In vitro phosphorylation of potential cyclin E–Cdk2 substrates by different cyclin–Cdk complexes
| Potential cyclin E–Cdk2 substrate | Phosphorylation sites and cyclin binding motifs in the RZPD clone | Phosphorylation of the RZPD clone by kinasesa | ||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Gene | ORF of RZPD clone | Name/biological process | min: S/TP | cons: S/TPXK/R | RXL | D1–K4 | D3–K4 | E–K2 | A–K2 | B–K1 | ||||||||
| NUMA1b | 245 aa C-term | Nuclear mitotic apparatus protein 1/ spindle apparatus organization | 7 | 1 | 0 | − | ± | + | + | − | ||||||||
| LOC201191 | 231 aa C-term | Hypothetical protein LOC201191/unknown | 6 | 1 | 2 | − | ± | + | + | − | ||||||||
| PIP5K1C | 200 aa C-term | Phosphatidylinositol-4-phosphate 5 kinase, type I γ/phosphatidylinositol signaling | 7 | 2 | 1 | ± | + | + | + | ± | ||||||||
| CCNL2 | 136 aa C-term | cyclin L2 / RNA processing | 7 | 3 | 2 | − | − | ± | + | ± | ||||||||
| RASL11B | 118 aa C-term | RAS-like, family 11, member B/signaling? | 2 | 1 | 0 | − | + | + | + | − | ||||||||
| SIRT2 | full-lengthc | Sirtuin (silent mating type information regulation 2 homologue) 2 (S. cerevisiae)/ NAD+-dependent histone/protein deacetylase | 3 | 1 | 1 | − | − | + | + | ± | ||||||||
| FLJ13111 | full-length | Hypothetical protein FLJ13111/unknown | 4 | 2 | 4 | − | ± | + | + | ± | ||||||||
| SUPT5Hb | 120 aa C-term | Suppressor of Ty 5 homologue (S. cerevisiae)/transcription | 1 | 1 | 1 | − | − | ± | − | − | ||||||||
| STMN2b | full-length | Stathmin-like 2/microtubule destabilization in neuronal growth | 2 | 1 | 1 | − | + | + | + | − | ||||||||
| LOC146909 | 114 aa C-term | Hypothetical protein LOC146909/unknown | 1 | 0 | 1 | − | + | + | + | − | ||||||||
| EEF1G | 211 aa C-term | Eukaryotic translation elongation factor 1γ/translation | 2 | 1 | 1 | − | ± | + | + | ± | ||||||||
| TRIT1 | 99 aa C-term | tRNA isopentenyltransferase 1/ RNA modification | 3 | 1 | 1 | − | + | + | + | ± | ||||||||
| LOC388799 | 89 aa C-term | Hypothetical protein LOC388799/unknown | 2 | 1 | 0 | − | ± | + | + | ± | ||||||||
| STUB1 | full-length | STIP1 homology and U-box–containing protein 1/protein turnover | 4 | 1 | 3 | − | + | + | + | ± | ||||||||
| SRRM2 | 98 aa central | Serine/arginine repetitive matrix 2/ RNA processing | 3 | 0 | 0 | − | − | + | + | + | ||||||||
| TALDO1 | full-length | Transaldolase 1/metabolism | 3 | 1 | 6 | − | + | + | + | − | ||||||||
| SFRS1 | 243 aa C-term | Splicing factor, arginine/serine-rich 1 (splicing factor 2, alternate splicing factor)/RNA processing | 4 | 1 | 1 | − | + | + | + | − | ||||||||
| SNAPAP | full-length | SNAP-associated protein/exocytosis of synaptic vesicles | 2 | 1 | 2 | + | ± | + | ||||||||||
| LOC339287 | 90 aa central | Hypothetical protein LOC339287/unknown | 5 | 4 | 1 | + | + | + | + | + | ||||||||
| FLJ12949 | 248 aa C-term | Hypothetical protein FLJ12949/unknown | 4 | 1 | 3 | − | + | + | − | − | ||||||||
| MAP2 | 376 aa C-term | MAP2/microtubule assembly in neurogenesis | 16 | 3 | 1 | − | + | + | + | − | ||||||||
| RPL14 | 211 aa C-term | Ribosomal protein L14/translation | 1 | 1 | 1 | − | ± | − | − | − | ||||||||
| FNBP3 | 245 aa C-term | Formin binding protein 3/splicing? | 2 | 1 | 1 | − | ± | ± | ± | − | ||||||||
| PRC1d | full-length | Protein regulating cytokinesis 1/ cytokinesis | 5 | 2 | 2 | − | + | + | + | ± | ||||||||
| FLJ13305 | 162 aa central | Hypothetical protein FLJ13305/unknown | 4 | 0 | 3 | − | + | + | ± | − | ||||||||
| ING5 | full-lengthc | Inhibitor of growth family, member 5/ p53 pathway, replication | 2 | 1 | 3 | − | − | + | + | − | ||||||||
| Potential cyclin E–Cdk2 substrate | Phosphorylation sites and cyclin binding motifs in the RZPD clone | Phosphorylation of the RZPD clone by kinases | ||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Gene | ORF of RZPD clone | Name/biological process | min: S/TP | cons: S/TPXK/R | RXL | D1–K4 | D3–K4 | E–K2 | A–K2 | B–K1 | ||||||||
