subject(s)
Adaptor Proteins, Signal Transducing/genetics , Alternative Splicing , Chickens/genetics , Adaptor Proteins, Signal Transducing/analysis , Adaptor Proteins, Signal Transducing/classification , Amino Acid Sequence , Animals , Apoptosis , Base Sequence , Chickens/growth & development , Cloning, Molecular , DNA, Complementary/genetics , HeLa Cells , Humans , Molecular Sequence Data , Phylogeny , Protein Isoforms/genetics , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , TransfectionABSTRACT
Bid protein, a member of the "BH3-only" subgroup of Bcl-2 family, plays a critical role in mammalian apoptosis regulation. In this study, we have cloned the chicken Bid gene, which encodes a 193 amino acid protein and shares 40% homology with human and mouse Bid proteins. Bid sequence comparison emphasises the conservation of both the functional domain BH3 and the proteolytic cleavage sites. An induction of apoptosis by chicken Bid and the cleavage of the protein, after TNFalpha treatment, were also demonstrated. In addition, mRNA Bid expression was detected along all embryo stages and tissues examined, suggesting a role for this protein in the developmental process. This is the first report demonstrating the functionality of a "BH3-only" protein in chicken.
Subject(s)
BH3 Interacting Domain Death Agonist Protein/metabolism , Amino Acid Sequence , Animals , Apoptosis/drug effects , BH3 Interacting Domain Death Agonist Protein/chemistry , BH3 Interacting Domain Death Agonist Protein/genetics , Chick Embryo , Chickens , Gene Expression Profiling , Gene Expression Regulation, Developmental , Humans , Molecular Sequence Data , Phylogeny , Protein Processing, Post-Translational , RNA, Messenger/genetics , RNA, Messenger/metabolism , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
The chicken (Gallus gallus) is one of the primary models for embryological and developmental studies. In order to begin to understand the molecular mechanisms underlying the normal and abnormal development of the chicken, we used 2-DE to construct a whole-embryo proteome map. Proteins were separated by IEF on IPG strips, and by 11% SDS-PAGE) gels. Protein identification was performed by means of PMF with MALDI-TOF-MS. In all, 105 protein spots were identified, 35 of them implicated in embryo development, 10 related with some diseases, and 16, finally, being proteins that have never been identified, purified or characterized in the chicken before. This map will be updated continuously and will serve as a reference database for investigators, studying changes at the protein level under different physiological conditions.
Subject(s)
Chick Embryo/growth & development , Proteome/analysis , Animals , Chick Embryo/metabolism , Electrophoresis, Gel, Two-Dimensional/methods , Gene Expression Regulation, Developmental/physiologyABSTRACT
In 1993 a destructive new Phytophthora pathogen of riparian Alnus trees was discovered in the UK and subsequently shown to be present in other parts of Europe. The new Phytophthora comprised a group of emergent heteroploid hybrids, probably between P. cambivora and a species related to P. fragariae. These included a common, near tetraploid standard hybrid, the presumptive allopolyploid; and four scarcer major variant types with chromosome numbers intermediate between diploid and tetraploid, named the Swedish, Dutch, German and UK variants. The standard hybrid type is formally designated here as Phytophthora alni subsp. alni. The Swedish variant is designated as P. alni subsp. uniformis; and the Dutch, German and UK variants collectively as P. alni subsp. multiformis. The properties of the Dutch, German and UK variants within subsp. multiformis are informally described. The problems of designating emergent species hybrids under the International Code of Botanical Nomenclature and the reasons for the taxonomic choices made are discussed.
Subject(s)
Alnus , Phytophthora/classification , Plant Diseases/microbiology , Base Sequence , DNA, Fungal/chemistry , DNA, Fungal/genetics , DNA, Intergenic/chemistry , DNA, Intergenic/genetics , Europe , Molecular Sequence Data , Phylogeny , Phytophthora/genetics , Phytophthora/growth & development , Phytophthora/ultrastructure , Sequence Analysis, DNAABSTRACT
We report a 16-week-gestation foetus obtained by voluntary abortion after prenatal diagnosis, in which a ring chromosome 22 was observed with deletion of the 22q13.3 region. A prenatal study of the amniotic fluid by standard chromosome technique with G bands and FISH (fluorescence in situ hybridisation) was performed. After the abortion, the anatomopathological study of the obtained foetus was carried out. Morphological and histological analysis of the foetus did not reveal severe physical abnormalities, although alterations of the nervous system were observed consisting of corpus callosum, fornix and septum pellucidum agenesia. It could be that the genes in this region that were involved in the development of the central nervous system were responsible for the alterations found in the morphological study. The wide range of manifestations observed in patients with this cytogenetic alteration is probably due to size differences in the deleted region.
