Get access

Regulation of late endosomal/lysosomal maturation and trafficking by cortactin affects Golgi morphology

Authors

  • Kellye C. Kirkbride,

    1. Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
    Search for more papers by this author
    • K.C.K. and N.H.H. contributed equally to this work.

  • Nan Hyung Hong,

    1. Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
    Search for more papers by this author
    • K.C.K. and N.H.H. contributed equally to this work.

  • Christi L. French,

    1. Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
    Search for more papers by this author
  • Emily S. Clark,

    1. Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
    Current affiliation:
    1. University of Miami Medical School, Miami, Florida 33101
    Search for more papers by this author
  • W. Gray Jerome,

    1. Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee
    Search for more papers by this author
  • Alissa M. Weaver

    Corresponding author
    1. Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
    2. Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee
    • Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee 37232
    Search for more papers by this author

  • Monitoring Editor: Bruce Goode

Abstract

Cortactin is a branched actin regulator and tumor-overexpressed protein that promotes vesicular trafficking at a variety of cellular sites, including endosomes and the trans-Golgi network. To better understand its role in secretory trafficking, we investigated its function in Golgi homeostasis. Here, we report that knockdown (KD) of cortactin leads to a dramatic change in Golgi morphology by light microscopy, dependent on binding the Arp2/3 actin-nucleating complex. Surprisingly, there was little effect of cortactin-KD on anterograde trafficking of the constitutive cargo vesicular stomatitis virus glycoprotein (VSVG), Golgi assembly from endoplasmic reticulum membranes upon Brefeldin A washout, or Golgi ultrastructure. Instead, electron microscopy studies revealed that cortactin-KD cells contained a large number of immature-appearing late endosomal/lysosomal (LE/Lys) hybrid organelles, similar to those found in lysosomal storage diseases. Consistent with a defect in LE/Lys trafficking, cortactin-KD cells also exhibited accumulation of free cholesterol and retention of the retrograde Golgi cargo mannose-6-phosphate receptor in LE. Inhibition of LE maturation by treatment of control cells with Rab7 siRNA or chloroquine led to a compact Golgi morphology similar to that observed in cortactin-KD cells. Furthermore, the Golgi morphology defects of cortactin-KD cells could be rescued by removal of cholesterol-containing lipids from the media, suggesting that buildup of cholesterol-rich membranes in immature LE/Lys induced disturbances in retrograde trafficking. Taken together, these data reveal that LE/Lys maturation and trafficking are highly sensitive to cortactin-regulated branched actin assembly and suggests that cytoskeletal-induced Golgi morphology changes can be a consequence of altered trafficking at late endosomes. © 2012 Wiley Periodicals, Inc.

Ancillary