Cloning and characterization of the Xenopus cyclin-dependent kinase inhibitor p27XIC1.

Abstract

We have isolated a gene encoding Xic-1, a 27-kDa cyclin-dependent kinase (Cdk) inhibitor from Xenopus ovary that shares significant homology with both mammalian CIP1 and Kipl/Kip2. The N-and C-terminal halves of Xic-1 are sufficient for interacting with Cdks and proliferating cell nuclear antigen, respectively. Recombinant Xic-1 inhibits Xenopus cyclin E/Cdk2, cyclin A/Cdk2, and cyclin B/Cdc2 activities, although with quite different IC50 values. Truncation of the N terminus of Xic-1 increases the IC50 value for cyclin A/Cdk2 50-fold with no effect on the inhibition of cyclin E/Cdk2 or cyclin B/Cdc2. Xic-1 inhibits both single-stranded and nuclear DNA synthesis in egg extracts, an effect reversed by proliferating cell nuclear antigen or cyclin E/Cdk2, respectively. These results suggest a function for Xic-1 in the control of DNA synthesis by cyclin E/Cdk2.Cyclin-dependent kinases (Cdks) are required for a number of key cell cycle transitions and are regulated by checkpoints that ensure particular cell cycle events are initiated only with genomic integrity and after the successful completion of prior events. A form of regulation discovered recently involves Cdk inhibitor proteins (CKIs) that bind to and inhibit a wide variety of Cdk complexes (for review, see ref.

Two for two: Cdk2 and its role in centrosome doubling

“…Cdk2-E activity was then specifically inhibited in these extracts by the addition of recombinant D34Xic-1, a NH 3 -terminal truncated form of the Xenopus cyclin dependent kinase inhibitor Xic-1 p27 . At the concentration used D34Xic-1 specifically inhibits the activity of Cdk2-E but not Cdk1-A or Cdk1-B (Su et al, 1995). Cdk2-cyclin A activity was not a factor in these experiments, because Cdk2 does not complex with cyclin A until after the mid-blastula transition in Xenopus .…”

Xenopus hairy2 functions in neural crest formation by maintaining cells in a mitotic and undifferentiated state

“…Antisense RNA probes for in situ hybridization were prepared from templates encoding Xhairy2b (Tsuji et al, 2003), p27 xic1 (Su et al, 1995), ntubulin (Chitnis et al, 1995), Sox2 (Mizuseki et al, 1998), X-Delta-1 (Chitnis et al, 1995), Keratin (Jonas et al, 1985), X-ngnr-1 (Ma et al, 1996), FoxD3 (Sasai et al, 2001), Slug (Mayor et al, 1995), Xtwist (Hopwood et al, 1989), and Xbmp4 (HemmatiBrivanlou and Thomsen, 1995).…”

Nuclear accumulation of cyclin E/Cdk2 triggers a concentration-dependent switch for the destruction of p27 Xic1

“…Xic1 also has a C-terminal QT domain highly homologous to that of p27 Kip1 , containing a homologous CDK phosphorylation site, T205 (Fig. 4B) (21), and five other potential CDK phosphorylation sites. To determine the in vitro cyclin E͞Cdk2 phosphorylation sites, we analyzed tryptic digests of Xic1 phosphorylated in vitro by cyclin E͞Cdk2 by MS.…”