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Organelle structure Nucleus Sarco(endo)plasmic reticulum Mitochondria Peroxisome Autophagosome.

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Presentation on theme: "Organelle structure Nucleus Sarco(endo)plasmic reticulum Mitochondria Peroxisome Autophagosome."— Presentation transcript:

1 Organelle structure Nucleus Sarco(endo)plasmic reticulum Mitochondria Peroxisome Autophagosome

2 Electron microscope Diffraction limit – Wave nature of light passing through lens – Optical: ~0.6 um, resolution ~0.2 um – Confocal, FT, nanohole arrays De Broglie wavelength – = h/mv – Electron: = 1.2(V - ½ ) nm (V: accelerating voltage) B~ /(2 NA)

3 EM imaging Essentially a CRT – Vacuum – Focus e-beam on specimen – Expand onto screen Sample prep – Dehydrate & fix – Heavy metal stain (uranium) – 90 nm slices (Graham Colm, wikipedia)

4 Structure Myofibril Nucleus Mitochondria Artifact Out of plane

5 Eisenberg et al., 1974

6 Common features Densely packed myofibrils Subsarcolemmal space – Nuclei – Mitochondria Intermyofibrillar I-band space – Mitochondria – SR/T-tubule triads

7 Fractional composition Stereology – Statistical model of 3-D structure – 2-D images & discrete sampling CompartmentGuinnea pigToadfish swimbladder Rattlesnake tail shaker Myofibrils85-90%70-80%32% SR3.5-4.5%26% Mitochondrial2-5% (18% SS, 3- 4% core) 1-4% (female- male) 26% T-system0.15-0.25% Nuclei0.9%

8 Rattlesnake tail muscle: prominent extensive glycogen & mitochondria; relatively few myofibrils Clark & Schultz, 1980

9 Nucleus Multiple – MND 3e4-5e4 um 3 /nucleus – Fiber volume 5e5-5e6 um3 (10-1000 nuclei) Cylindrical, ~30 um 3 Nucleolus Cajal bodies DNA RNA processing Terada et al., 2010

10 RNA processing Nucleolus: rRNA synthesis & ribosome assy Cajal bodies: Spliceosome assembly Speckles: RNA splicing Pollard & Earnshaw, 2008

11 Ribosomal biogenesis Multiple (300+) copies of rRNA strand – Transcribed – Cleaved to 28S, 18S, 5.8S strands – Ribose methylation – Uridine isomerization to pseudouridine Ribosomal proteins – Imported to nucleus – Anneal to rRNA in nucleolus

12 Nuclear Envelope Dual membrane Control nuclear import/export Nuclear pore complex

13 Nuclear membrane proteins Chromatin anchors – Lamin, lamin B receptor, emerin Transport proteins – Importin, Exportin, Ran

14 Mitochondria Dual membrane Oxidative ATP synthesis – Citric acid cycle – Electron transport chain Separate genome – Most Mt proteins come from nuclear DNA

15 Mitochondrial syncytium Network, not bacterium-like Regulated growth – Division – Fusion Apoptosis Manella, 2000

16 Muscle mitochondria Extensive subsarcolemmal syncitia Elongated tubules, perpendicular to fiber – Frequently doubled around Z-disks Ogata & Yamasaki, 1997 Subsarcolemmal mitochondria Intermyofibrillar mitochondria

17 Inner membrane Cristae – “Shelf-like” – Tubular network Manella, 2008

18 Oxidative ATP Synthesis Electron transport chain – NADH – FADH2 – Ubiquinone – Cytochrome C Reactive oxygen

19 Peroxisome Single membrane Controlled H2O2 chemistry – Oxidases generate H2O2 – Peroxidases degrade H2O2 – Fatty acid metabolism – Cholesterol & bile synthesis Peroxins (PEX) control protein composition and biogenesis

20 Sarcoplasmic Reticulum Single membrane – Lamellar, fenestrated – Terminal cisternae Synthesis of transmembrane and extracellular proteins Calcium storage

21 SR T-tubule communication Terminal cisternae wrap T-tubule Communication of membrane events to SR Ogata & Yamasaki, 1997 SEM of I-band region of a myofiber with myofibrils extracted, leaving only membrane-bounded skeletons of T- tubule (T), SR (blue), terminal cisternae (arrowheads), and mitochondria. Fiber axis is vertical.

22 Triad Physical association between T-Tubule and SR “Feet” – Dihydropyridine receptor (DHPR) – Ryanodine Receptor (RyR) Franzini-Armstrong, 1970

23 “white” vs “red” muscle Red: larger, more extensive mitochondria White: (slightly) more extensive SR

24 Autophagosome Double membrane – Small (1 um), spherical Self-engulfment Recruit lysosome & degrade Mizushima & al., 2002

25 Autolysis Degradation of large structures Starvation defense Infection defense Targeting – Apg5/Apg12 targeting complex – LC3 microtubule associated light chain – PI3k, FOXO3

26 Summary Specialized chemistry is constrained to isolated cellular domains Nucleus Mitochondria ER/SR Autophagosome Peroxisome

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