FF domains of CA150 bind transcription and splicing factors through multiple weak interactions.
The human transcription factor CA150 modulates human immunodeficiency virus type 1 gene transcription and contains numerous signaling elements, including six FF domains. Repeated FF domains are present in several transcription and splicing factors and can recognize phosphoserine motifs in the C-terminal domain (CTD) of RNA polymerase II (RNAPII). Using mass spectrometry, we identify a number of nuclear binding partners for the CA150 FF domains and demonstrate a direct interaction between CA150 and Tat-SF1, a protein involved in the coupling of splicing and transcription. CA150 FF domains recognize multiple sites within the Tat-SF1 protein conforming to the consensus motif (D/E)(2/5)-F/W/Y-(D/E)(2/5). Individual FF domains are capable of interacting with Tat-SF1 peptide ligands in an equivalent and noncooperative manner, with affinities ranging from 150 to 500 microM. Repeated FF domains therefore appear to bind their targets through multiple weak interactions with motifs comprised of negatively charged residues flanking aromatic amino acids. The RNAPII CTD represents a consensus FF domain-binding site, contingent on generation of the requisite negative charges by phosphorylation of serines 2 and 5. We propose that CA150, through the dual recognition of acidic motifs in proteins such as Tat-SF1 and the phosphorylated CTD, could mediate the recruitment of transcription and splicing factors to actively transcribing RNAPII.
Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Ave., Toronto, Ontario, Canada, M5G 1X5.
Matthew J
MJ
Smith
Sarang
S
Kulkarni
Tony
T
Pawson
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D006801:Humans; D007700:Kinetics; D008024:Ligands; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry / Q000378:metabolism; D012326:RNA Splicing / Q000235:genetics; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D035602:Transcriptional Elongation Factors
D008024:Ligands; D009687:Nuclear Proteins; D010446:Peptide Fragments; C493526:Tat-specific factor 1, mouse; C473940:Tcerg1 protein, mouse; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D010768:Phosphoserine; D012319:RNA Polymerase II