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WBPicture0000009601 Description: Fig. 5. SAD-1 Expression and Protein Localization. (A-D) sad-1 regulatory sequences were used to drive the expression of GFP (A). GFP was fused in frame to the second exon of sad-1. psad-1GFP expression was seen exclusively in the nervous system beginning in the late embryo (B) and continuing through the life of the animal (C, D). (E, G) Anti-SAD-1 antibody staining of animals overexpressing SAD-1 from the endogenous promoter. (F, H) pstr-3SNB-1::GFP expression in the same animals. (E) In some animals, fluorescence was confined to axons of the nerve ring (thick arrow) and ventral (white arrowhead) and dorsal (yellow arrowhead) cords and to cell bodies (asterisk). No staining was observed in the sensory dendrites (thin arrow) or axon commissures. In these animals, ASI neuron morphology was normal (F). (G) In other animals, fluorescence was present in the dendrites (thin arrow) and commissures (yellow arrow) in addition to the axon and cell body staining seen in (E). (H) In these animals, axons were terminated (thick arrow) or otherwise defective, and some vesicle clusters appeared in the dendrite (thin arrow). (I) A full-length SAD-1::GFP fusion protein was expressed in the nervous system using the unc-115 promoter. GFP was inserted in frame at amino acid 566 of the SAD-1 protein. Sequences 5' to GFP were from the sad-1 cDNA, and sequences 3' were from the sad-1 genomic clone. Shading of domain structure as in Fig. 4(B), GFP in green. (J) SAD-1::GFP localized to puncta within the axons of the nerve ring (thick arrow) and ventral (white arrowhead) and dorsal (yellow arrowhead) nerve cords but was faint or absent in the dendrites (thin arrow) and synapse-poor commissures. Additionally, SAD-1::GFP was observed in cell bodies, where it was excluded from the nucleus (asterisk). These animals were fixed with formaldehyde to eliminate gut autofluorescence. (K) A higher magnification of the ventral cord in animals expressing punc-115SAD-1::GFP (green) and costained with anti-SYD-2 (red) antibodies. Little colocalization of SAD-1::GFP and SYD-2 was evident (e.g., arrowhead). (L) A higher magnification of the dorsal cord in animals overexpressing SAD-1 from its own promoter and costained with anti-SAD-1 (red) and anti-SNT-1 (green) antibodies. Many SAD-1 and SNT-1 puncta overlapped (yellow color/white arrowheads) or were adjacent (red arrowheads). Most SNT-1 puncta were associated with SAD-1, but some SAD-1 puncta were not associated with SNT-1 (yellow arrowheads).
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  Acknowledgment: Template: Reprinted from <Journal_URL>, <Article_URL>, Copyright <Publication_year>, with permission from <Publisher_URL>.
    Publication_year: 2001
    Article_URL: DOI id 10.1016/S0896-6273(01)00184-2
    Journal_URL: Neuron
    Publisher_URL: Elsevier
  Reference: WBPaper00004523