Organelle Atlas APPENDIX TO CHAPTER 4 4A.1

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1 APPENDIX TO CHAPTER 4 This atlas contains images of cellular organelles visualized using a wide variety of reagents and techniques and demonstrates the diversity of methods for exploring the cell. The atlas consists of the figures listed in Table 4A.1. Table 4A.1 Figures in a Figure Organelle(s)/structure(s) Visualized using: Fig. 4A.1 Mitotic spindle and chromosomes γ-tubulin-gfp, anti-β-tubulin antibody, Hoechst Fig. 4A.2 DNA and β-tubulin Hoechst 33342, anti-β-tubulin antibody Fig. 4A.3 Nuclear pore complexes POM121-GFP Fig. 4A.4 Nuclear envelope Lamin B receptor GFP Fig. 4A.5 Myosin, actin stress fibers Anti-myosin antibody, rhodamineconjugated phalloidin Fig. 4A.6 Focal adhesions Anti-vinculin antibody Fig. 4A.7 Intermediate filaments Anti-vimentin antibody Fig. 4A.8 Microtubules and Golgi complex Anti-tubulin and antigalactosyltransferase antibodies Fig. 4A.9 Endoplasmic reticulum DiOC 6 Fig. 4A.10 Endoplasmic reticulum Anti-ER-resident protein antibodies Fig. 4A.11 Lysosomes Rhodamine-ovalbumin Fig. 4A.12 Lysosomes Anti-LEP100 antibodies Fig. 4A.13 Lysosomes, endosomes Rhodamine-conjugated wheat germ agglutinin Fig. 4A.14 Golgi complex Anti mannosidase II antibodies, brefeldin A, nocodazole Fig. 4A.15 Mitochondria TMRE, calcium green AM Fig. 4A.16 Endosomes Anti-transferrin antibodies Fig. 4A.17 Trans-Golgi network Anti-TGN38 antibodies Fig. 4A.18 Trans-Golgi network, endosomes Monoclonal anti-ap1 antibodies (100/3) Fig. 4A.19 Trans-Golgi network, endosomes Anti-TGN46 and anti-ap3 antibodies Fig. 4A.20 Plasma membrane clathrin-coated pits, clathrin-coated vesicles Anti-AP2 antibodies Fig. 4A.21 Plasma membrane clathrin-coated pits, trans-golgi network, endosomes, clathrin-coated vesicles Fibronectin (an extracellular matrix protein), nuclei Monoclonal anti-clathrin antibodies (X22) Fig. 4A.22 Rhodamine-conjugated anti-fibronectin antibody, DAPI a Abbreviations: DAPI, 4,6-diamidino-2-phenylindole; DiOC6, dicarbocyanine dye; GFP, green fluorescent protein; TGN, trans-golgi network; TMRE, tetramethylrhodamine ethylester. (1998) 4A.1-4A.12 Copyright 1998 by John Wiley & Sons, Inc. 4A.1

2 Figure 4A.1 Mitotic NRK cell in metaphase. Red, mouse anti-β-tubulin antibody; green, γ-tubulin-gfp; blue, Hoechst Figure provided by Jan Ellenberg, NICHD/NIH. Figure 4A.2 Distribution of DNA (Hoechst 33342, blue) and β- tubulin (anti-β-tubulin antibody, red) in interphase NRK cells. Figure provided by Jan Ellenberg, NICHD/NIH. 4A.2

3 Figure 4A.3 Nuclear pore complexes in PTK 2 cell stained with POM121 tagged with green fluorescent protein (GFP). Figure provided by Jan Ellenberg, NICHD/NIH. 4A.3

4 Figure 4A.4 Nuclear envelope in COS cell labeled with lamin B receptor tagged with green fluorescent protein (GFP). Figure provided by Jan Ellenberg, NICHD/NIH. 4A.4

