Who Needs Microtubules? Myogenic Reorganization of MTOC, Golgi Complex and ER Exit Sites Persists Despite Lack of Normal Microtubule Tracks

University of Colorado, Boulder, United States of America
PLoS ONE (Impact Factor: 3.23). 12/2011; 6(12):e29057. DOI: 10.1371/journal.pone.0029057
Source: PubMed


A wave of structural reorganization involving centrosomes, microtubules, Golgi complex and ER exit sites takes place early during skeletal muscle differentiation and completely remodels the secretory pathway. The mechanism of these changes and their functional implications are still poorly understood, in large part because all changes occur seemingly simultaneously. In an effort to uncouple the reorganizations, we have used taxol, nocodazole, and the specific GSK3-β inhibitor DW12, to disrupt the dynamic microtubule network of differentiating cultures of the mouse skeletal muscle cell line C2. Despite strong effects on microtubules, cell shape and cell fusion, none of the treatments prevented early differentiation. Redistribution of centrosomal proteins, conditional on differentiation, was in fact increased by taxol and nocodazole and normal in DW12. Redistributions of Golgi complex and ER exit sites were incomplete but remained tightly linked under all circumstances, and conditional on centrosomal reorganization. We were therefore able to uncouple microtubule reorganization from the other events and to determine that centrosomal proteins lead the reorganization hierarchy. In addition, we have gained new insight into structural and functional aspects of the reorganization of microtubule nucleation during myogenesis.

Download full-text


Available from: Tan Zhang
  • Source
    • "Interestingly, in short-time MT regrowth experiments, new MTs not only formed at the perinuclear ring but also appeared as asters growing from cytoplasmic nucleating sites. More than 85% of these MT regrowth foci were identified as Golgi elements that in fused myotubes not only surrounded nuclei but also extended between them [83–85]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: A shared feature among all microtubule (MT)-dependent processes is the requirement for MTs to be organized in arrays of defined geometry. At a fundamental level, this is achieved by precisely controlling the timing and localization of the nucleation events that give rise to new MTs. To this end, MT nucleation is restricted to specific subcellular sites called MT-organizing centres. The primary MT-organizing centre in proliferating animal cells is the centrosome. However, the discovery of MT nucleation capacity of the Golgi apparatus (GA) has substantially changed our understanding of MT network organization in interphase cells. Interestingly, MT nucleation at the Golgi apparently relies on multiprotein complexes, similar to those present at the centrosome, that assemble at the cis-face of the organelle. In this process, AKAP450 plays a central role, acting as a scaffold to recruit other centrosomal proteins important for MT generation. MT arrays derived from either the centrosome or the GA differ in their geometry, probably reflecting their different, yet complementary, functions. Here, I review our current understanding of the molecular mechanisms involved in MT nucleation at the GA and how Golgi- and centrosome-based MT arrays work in concert to ensure the formation of a pericentrosomal polarized continuous Golgi ribbon structure, a critical feature for cell polarity in mammalian cells. In addition, I comment on the important role of the Golgi-nucleated MTs in organizing specialized MT arrays that serve specific functions in terminally differentiated cells.
    Full-text · Article · Sep 2014 · Philosophical Transactions of The Royal Society B Biological Sciences
  • Source
    • "Accordingly, in addition to labeling the endogenous Yip1B protein with the polyclonal antibody we generated, we stained muscle fast fibers from the tibialis anterior and slow fibers from soleus muscles with a monoclonal antibody against the GM130, used as cis-Golgi marker. Confocal analysis revealed the cis-Golgi protein GM130 as trains of spots along the fiber in the slow-twitch soleus muscle and in pairs of spots in the fast-twitch tibialis anterior muscle, as expected (Zaal et al. 2011). Staining of the same muscles with Yip1B revealed spots resembling the GM130 pattern (Fig. 3). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The mechanism of endoplasmic reticulum (ER)-Golgi complex (GC) traffic is conserved from yeast to higher animals, but the architectures and the dynamics of vesicles’ traffic between ER and GC vary across cell types and species. Skeletal muscle is a unique tissue in which ER and GC undergo a structural reorganization during differentiation that completely remodels the secretory pathway. In mature skeletal muscle, the ER is turned into sarcoplasmic reticulum, which is composed of junctional and longitudinal regions specialized, respectively, in calcium release and uptake during contraction. During skeletal muscle differentiation, GC acquires a particular fragmented organization as it appears as spots both at the nuclear poles and along the fibers. The ubiquitary-expressed Yip1A isoform has been proposed to be involved in anterograde trafficking from the ER exit sites to the cis-side of the GC and in ER and GC architecture organization. We investigated the role of Yip1 in skeletal muscle. Here we report that, following skeletal muscle development, the expression of the Yip1A decreases and is replaced by the muscle-specific Yip1B isoform. Confocal microscope analysis revealed that in adult skeletal muscle the Yip1B isoform is localized in the ER-Golgi intermediate and cis-Golgi compartments. Finally, skeletal muscle knockdown experiments in vitro and in vivo suggested that Yip1B is not involved in GC structure maintenance.
    Full-text · Article · Mar 2014 · Histochemie
  • Source
    • "During muscle differentiation the MTOC redistributes from centrosomes to nuclear membranes (Tassin et al., 1985a; Lu et al., 2001; Bugnard et al., 2005; Zaal et al., 2011). EB3-GFP and immobile, highlighting a frame that remains unchanged for minutes (Fig. 1 D1 and Video 3); the color-coded projections are mostly white (Fig. 1 D2). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Skeletal muscle microtubules (MTs) form a nonclassic grid-like network, which has so far been documented in static images only. We have now observed and analyzed dynamics of GFP constructs of MT and Golgi markers in single live fibers and in the whole mouse muscle in vivo. Using confocal, intravital, and superresolution microscopy, we find that muscle MTs are dynamic, growing at the typical speed of ∼9 µm/min, and forming small bundles that build a durable network. We also show that static Golgi elements, associated with the MT-organizing center proteins γ-tubulin and pericentrin, are major sites of muscle MT nucleation, in addition to the previously identified sites (i.e., nuclear membranes). These data give us a framework for understanding how muscle MTs organize and how they contribute to the pathology of muscle diseases such as Duchenne muscular dystrophy.
    Full-text · Article · Oct 2013 · The Journal of Cell Biology
Show more