Description:
Aktisaserine/threonineproteinkinasethatplaysimportantrolesinmediatingsignalsfornumerousofcellfunctionsincludingcellgrowth,differentiation,cellcycle,transcription,andglucosemetabolism.TheenzymeisactivatedbyPI3K,andactivatedAktleadstophosphorylationofawiderangeofproteinsubstratessuchasGSK-3,IKK-ß,BAD,MDM2,mTOR,CASP9,p21Cip1,p27Kip1.DifferentialexpressionofAktsubstratescontributesdifferentphenotypicoutcomes.ApplicableGrid:
ListofApplicableGenes
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | |
| A | 14-3-3sigma | CCND1 | MDM2 | 14-3-3sigma | CCND1 | MDM2 | 14-3-3sigma | CCND1 | MDM2 | 14-3-3sigma | CCND1 | MDM2 |
| B | 4E-BP | eNOS | mTOR | 4E-BP | eNOS | mTOR | 4E-BP | eNOS | mTOR | 4E-BP | eNOS | mTOR |
| C | Akt1 | FOXO1 | p21 | Akt1 | FOXO1 | p21 | Akt1 | FOXO1 | p21 | Akt1 | FOXO1 | p21 |
| D | Akt2 | FOXO3 | p27,kip1 | Akt2 | FOXO3 | p27,kip1 | Akt2 | FOXO3 | p27,kip1 | Akt2 | FOXO3 | p27,kip1 |
| E | Akt3 | GAPDH | p70-S6K | Akt3 | GAPDH | p70-S6K | Akt3 | GAPDH | p70-S6K | Akt3 | GAPDH | p70-S6K |
| F | B-actin | GSK-3A | PDK1 | B-actin | GSK-3A | PDK1 | B-actin | GSK-3A | PDK1 | B-actin | GSK-3A | PDK1 |
| G | Bad | IGF1R | PI3K | Bad | IGF1R | PI3K | Bad | IGF1R | PI3K | Bad | IGF1R | PI3K |
| H | CASP9 | IKK | PTEN | CASP9 | IKK | PTEN | CASP9 | IKK | PTEN | CASP9 | IKK | PTEN |
Principle:
Signosis’proprietaryCDNAplatearrayisaplate-basedhybridizationprofilinganalysisformonitoringtheexpressionofdozensofgenesthroughreversetranscriptionofmRNAintocDNA.Likearrayanalyses,totalRNAisfirstreversetranscribedintocDNAinthepresenceofbiotin-dUTPintheassay.Targetedgenesarethenspecificallycapturedontoindividualwellsonaplate,insteadofmembranes,throughapre-coatedgene-specificoligonucleotide.ThecapturedcDNAsarefurtherdetectedwithstreptavidin-HRP.Luminescenceisreportedasrelativelightunits(RLUs)onamicroplateluminometer.Theexpressionlevelofgenesisdirectlyproportionaltotheluminescentintensity. |
Data:
HumanAktpathwaycDNAplatearrayanalysis. HeLacellsweretreatedwithandwithout100ng/mlIGFfor4hours,RNAsprepared,cDNAsynthesizedwithbiotinlabelandsubjectedtocDNAplatearrayhybridizationanddetection.
