A dynactin subunit with a highly conserved cysteine-rich motif interacts directly with Arp1.
Dynactin is a multisubunit complex and a required cofactor for most, or all, of the cellular processes powered by the microtubule-based motor cytoplasmic dynein. Using a dynein affinity column, the previously uncharacterized p62 subunit of dynactin was isolated and microsequenced. Two peptide sequences were used to clone human cDNAs encoding p62. Sequence analysis of the predicted human polypeptide of 53 kDa revealed a highly conserved pattern of eleven cysteine residues, eight of which fit the consensus sequence for a Zn(2+)-binding RING domain. We have characterized p62 as an integral component of 20 S dynactin by biochemical and immunocytochemical methods. Affinity chromatography experiments demonstrate that p62 binds directly to the Arp1 subunit of dynactin. Immunocytochemistry with antibodies to p62 demonstrates that this polypeptide has a punctate cytoplasmic distribution as well as centrosomal distribution typical of dynactin. In transfected cells, overexpression of p62 did not disrupt microtubule organization or the integrity of the Golgi but did cause both cytosolic and nuclear distribution of the protein, suggesting that this polypeptide may be targeted to the nucleus at very high expression levels.
Karki
S
S
Department of Animal Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Tokito
M K
MK
Holzbaur
E L
EL
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D051837:COUP Transcription Factor II; D051836:COUP Transcription Factors; D002460:Cell Line; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003545:Cysteine / Q000032:analysis*; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000072159:Dynactin Complex; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011987:Receptors, Steroid; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
D051837:COUP Transcription Factor II; D051836:COUP Transcription Factors; D003545:Cysteine; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins; D011987:Receptors, Steroid; D014157:Transcription Factors; C095569:NR2F2 protein, human