5 Figure 4A.5 (A) Myosin localization in formaldehyde-fixed NRK cell labeled with mouse antimyosin antibodies. (B) Stress fibers in the same formaldehyde-fixed NRK cell labeled with rhodamine-conjugated phalloidin. Figure provided by Julie Donaldson, NHBLI/NIH. Figure 4A.6 Focal adhesions in formaldehyde-fixed NRK cell labeled with mouse anti-vinculin antibodies. Figure provided by Julie Donaldson, NHBLI/NIH. Figure 4A.8 Microtubules and Golgi complex in HeLa cell stained with anti-tubulin antibodies (green) and antibodies to the Golgi complex enzyme galactosyl transferase (yellow-orange). Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. Figure 4A.7 Intermediate filaments in formaldehyde-fixed NRK cell labeled with rabbit anti-vimentin antibodies. Figure provided by Julie Donaldson, NHBLI/NIH. 4A.5

6 Figure 4A.9 Endoplasmic reticulum (ER) in frog skin cell labeled with DiOC 6. Figure provided by Mark Terasaki, University of Connecticut Health Center. 4A.6

7 Figure 4A.10 Endoplasmic reticulum (ER) in NRK cell labeled with antibodies to ER-resident proteins (from D. Louvard). Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. Figure 4A.11 Lysosomes in chicken fibroblast labeled with rhodamine-conjugated ovalbumin taken up by the cells over 4 hr. Figure provided by Jennitfer Lippincott- Schwartz, NICHD/NIH. Figure 4A.12 Lysosomes in chicken fibroblast labeled with antibodies to LEP100, a 100-kDa lysosomal membrane protein. Figure provided by Jennifer Lippincott- Schwartz, NICHD/NIH. 4A.7

8 Figure 4A.13 HeLa cell stained with rhodamine-conjugated wheat germ agglutinin (WGA), which binds to sialic acid and N-acetylglucosaminyl residues on glycoproteins. Glycoproteins enriched in these residues are found in lysosomes and endosomes. Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. Figure 4A.14 Distribution of antibodies to the Golgi enzyme mannosidase II in fixed and permeabilized HeLa cells (A) untreated; (B) treated with brefeldin A; or (C) treated with nocodazole, which depolymerizes microtubules. Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. 4A.8

9 Figure 4A.15 Neuroblastoma cell stained with TMRE to label mitochondria (orange) and with calcium green AM to monitor calcium. Figure provided by Ion Moran, University of Connecticut Health Center. Figure 4A.17 Trans-Golgi network (TGN) in NRK cell labeled with antibodies to TGN-38, a membrane protein. Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. Figure 4A.16 Distribution of endosomes in NRK cell incubated in the presence of iron-saturated transferrin and labeled with fluorescent antibodies to transferrin. Figure provided by Jennifer Lippincott-Schwartz, NICHD/NIH. 4A.9

10 Figure 4A.18 Distribution of the adaptor complex AP1 in human fibroblasts stained with monoclonal 100/3 in the trans-golgi network and endosomal structures. Figure provided by Juan Bonifacino, NICHD/NIH. Figure 4A.19 Distribution of the trans-golgi network and the adaptor complex AP3 in human fibroblasts labeled with antibodies to TGN46 (green) and AP3 (red). Figure provided by Juan Bonifacino, NICHD/NIH. Figure 4A.20 Localization of adaptor complex AP2 to plasma membrane clathrin-coated pits and clathrin-coated vesicles in HeLa cells labeled with anti-ap2 antibodies. Figure provided by Juan Bonifacino, NICHD/NIH. 4A.10

11 Figure 4A.21 Clathrin distribution in human fibroblasts labeled with monoclonal antibody X22 (Affinity Bioreagents). Figure provided by Juan Bonifacino, NICHD/NIH. 4A.11

12 Figure 4A.22 Distribution of the extracellular matrix protein fibronectin in cells labeled with rhodamineconjugated antibody to fibronectin (red) and DAPI (blue). Figure provided by Kenneth Yamada, NIDR/NIH. 4A.12