Subject(s)
Dog Diseases/diagnostic imaging , Lung Diseases, Interstitial/veterinary , Lung Neoplasms/veterinary , Mast-Cell Sarcoma/veterinary , Tomography, X-Ray Computed/veterinary , Animals , Diagnosis, Differential , Dogs , Female , Lung Diseases, Interstitial/diagnostic imaging , Lung Neoplasms/diagnostic imaging , Mast-Cell Sarcoma/diagnostic imaging , Pulmonary Alveoli/diagnostic imagingABSTRACT
Pulsed laser cross-linking provides a means of introducing a covalent bond between proteins and the nucleic acids to which they are bound. This rapid cross-linking effectively traps the equilibrium that exists at the moment of irradiation and thus allows examination of the protein-nucleic acid interactions that existed. Laser irradiation may also induce photodestruction of protein and we have used the bacteriophage T4 gene 32 protein to investigate this phenomenon. Our results show that both nonspecific and specific photoproducts can occur, specifically at wavelengths where the peptide backbone of proteins is known to absorb. These results demonstrate that nonspecific photodegradation can be correlated with the formation of a specific photodegradation product. The formation of this product was monitored to show that product yield is nonlinearly dependent on laser power and wavelength. We have also investigated an unexpected photoproduct whose formation is dependent on the length of the polynucleotide to which the gene 32 protein binds and that further demonstrates the complexities of analyzing protein-nucleic acid interactions through the use of UV laser cross-linking. These data provide essential information for the establishment of appropriate conditions for future studies that use UV cross-linking of protein-nucleic acid complexes.
Subject(s)
DNA, Viral/radiation effects , DNA-Binding Proteins/radiation effects , Viral Proteins/radiation effects , Cross-Linking Reagents , DNA, Viral/chemistry , DNA-Binding Proteins/chemistry , Lasers , Oligodeoxyribonucleotides/chemistry , Oligodeoxyribonucleotides/radiation effects , Photochemistry , Viral Proteins/chemistryABSTRACT
The function of the extramacrochaetae (emc) gene is required to establish the normal spatial pattern of adult sensory organs in Drosophila. emc acts to suppress sensory organ development in certain regions of the body surface, apparently by antagonizing the function of the achaete and scute genes of the achaetescute complex (AS.C). We have found that emc encodes a novel member of the helix-loop-helix (HLH) family of proteins. The emc protein shares the dimerization domain of other HLH proteins but lacks their DNA binding motif. We propose a model in which the emc protein negatively regulates sensory organ determination by forming heterodimers with the HLH proteins encoded by the AS-C and/or daughterless, thereby altering or interfering with their activity.
Subject(s)
DNA-Binding Proteins/genetics , Drosophila Proteins , Drosophila/genetics , Genes, Regulator , Repressor Proteins , Alleles , Amino Acid Sequence , Animals , Base Sequence , Basic Helix-Loop-Helix Transcription Factors , Cloning, Molecular , DNA/genetics , DNA/isolation & purification , DNA Transposable Elements , DNA-Binding Proteins/physiology , Drosophila/embryology , Drosophila/growth & development , Embryo, Nonmammalian/physiology , Gene Expression Regulation , Molecular Sequence Data , Mutation , Nucleic Acid Hybridization , Oligonucleotide Probes , RNA/genetics , RNA/isolation & purification , Restriction Mapping , Sequence Homology, Nucleic Acid , Transcription, GeneticABSTRACT
A radioreceptor assay using [3H]2-beta-endorphin (B-EP) was used to examine binding in the brains of rats that had been exercised on a treadmill for 5 months. The binding of [3H]2-beta-EP to brain homogenates gave an average Kd of 0.830 +/- 0.055 nM for the controls and 0.826 + 0.022 nM for the runners, and respective Bmax's of 166 +/- 20 pM and 159 +/- 27 pM.