| NUMA1 | 245 aa C-term | Nuclear mitotic apparatus protein 1/ spindle apparatus organization | 7 | 1 | 0 | − | ± | + | + | − | ||||||||
| LOC201191 | 231 aa C-term | Hypothetical protein LOC201191/unknown | 6 | 1 | 2 | − | ± | + | + | − | ||||||||
| PIP5K1C | 200 aa C-term | Phosphatidylinositol-4-phosphate 5 kinase, type I γ/phosphatidylinositol signaling | 7 | 2 | 1 | ± | + | + | + | ± | ||||||||
| CCNL2 | 136 aa C-term | cyclin L2 / RNA processing | 7 | 3 | 2 | − | − | ± | + | ± | ||||||||
| RASL11B | 118 aa C-term | RAS-like, family 11, member B/signaling? | 2 | 1 | 0 | − | + | + | + | − | ||||||||
| SIRT2 | full-length | Sirtuin (silent mating type information regulation 2 homologue) 2 ( | 3 | 1 | 1 | − | − | + | + | ± | ||||||||
| FLJ13111 | full-length | Hypothetical protein FLJ13111/unknown | 4 | 2 | 4 | − | ± | + | + | ± | ||||||||
| SUPT5H | 120 aa C-term | Suppressor of Ty 5 homologue ( | 1 | 1 | 1 | − | − | ± | − | − | ||||||||
| STMN2 | full-length | Stathmin-like 2/microtubule destabilization in neuronal growth | 2 | 1 | 1 | − | + | + | + | − | ||||||||
| LOC146909 | 114 aa C-term | Hypothetical protein LOC146909/unknown | 1 | 0 | 1 | − | + | + | + | − | ||||||||
| EEF1G | 211 aa C-term | Eukaryotic translation elongation factor 1γ/translation | 2 | 1 | 1 | − | ± | + | + | ± | ||||||||
| TRIT1 | 99 aa C-term | tRNA isopentenyltransferase 1/ RNA modification | 3 | 1 | 1 | − | + | + | + | ± | ||||||||
| LOC388799 | 89 aa C-term | Hypothetical protein LOC388799/unknown | 2 | 1 | 0 | − | ± | + | + | ± | ||||||||
| STUB1 | full-length | STIP1 homology and U-box–containing protein 1/protein turnover | 4 | 1 | 3 | − | + | + | + | ± | ||||||||
| SRRM2 | 98 aa central | Serine/arginine repetitive matrix 2/ RNA processing | 3 | 0 | 0 | − | − | + | + | + | ||||||||
| TALDO1 | full-length | Transaldolase 1/metabolism | 3 | 1 | 6 | − | + | + | + | − | ||||||||
| SFRS1 | 243 aa C-term | Splicing factor, arginine/serine-rich 1 (splicing factor 2, alternate splicing factor)/RNA processing | 4 | 1 | 1 | − | + | + | + | − | ||||||||
| SNAPAP | full-length | SNAP-associated protein/exocytosis of synaptic vesicles | 2 | 1 | 2 | + | ± | + | ||||||||||
| LOC339287 | 90 aa central | Hypothetical protein LOC339287/unknown | 5 | 4 | 1 | + | + | + | + | + | ||||||||
| FLJ12949 | 248 aa C-term | Hypothetical protein FLJ12949/unknown | 4 | 1 | 3 | − | + | + | − | − | ||||||||
| MAP2 | 376 aa C-term | MAP2/microtubule assembly in neurogenesis | 16 | 3 | 1 | − | + | + | + | − | ||||||||
| RPL14 | 211 aa C-term | Ribosomal protein L14/translation | 1 | 1 | 1 | − | ± | − | − | − | ||||||||
| FNBP3 | 245 aa C-term | Formin binding protein 3/splicing? | 2 | 1 | 1 | − | ± | ± | ± | − | ||||||||
| PRC1 | full-length | Protein regulating cytokinesis 1/ cytokinesis | 5 | 2 | 2 | − | + | + | + | ± | ||||||||
| FLJ13305 | 162 aa central | Hypothetical protein FLJ13305/unknown | 4 | 0 | 3 | − | + | + | ± | − | ||||||||
| ING5 | full-length | Inhibitor of growth family, member 5/ p53 pathway, replication | 2 | 1 | 3 | − | − | + | + | − | ||||||||
The kinases used are abbreviated (e.g., D1–K4 for cyclin D1–Cdk4). The substrates are phosphorylated strongly (+), weakly (±), or not at all (−) by the respective kinase.
Previously reported Cdk substrate (Stachora et al., 1997; Gavet et al., 1998; Sun and Schatten, 2006).
For ING5 and SIRT2, GST-tagged full-length proteins were used in the kinase assays.
Previously reported cyclin E–Cdk2 substrate (Jiang et al., 1998).
Sharing content requires targeting cookies to be enabled. Please update your cookie preferences to use this feature.