Phosphorylation of the Fas associated factor FAF1 by protein kinase CK2 and identification of serines 289 and 291 as the in vitro phosphorylation sites

We previously identified the human Fas associated factor (FAF1) as one of t he interacting partners of protein kinase CK2 beta subunit. Since FAF1 is a phosphoprotein we investigated whether it is: a substrate for CK2. Here, w e report the full length human FAF1 cDNA sequence, expnsslon of FAF1 in Esc herichia coli and purification and characterization of FAF1 as a substrate for CK2. FAF1 as well as an N-terminal 40 kDa degradation product serve as substrates for both the recombinant CK2 holoenzyme (k(m), 100 muM) and the isolated catalytic alpha subunit (k(m) 200 muM). Despite the high k(m) valu es, we obtained evidence that CK2 is the major cellular kinase responsible for FAF1 phosphorylation, using tissue extracts as kinase sources. By MALDI -MS we identified the two serine residues at positions 289 and 291 as the m ajor in vitro CK2 phosphorylation sizes. These data may help us elucidate t he functions of FAF1 and the involvement of CK2 mediated phosphorylation in processes such as apoptotic signaling, ubiquitination, nuclear translocati on and embryonic development. (C) 2001 Elsevier Science Ltd. All rights res erved.