ABSTRACT
BACKGROUND: Owing to their complex nature, social interaction tests normally require the observation of video data by a human researcher, and thus are difficult to use in large-scale studies. We previously established a statistical method, a hidden Markov model (HMM), which enables the differentiation of two social states ("interaction" and "indifference"), and three social states ("sniffing", "following", and "indifference"), automatically in silico. NEW METHOD: Here, we developed freeware called DuoMouse for the rapid evaluation of social interaction behavior. This software incorporates five steps: (1) settings, (2) video recording, (3) tracking from the video data, (4) HMM analysis, and (5) visualization of the results. RESULTS: Using DuoMouse, we mapped a genetic locus related to social interaction. We previously reported that a consomic strain, B6-Chr6C(MSM), with its chromosome 6 substituted for one from MSM/Ms, showed more social interaction than C57BL/6 (B6). We made four subconsomic strains, C3, C5, C6, and C7, each of which has a shorter segment of chromosome 6 derived from B6-Chr6C, and conducted social interaction tests on these strains. DuoMouse indicated that C6, but not C3, C5, and C7, showed higher interaction, sniffing, and following than B6, specifically in males. COMPARISON WITH EXISTING METHOD: The data obtained by human observation showed high concordance to those from DuoMouse. The results indicated that the MSM-derived chromosomal region present in C6-but not in C3, C5, and C7-associated with increased social behavior. CONCLUSIONS: This method to analyze social interaction will aid primary screening for difference in social behavior in mice.
Subject(s)
Chromosome Mapping , Genetic Variation , Interpersonal Relations , Markov Chains , Pattern Recognition, Automated , Animals , Behavioral Research/instrumentation , Behavioral Research/methods , Chromosomes, Human, Pair 6/genetics , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Quantitative Trait LociABSTRACT
The Phox(S) strain of Drosophila melanogaster is an electrophoretically slow variant found in a wild population at Victoria, Australia. Prophenol oxidase isoform A(1) from PHOX-S was purified and characterized biochemically and genetically. The purified fraction of A(1) from PHOX-S showed a homodimer with a molecular weight of the subunit of approximately 77 kDa. The Phox(S) strain was temperature sensitive in vivo in culture, and the purified protein was thermolabile in vitro. By the deletion mapping method, the Phox(S) locus was cytologically estimated to be at the location 55-A on the right arm of the second chromosome and 79.6 genetically. These data show that PHOX-S is an electrophoretic variant of MOX and that PHOX-S is the first thermolabile protein found in invertebrate prophenol oxidase.
Subject(s)
Catechol Oxidase/genetics , Catechol Oxidase/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Enzyme Precursors/genetics , Enzyme Precursors/metabolism , Animals , Catechol Oxidase/isolation & purification , Chromosome Mapping , Cold Temperature , Drosophila Proteins/isolation & purification , Drosophila melanogaster/enzymology , Drosophila melanogaster/genetics , Electrophoresis, Polyacrylamide Gel , Enzyme Precursors/isolation & purification , Female , Hot Temperature , Isoenzymes/genetics , Isoenzymes/isolation & purification , Isoenzymes/metabolism , Larva/enzymology , Larva/genetics , Male , Pupa/enzymology , Pupa/geneticsABSTRACT
Circular dichroism (CD) of purified Drosophila melanogaster prophenol oxidase has been measured in the range of 195-245 nm. So far, few investigations about the interaction on higher-order structures have been performed. CD spectra of Drosophila prophenol oxidase with 2-propanol activator showed fluctuation of alpha-helices. At a high temperature of 80 degrees C, prophenol oxidase was partially denatured. However, it showed reversible recovery by renaturation after returning to low temperature at 30 degrees C. The conformational changes and reversible denaturation-renaturation interaction of the prophenol oxidase protein are discussed.
Subject(s)
Catechol Oxidase/chemistry , Drosophila melanogaster/enzymology , Enzyme Precursors/chemistry , 2-Propanol/pharmacology , Animals , Catechol Oxidase/metabolism , Circular Dichroism , Enzyme Activation/drug effects , Enzyme Precursors/metabolism , Enzyme Stability , Protein Conformation , TemperatureSubject(s)
Drosophila melanogaster/enzymology , Drosophila melanogaster/growth & development , Isoenzymes/metabolism , Monophenol Monooxygenase/metabolism , Animals , Drosophila melanogaster/genetics , Electrophoresis, Polyacrylamide Gel , Insect Proteins/biosynthesis , Isoenzymes/genetics , Larva/enzymology , Larva/genetics , Larva/growth & development , Monophenol Monooxygenase/genetics , Mutation , Time FactorsABSTRACT
Phenol oxidase exists in Drosophila hemolymph as a prophenol oxidase, A1 and A3, that is activated in vivo with a native activating system, AMM-1, by limited proteolysis with time. The polypeptide in purified prophenol oxidase A3 has a molecular weight of approximately 77,000 Da. A PCR-based cDNA sequence coding A3 has 2501 bp encoding an open reading frame of 682 amino acid residues. The potential copper-binding sites, from Trp-196 to Tyr-245, and from Asn-366 to Phe-421, are highly homologous to the corresponding sites in other invertebrates. The availability of prophenol oxidase cDNA should be useful in revealing the biochemical differences between A1 and A3 isoforms in Drosophila melanogaster that are refractory or unable to activate prophenol oxidase.
