PKN interacts with a paraneoplastic cerebellar degeneration-associated antigen, which is a potential transcription factor.
PKN is a fatty acid-activated serine/threonine protein kinase, having a catalytic domain homologous to protein kinase C family. PKN has been recently reported to interact with a small GTP-binding protein Rho and cytoskeletal proteins such as neurofilament and alpha-actinin. To identify the new components of the PKN-signaling pathway, the yeast two-hybrid system was employed. Using the amino-terminal regulatory domain of PKN as a bait, cDNA encoding a neural antigen PCD17, which is recognized by characteristic antibodies of patients with paraneoplastic cerebellar degeneration, was isolated from a human brain cDNA library. The interaction between PKN and PCD17 was also determined by the in vitro binding analysis. PCD17 was coimmunoprecipitated with PKN from the lysate of COS7 cells transfected with both expression constructs for PKN and the amino-terminal region of PCD17. PCD17 was phosphorylated by PKN, and the extent of this phosphorylation was enhanced by addition of 40 microM arachidonic acid. The amino-terminal region of PCD17 could form a homodimer in vitro, and PCD17 fused to the Gal4 DNA binding domain showed the transcriptional transactivation of the chloramphenicol acetyltransferase reporter gene linked to 5 Gal4 binding sites and minimal promoter in rat C6 glioma cells. These results suggest the participation of PCD17 in gene expression and lead to a clue for elucidating the PKN signaling pathway from the cytosol to the nucleus.
Takanaga
H
H
Faculty of Science, Kobe University, Kobe, 657, Japan.
Mukai
H
H
Shibata
H
H
Toshimori
M
M
Ono
Y
Y
D000818:Animals; D019556:COS Cells; D018014:Gene Transfer Techniques; D006801:Humans; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D011485:Protein Binding; D011493:Protein Kinase C; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
C095184:CDR2 protein, human; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; C061570:protein kinase N; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C