Microtubule-dependent transport

This is indicated by a linear or saturating behavior of as a function of time. In this phase, the particles entrapped in intracellular vesicles diffuse in the cytosol, waiting for an encounter with a motor protein as a microtubule-dependent transporting system. Phase III started with a dramatic increase in particle velocity and directed movement over long distances (several micrometer) within the cell along microtubules. The directed movement is demonstrated by the quadratic dependence of as a function of time with transport velocities up to 4 j,m... 

Figure 4 The Sertoli cell provides trophic factors to various germ cells (spermatogonia, pachytene spermatocyte, round spermatid, and elongate spermatid) by secreting a seminiferous tubule fluid. Microtubule-dependent transport facilitates this supportive role, using microtubule motors to move vesicles along the abundant, radially oriented, Sertoli cell cytoplasmic microtubules. In this model, a secretory granule is moving toward the microtubule (—) end, translocating from the perinuclear Golgi apparatus to the seminiferous tubule lumen. Cytoplasmic dynein, an abundant Sertoli cell protein (3 3 ), is the presumptive motor to catalyze this basal-to-lumenal microtubule-dependent transport.

Although indirect evidence supports the assertion that 2,5-HD alters Sertoli cell microtubule-dependent transport and inhibits seminiferous tubule fluid formation, the molecular connections between these processes remain to be elucidated. 

Kinesins and other molecular motors. Before considering further how the myosin motor may work, we should look briefly at the kinesins, a different group of motor molecules,1683 which transport various cellular materials along microtubule "rails." They also participate in organization of the mitotic spindle and other microtubule-dependent activities.168a/b/C See Section C,2 for further discussion. More than 90 members of the family have been identified. Kinesin heads have much shorter necks than do the myosin heads. 

Figure 28.3. Transport of bile acids and other constituents across the hepatocyte. The Na+ dependent bile salt (taurocholate) transporter (BA-) is shown on the sinusoidal membrane that utihzes the Na+ gradient maintained by the NAK pump, shown here on the lateral aspect of the plasmalemma. Bile salt transcellular transport involves microtubules, which then dehver substrate to the canahcular bile salt transporter (1). Bilary excretion of GSH, gluc-uronate (GluA), and sulfate conjugates of compounds such as 17P-estradiol (E2), bilirubin, and bromosulfothalein (BSP) is catalyzed by the multispecific organic anion transporter (MOAT 2). Both 1 and 2 are members of the ABC family of ATP-dependent transporters that also includes P-glycoprotein (3), another canalicular transporter catalyzing excretion of hpophihc compounds such as the chemotherapeutic drug, daunorubicin.

Cole RW, Ault JG, Hayden JH, et al. 1991. Crocidolite asbestos libers undergo size-dependent microtubule-mediated transport after endocytosis in vertebrate lung epithelial cells. Cancer Res 51 4942-4947. 

Lippincott-Schwartz J, Donaldson JG, Schweizer A, et al. (1990) Microtubule-dependent retrograde transport of proteins into the ER in the presence of brefeldin A suggests an ER recycling pathway. In Cell 60 821 -836. 

Actively transported PEI/DNA nanocomplexes exhibited an average velocity of 0.2 pm/sec [118], a value on the same order of magnitude as motor-protein driven motion. Transport was revealed to be microtubule dependent, because both active transport and perinuclear accumulation were abolished upon microtubule depolymerization. Experiments utilizing MPT to quantify the other intracellular barriers to gene delivery are under way. 

Microtubules provide routes for active transport of molecules and vesicles through the cell. The transport is carried out by microtubule-dependent motor proteins, the cytoplasmid dyneins and kinesins. Cytoplasmic dyneins are closely related to the ciliary dyneins found in cilia and flagella. Different kinesins and to, some extent, dyneins are involved in different cellular processes, such as the separation of chromosomes in mitosis, as well as transport through the cytoplasm. During cytoplasmic transport, one part of the motor protein is attached to the microtubule... 

The membrane tubules and lamellae of the endoplasmic reticulum (ER) are extended in the cell with the use of MTs and actin filaments. Kinesin motors are required for stretching out the ER, whereas depolymerization of microtubules causes the retraction of the ER to the cell centre in an actin-dependent manner. Newly synthesized proteins in the ER are moved by dynein motors along MTs to the Golgi complex (GC), where they are modified and packaged. The resulting vesicles move along the MTs to the cell periphery transported by kinesin motors. MTs determine the shape and the position also of the GC. Their depolymerization causes the fragmentation and dispersal of the GC. Dynein motors are required to rebuild the GC. 

Even though dynein, kinesin, and myosin serve similar ATPase-dependent chemomechanical functions and have structural similarities, they do not appear to be related to each other in molecular terms. Their similarity lies in the overall shape of the molecule, which is composed of a pair of globular heads that bind microtubules and a fan-shaped tail piece (not present in myosin) that is suspected to carry the attachment site for membranous vesicles and other cytoplasmic components transported by MT. The cytoplasmic and axonemal dyneins are similar in structure (Hirokawa et al., 1989 Holzbaur and Vallee, 1994). Current studies on mutant phenotypes are likely to lead to a better understanding of the cellular roles of molecular motor proteins and their mechanisms of action (Endow and Titus, 1992). 

Several aspects of intracellular trafficking should be kept in mind in the intracellular trafficking section. The first is the dependence of acidification of endosomes on the uptake of liposomes. This aspect is sometimes discussed when analyzing clathrin uptake. However, several other pathways are also in need of acidic compartments as a destination of uptake so, we list this factor as an individual aspect. Other aspects of intracellular trafficking that are of interest are the transport from early endosomes to late endosomes, the dependence of actin filaments and dynamin, and/or microtubules. Furthermore, the energy dependence of liposome uptake is discussed. 

This family of ATP-dependent motors [EC 3.6.1.33] that forms ADP and orthophosphate during its energy-dependent translocation along the surface of microtubules. Unique cargo sites on dynein molecules allow for the specific transport of cellular organelles and other macro-molecular components. 

Microtubules in the long axons of nerve cells function as "rails" for the "fast transport" of proteins and other materials from the cell body down the axons. In fact, microtubules appear to be present throughout the cytoplasm of virtually all eukaryotic cells (Fig. 7-32) and also in spirochetes.311 Motion in microtubular systems depends upon motor proteins such as kinesin, which moves bound materials toward what is known as the "negative" end of the microtubule,312 dyneins which move toward the positive end.310 These motor proteins are driven by the Gibbs energy of hydrolysis of ATP or GTP and in this respect, as well as in some structural details (Chapter 19), resemble the muscle protein myosin. Dynein is present in the arms of the microtubules of cilia (Fig. 1-8) whose motion results from the sliding of the microtubules driven by the action of this protein (Chapter 19). 

As gene carriers are internalized by endocytosis, the motion characteristics inside the cell resembles the movement of the endosomal compartments within the cell and the formed vesicles are transported along the microtubule network [38]. Suh et al. [41] quantified the transport of individual internalized polyplexes by multiple-particle tracking and showed that the intracellular transport characteristics of polyplexes depend on spatial location and time posttransfection. Within 30 min, polyplexes accumulated around the nucleus. An average of the transport modes over a 22.5 h period after transfection showed that the largest fraction of polyplexes with active transport was found in the peripheral region of the cells whereas polyplexes close to the nucleus were largely diffusive and subdiffusive. Disruption of the microtubule network by nocodazole completely inhibits active transport and also the perinuclear accumulation of polyplexes [37, 40, 47]. 

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