ABSTRACT
BACKGROUND: During the development of the central nervous system (CNS), patterning processes along the dorsoventral (DV) axis of the neural tube generate different neuronal subtypes. As development progresses these neurons are arranged into functional units with varying cytoarchitecture, such as laminae or nuclei for efficient relaying of information. Early in development ventral and dorsal regions are similar in size and structure. Different proliferation rates and cell migration patterns are likely to result in the formation of laminae or nuclei, eventually. However, the underlying molecular mechanisms that establish these different structural arrangements are not well understood.We undertook a differential display polymerase chain reaction (DD-PCR) screen to identify genes with distinct expression patterns between dorsal and ventral regions of the chick midbrain in order to identify genes which regulate the sculpturing of such divergent neuronal organisation. We focused on the DV axis of the early chick midbrain since mesencephalic alar plate and basal plate develop into laminae and nuclei, respectively. RESULTS: We identified 53 differentially expressed bands in our initial screen. Twenty-six of these could be assigned to specific genes and we could unambiguously show the differential expression of five of the isolated cDNAs in vivo by in situ mRNA expression analysis. Additionally, we verified differential levels of expression of a selected number of genes by using reverse transcriptase (RT) PCR method with gene-specific primers.One of these genes, QR1, has been previously cloned and we present here a detailed study of its early developmental time course and pattern of expression providing some insights into its possible function. Our phylogenetic analysis of QR1 shows that it is the chick orthologue of Sparc-like 1/Hevin/Mast9 gene in mice, rats, dogs and humans, a protein involved in cell adhesion. CONCLUSION: This study reveals some possible networks, which might be involved in directing the difference in neuronal specification and cytoarchitecture observed in the brain.
Subject(s)
Avian Proteins/genetics , Gene Expression Regulation, Developmental , Mesencephalon/embryology , Animals , Avian Proteins/metabolism , Body Patterning , Chick Embryo , Chickens , In Situ Hybridization , Mesencephalon/cytology , Mesencephalon/metabolism , Phylogeny , Polymerase Chain Reaction , RNA, Messenger/metabolismABSTRACT
A subclass of zinc finger proteins containing a unique protein motif called the positive regulatory (PR) domain has been described. The members include the PRDI-BF1/Blimp-1 protein, the Caenorhabditis elegans egl-43 and EVI1 gene products, and the retinoblastoma interacting protein RIZ. Here we describe a member of this family, SC-1, that exhibits several distinctive features. First, SC-1 interacts with the p75 neurotrophin receptor and is redistributed from the cytoplasm to the nucleus after nerve growth factor (NGF) treatment of transfected COS cells. The translocation of SC-1 to the nucleus was specific for p75, as NGF binding to the TrkA receptor did not lead to nuclear localization of SC-1. Thus, SC-1 provides a downstream transducer for the effects of NGF through the p75 neurotrophin receptor. Under normal growth conditions, SC-1 was found predominantly in the cytoplasm. On serum-starvation, SC-1 also translocated into the nucleus. A direct correlation between nuclear expression of SC-1 with the loss of BrdUrd incorporation was observed. These results imply that SC-1 may be involved in events associated with growth arrest.
Subject(s)
Carrier Proteins/chemistry , Carrier Proteins/metabolism , Intracellular Signaling Peptides and Proteins , Nerve Growth Factors/metabolism , Zinc Fingers , Amino Acid Sequence , Animals , Brain-Derived Neurotrophic Factor/metabolism , Bromodeoxyuridine/metabolism , COS Cells , Cell Line , Cell Nucleus/metabolism , Culture Media, Serum-Free/metabolism , Cytoplasm/metabolism , DNA, Complementary/metabolism , DNA-Binding Proteins , Humans , Models, Genetic , Molecular Sequence Data , Neurotrophin 3 , Nuclear Proteins , Proto-Oncogene Proteins/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , Receptor, Nerve Growth Factor , Receptor, trkA , Receptors, Nerve Growth Factor/metabolism , Sequence Homology, Amino Acid , Time Factors , Transcription FactorsABSTRACT
The mechanism of action of NGF has continued to provide a challenging and formidable problem in signal transduction. NGF can bind independently to two different receptors, the trkA tyrosine kinase receptor and the p75 neurotrophin receptor, which are involved in many different signaling events. In addition to promoting cell differentiation survival, NGF can paradoxically be an inducer of cell death. Several receptor mediated mechanisms are proposed to explain how NGF might act as a trophic factor and as a cell killer. The survival and cell death properties of the receptors are dependent upon the relative ratio of receptors and the persistent nature of the signaling events.
Subject(s)
Brain/physiology , Nerve Growth Factors/physiology , Proto-Oncogene Proteins/physiology , Receptor Protein-Tyrosine Kinases/physiology , Receptors, Nerve Growth Factor/physiology , Animals , Brain/cytology , Cell Death , Cell Survival , Humans , Neurons/cytology , Neurons/physiology , Receptor, Nerve Growth Factor , Receptor, trkAABSTRACT
VILIP is a member of the visinin/recoverin family of neuronal EF-hand Ca(2+)-binding proteins. Cell fractionation revealed cytoplasmic, membrane- and cytoskeleton-associated pools of VILIP. The association with the cytoskeletal protein fraction is Ca(2+)-dependent and may be mediated by direct interaction with actin. This is concluded from the observations that (i) Ca(2+)-loaded recombinant VILIP binds actin in an overlay assay; (ii) in the presence of Ca(2+), beta-actin co-immunoprecipitates with native VILIP from brain extracts, and (iii) actin and VILIP are co-localized in PC12 cells stably transfected with VILIP cDNA. The interaction of VILIP with the cortical cytoskeleton through actin may facilitate a Ca(2+)-dependent recruitment of VILIP to the cell membrane.
Subject(s)
Actins/metabolism , Calcium-Binding Proteins/metabolism , Cell Membrane/metabolism , Cytoskeleton/metabolism , Nerve Tissue Proteins/metabolism , Receptors, Calcium-Sensing , Animals , Animals, Newborn , Brain/metabolism , Calcium-Binding Proteins/genetics , Chickens , In Vitro Techniques , Nerve Tissue Proteins/genetics , Neurocalcin , PC12 Cells , Protein Binding , Rats , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Subcellular Fractions/metabolism , TransfectionABSTRACT
Growth of the normal and malignant prostate is known to be regulated by androgens. Part of their effect has been suggested to be mediated through coordinated regulation of secreted growth factors with autocrine function. We now examine the biological role of preferentially paracrine acting factors in growth control of prostate cancer, i.e. fibroblast growth factor(s) (FGF). Coculture experiments using the androgen-responsive human prostate carcinoma cell line LNCaP as feeder cells and the FGF-dependent human adrenal carcinoma SW-13 cell line as target cells show that (i) LNCaP cells induce growth of SW-13 cells, (ii) even higher stimulation of SW-13 cells is seen in the presence of androgen treated LNCaP cells and (iii) a specific anti-bFGF antibody inhibits growth of SW-13 cells induced by androgen treated LNCaP cells; no proliferation of SW-13 cells occurs in the absence of LNCaP cells. Partial purification of the secretory products of LNCaP cells was performed by affinity chromatography using a heparin sepharose column. Fractions were tested for biological activity in a soft agar assay with SW-13 cells. Several activities could be detected, the main activity was eluted with about 1.5 M NaCl. These data suggest that androgen treatment of LNCaP cells leads to enhanced secretion of proteins which belong to the FGF-family.