GSBS Student Publications

Title

Molecular characterization of the 50-kD subunit of dynactin reveals function for the complex in chromosome alignment and spindle organization during mitosis

GSBS Program

Biochemistry & Molecular Pharmacology

UMMS Affiliation

Graduate School of Biomedical Sciences; Department of Cell Biology

Date

2-1-1996

Document Type

Article

Medical Subject Headings

Amino Acid Sequence; Animals; Base Sequence; Cattle; Cell Line; *Chromosomes; Cloning, Molecular; DNA, Complementary; Dynein ATPase; Humans; Male; Microtubule Proteins; *Microtubule-Associated Proteins; *Mitosis; Molecular Sequence Data; Rats; Rats, Sprague-Dawley; Sequence Analysis

Disciplines

Life Sciences | Medicine and Health Sciences

Abstract

Dynactin is a multi-subunit complex which has been implicated in cytoplasmic dynein function, though its mechanism of action is unknown. In this study, we have characterized the 50-kD subunit of dynactin, and analyzed the effects of its overexpression on mitosis in living cells. Rat and human cDNA clones revealed p50 to be novel and highly conserved, containing three predicted coiled-coil domains. Immunofluorescence staining of dynactin and cytoplasmic dynein components in cultured vertebrate cells showed that both complexes are recruited to kinetochores during prometaphase, and concentrate near spindle poles thereafter. Overexpression of p50 in COS-7 cells disrupted mitosis, causing cells to accumulate in a prometaphase-like state. Chromosomes were condensed but unaligned, and spindles, while still bipolar, were dramatically distorted. Sedimentation analysis revealed the dynactin complex to be dissociated in the transfected cultures. Furthermore, both dynactin and cytoplasmic dynein staining at prometaphase kinetochores was markedly diminished in cells expressing high levels of p50. These findings represent clear evidence for dynactin and cytoplasmic dynein codistribution within cells, and for the presence of dynactin at kinetochores. The data also provide direct in vivo evidence for a role for vertebrate dynactin in modulating cytoplasmic dynein binding to an organelle, and implicate both dynactin and dynein in chromosome alignment and spindle organization.

Rights and Permissions

Citation: J Cell Biol. 1996 Feb;132(4):617-33.

Related Resources

Link to article in PubMed

Journal Title

The Journal of cell biology

PubMed ID

8647893