title: Regulation of late endosomal/lysosomal maturation and trafficking by cortactin affects Golgi morphology.
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.
authors.affiliation: Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA.
authors.forename: Kellye C
authors.initials: KC
authors.lastname: Kirkbride
authors.forename: Nan Hyung
authors.initials: NH
authors.lastname: Hong
authors.forename: Christi L
authors.initials: CL
authors.lastname: French
authors.forename: Emily S
authors.initials: ES
authors.lastname: Clark
authors.forename: W Gray
authors.initials: WG
authors.lastname: Jerome
authors.forename: Alissa M
authors.initials: AM
authors.lastname: Weaver
mesh_terms: D000199:Actins / Q000378:metabolism*; D045744:Cell Line, Tumor; D051356:Cortactin / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission
chemical_list: D000199:Actins; D051356:Cortactin