docPrednáška 3 Mikrofilamenty III
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Prednáška 3 Mikrofilamenty III
Prednáška 3 Mikrofilamenty III Roles of microfilaments in eukaryotic cell: Cell division Cell adhesion Cell movement Cell shape Actin filaments in cell movement The acrosome reaction in echinoderm sperm Acrosomal process Polymerized actin Acrosomal vesicle Fascin Cross-links Acromere Periacrosomal region containing profilin-actin complexes Nucleus UNACTIVATED ACTIVATED Movement of Lysteria in infected fibroblasts Actin filaments assemble at the leading edge of migrating cells Actin filaments assemble at the leading edge of migrating cells Microfilaments & cell motility A model of cell movement mediated by the actin-rich cortex actin cortex lamellipodium substratum Cell signaling and cytoskeleton organization Growth factors Cdc42 Filopodia LPA Rac Rho Cofilin Myosin II Lamellipodia Stress fibres LPA = lysophosphatidic acid The effects of Cdc42, Rac and Rho activation on actin organization in fibroblasts Quiescent cells +Cdc42 +Rac +Rho Involvement of microfilaments in neutrophil polarization and chemotaxis formyl-Met-Leu-Phe Myosin – dependent dynamics of actin cytoskelet Rotary movement of mitochondrial F1FOATPase – an example of molecular motor in action ADP+Pi → ATP H+ matrix H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+H+ H+ H+ H+H+ H+ H+ H+ Rotary movement of mitochondrial F1FOATPase – experimental demonstration Rotary movement of mitochondrial F1FOATPase – experimental demonstration Noji et al. (1997). Nature 386: 299-302 Myosin – dependent dynamics of actin cytoskelet Myosin – another type of a molecular motor Head Neck Tail MYOSIN I Calmodulin light chain MYOSIN II 130 nm Essential light chain Vesicular transport Actin-plasma membrane interactions Muscle contraction Cytokinesis Regulatory light chain MYOSIN V Calmodulin light chain Vesicular transport Actin-plasma membrane interactions 3D structure of the myosin head domain Myosin movement along an actin filament Myosin movement along an actin filament Myosin cycle – Animation I The effect of lever arm length on the step size The sliding-filament assay The sliding-filament assay 0 sec 0.6 sec 1.2 sec 1.8 sec 4 µm/sec The Sliding-Filament Assay – Animation Some numbers: Rates of myosin-driven actin filament movement: ~0.04-4.5 µm/sec Length of one myosin chain: ~100 nm If scaled up to a human dimensions (~180 cm) …then it moves ~600 m/sec (2000 km/hour) How to determine myosin-generated force? Actin Myosin Optical trap „off“ Optical trap „on“ Latex bead Optical traps: Carlos Bustamante, http://alice.berkeley.edu/ http://www.st-andrews.ac.uk/~atomtrap/Resources.htm Roles of myosin-I, -II and -V in a eukaryotic cell Skeletal muscle myofibrils Skeletal muscle myofibrils Stabilization of the actin thin filaments by capping proteins Z disk CapZ Myosin thick filament Actin thin filament Tropomodulin Z disk Contraction of the striated muscle by sliding-filament Myosin cycle – Animation II Involvement of myosin in cell locomotion Actin – blue; myosin II – pink A model of cell movement mediated by the actin-rich cortex actin cortex lamellipodium substratum cortex under tension actin polymerization at plus end movement of unpolymerized actin retraction focal contacts Cell Motility Animation Actin and myosin in nonmuscle cells contractile ring Myosin II is required for cytokinesis Polarized mRNA localization in the yeast bud tip ash mRNA a a switching allowed a or α a switching not allowed a Actin dynamics in Saccharomyces cerevisiae Polarized mRNA localization in the yeast bud tip Unexpected roles of actin in a nucleus SRF = serum response factor MAL = co-activator of SRF Crm1 = exportin Vartiainen et al. (2007). Science 316: 1749-1752. Unexpected roles of actin in a nucleus CTD, COOH-terminal domain of the largest subunit of RNA polymerase II; HAT, histone acetyltransferase; hnRNP, heterogeneous nuclear RNP; J. Cell Biol. 172: 967-971 (2006). Various forms of actin microfilaments in metazoan cells Filopodium Lamellipodium Lamellum Microvillus Cortical actin Podosome Endosome Phagocytic cup Endocytic pit Golgi ass.-actin Nuclear actin Ruffle Stress fibre Cadherin-based adherent junction Chhabra & Higgs (2007). Nature Cell Biol. 9: 1110-1121.
