Transforming acidic coiled-coil-containing protein 4 interacts with centrosomal AKAP350 and the mitotic spindle apparatus.
AKAP350 is a multiply spliced family of 350-450-kDa protein kinase A-anchoring proteins localized to the centrosomes and the Golgi apparatus. Using AKAP350A as bait in a yeast two-hybrid screen of a rabbit parietal cell library, we have identified a novel AKAP350-interacting protein, transforming acidic coiled-coil-containing protein 4 (TACC4). Two-hybrid binary assays demonstrate interaction of both TACC3 and TACC4 with AKAP350A and AKAP350B. Antibodies raised to a TACC4-specific peptide sequence colocalize TACC4 with AKAP350 at the centrosome in interphase Jurkat cells. Mitotic cell staining reveals translocation of TACC4 from the centrosome to the spindle apparatus with the majority of TACC4 at the spindle poles. Truncated TACC4 proteins lacking the AKAP350 minimal binding domain found in the carboxyl coiled-coil region of TACC4 could no longer target to the centrosome. Amino-truncated TACC4 proteins could no longer target to the spindle apparatus. Further, overexpression of TACC4 fusion proteins that retained spindle localization in mitotic cells resulted in an increased proportion of cells present in prometaphase. We propose that AKAP350 is responsible for sequestration of TACC4 to the centrosome in interphase, whereas a separate TACC4 domain results in functional localization of TACC4 to the spindle apparatus in mitotic cells.
Steadman
Brent T
BT
Department of Medicine, Institute of Molecular Medicine and Genetics, Medical College of Georgia and the Augusta Veterans Affairs Medical Center, Augusta, Georgia 30912, USA.
Schmidt
P Henry
PH
Shanks
Ryan A
RA
Lapierre
Lynne A
LA
Goldenring
James R
JR
D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D018385:Centrosome / Q000378:metabolism*; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins; D004247:DNA; D018076:DNA, Complementary; D006801:Humans; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D008941:Spindle Apparatus / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D054758:A Kinase Anchor Proteins; C119719:AKAP9 protein, human; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D018076:DNA, Complementary; D004247:DNA