Immunohistochemical study of Ca2+/calmodulin-dependent protein kinase II in the Drosophila brain using a specific monoclonal antibody.
To analyze the distribution of Drosophila calcium/calmodulin-dependent protein kinase II (dCaMKII) in the adult brain, we generated monoclonal antibodies against the bacterially expressed 490-amino acid (a.a.) form of dCaMKII. One of those, named #18 antibody, was used for this study. Western blot analysis of the adult head extracts showed that the antibody specifically detects multiple bands between 55 and 60 kDa corresponding to the molecular weights of the splicing isoforms of dCaMKII. Epitope mapping revealed that it was in the region between 199 and 283 a.a. of dCaMKII. Preferential dCaMKII immunoreactivity in the embryonic nervous system, adult thoracic ganglion and gut, and larval neuro-muscular junction (NMJ) was consistent with previous observations by in situ hybridization and immunostaining with a polyclonal antibody at the NMJ, indicating that the antibody is applicable to immunohistochemistry. Although dCaMKII immunoreactive signal was low in the retina, it was found at regular intervals in the outer margin of the compound eye. These signals were most likely to be interommatidial bristle mechanosensory neurons. dCaMKII immunoreactivity in the brain was observed in almost all regions and relatively higher staining was found in the neuropilar region than in the cortex. Higher dCaMKII immunoreactivity in the mushroom body (MB) was found in the entire gamma lobe including the heel, and dorsal tips of the alpha and alpha' lobes, while cores of alpha and beta lobes were stained light. Finding abundant dCaMKII accumulation in the gamma lobe suggested that this lobe might especially require high levels of dCaMKII expression to function properly among MB lobes.
Department of Brain Structure, Tokyo Metropolitan Institute for Neuroscience, 2-6 Musashidai, Fuchu, Japan. yoshiki@tmin.ac.jp
Yoshiki
Y
Takamatsu
Yasuko
Y
Kishimoto
Shunji
S
Ohsako
D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D001923:Brain Chemistry / Q000502:physiology*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D015816:Cell Adhesion Molecules, Neuronal / Q000502:physiology; D004330:Drosophila / Q000378:metabolism*; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007858:Learning / Q000502:physiology; D008568:Memory / Q000502:physiology; D010766:Phosphorylation; D012160:Retina / Q000378:metabolism; D016707:Tissue Fixation; D014162:Transfection
D000911:Antibodies, Monoclonal; D015816:Cell Adhesion Molecules, Neuronal; C051642:fasciclin II; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases