Application of active and kinase-deficient kinome collection for identification of kinases regulating hedgehog signaling.
To allow genome-scale identification of genes that regulate cellular signaling, we cloned >90% of all human full-length protein kinase cDNAs and constructed the corresponding kinase activity-deficient mutants. To establish the utility of this resource, we tested the effect of expression of the kinases on three different cellular signaling models. In all screens, many kinases had a modest but significant effect, apparently due to crosstalk between signaling pathways. However, the strongest effects were found with known regulators and novel components, such as MAP3K10 and DYRK2, which we identified in a mammalian Hedgehog (Hh) signaling screen. DYRK2 directly phosphorylated and induced the proteasome-dependent degradation of the key Hh pathway-regulated transcription factor, GLI2. MAP3K10, in turn, affected GLI2 indirectly by modulating the activity of DYRK2 and the known Hh pathway component, GSK3beta. Our results establish kinome expression screening as a highly effective way to identify physiological signaling pathway components and genes involved in pathological signaling crosstalk.
Varjosalo
Markku
M
Department of Molecular Medicine, National Public Health Institute (KTL), FI00290 Helsinki, Finland.
Björklund
Mikael
M
Cheng
Fang
F
Syvänen
Heidi
H
Kivioja
Teemu
T
Kilpinen
Sami
S
Sun
Zairen
Z
Kallioniemi
Olli
O
Stunnenberg
Hendrik G
HG
He
Wei-Wu
WW
Ojala
Päivi
P
Taipale
Jussi
J
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D015723:Gene Library; D053823:Hedgehog Proteins / Q000378:metabolism*; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D008322:Mammals; D051379:Mice; D041681:NIH 3T3 Cells; D015513:Oncogene Proteins / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D015398:Signal Transduction; D015534:Trans-Activators / Q000378:metabolism; D014709:Vero Cells; D000071676:Zinc Finger Protein GLI1; D000075746:Zinc Finger Protein Gli2; D000097603:Dyrk Kinases
D053823:Hedgehog Proteins; D051741:Kruppel-Like Transcription Factors; D020930:MAP Kinase Kinase Kinases; D015513:Oncogene Proteins; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D015534:Trans-Activators; D000071676:Zinc Finger Protein GLI1; D000075746:Zinc Finger Protein Gli2; D000097603:Dyrk Kinases; C000618551:Gli2 protein, mouse