1-NM-PP1 - CAS 221244-14-0
Not Intended for Therapeutic Use. For research use only.
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CAS 221244-14-0 1-NM-PP1

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Reference Reading

1.Analysis of 3-phosphoinositide-dependent kinase-1 signaling and function in ES cells.
Tamgüney T1, Zhang C, Fiedler D, Shokat K, Stokoe D. Exp Cell Res. 2008 Jul 1;314(11-12):2299-312. doi: 10.1016/j.yexcr.2008.04.006. Epub 2008 Apr 23.
3-phosphoinositide-dependent kinase-1 (PDK1) phosphorylates and activates several kinases in the cAMP-dependent, cGMP-dependent and protein kinase C (AGC) family. Many putative PDK1 substrates have been identified, but have not been analyzed following transient and specific inhibition of PDK1 activity. Here, we demonstrate that a previously characterized PDK1 inhibitor, BX-795, shows biological effects that are not consistent with PDK1 inhibition. Therefore, we describe the creation and characterization of a PDK1 mutant, L159G, which can bind inhibitor analogues containing bulky groups that hinder access to the ATP binding pocket of wild type (WT) kinases. When expressed in PDK1(-/-) ES cells, PDK1 L159G restored phosphorylation of PDK1 targets known to be hypophosphorylated in these cells. Screening of multiple inhibitor analogues showed that 1-NM-PP1 and 3,4-DMB-PP1 optimally inhibited the phosphorylation of PDK1 targets in PDK1(-/-) ES cells expressing PDK1 L159G but not WT PDK1.
2.Synchronized fission yeast meiosis using an ATP analog-sensitive Pat1 protein kinase.
Cipak L1, Polakova S2, Hyppa RW3, Smith GR4, Gregan J5. Nat Protoc. 2014 Jan;9(1):223-31. doi: 10.1038/nprot.2014.013. Epub 2014 Jan 2.
Synchronous cultures are often indispensable for studying meiosis. Here we present an optimized protocol for induction of synchronous meiosis in the fission yeast Schizosaccharomyces pombe. Chemical inactivation of an ATP analog-sensitive form of the Pat1 kinase (pat1-as2) by adding the ATP analog 1-NM-PP1 in G1-arrested cells allows the induction of synchronous meiosis at optimal temperature (25°C). Importantly, this protocol eliminates detrimental effects of elevated temperature (34°C), which is required to inactivate the commonly used temperature-sensitive Pat1 kinase mutant (pat1-114). The addition of the mat-Pc gene to a mat1-M strain further improves chromosome segregation and spore viability. Thus, our protocol offers highly synchronous meiosis at optimal temperature, with most characteristics similar to those of wild-type meiosis. The synchronization protocol can be completed in 5 d (not including strain production, which may take as long as 2 or 3 months).
3.Mps1 kinase activity restrains anaphase during an unperturbed mitosis and targets Mad2 to kinetochores.
Tighe A1, Staples O, Taylor S. J Cell Biol. 2008 Jun 16;181(6):893-901. doi: 10.1083/jcb.200712028. Epub 2008 Jun 9.
Mps1 is an upstream component of the spindle assembly checkpoint, which, in human cells, is required for checkpoint activation in response to spindle damage but not apparently during an unperturbed mitosis. Mps1 also recruits Mad1 and Mad2 to kinetochores. However, whether the enzymatic activity of Mps1 is required for these processes is unclear. To address this question, we established an RNA interference (RNAi) complementation assay. Repression of Mps1 triggers premature anaphase, often with unaligned or maloriented chromosomes. This phenotype is rescued by an RNAi-resistant wild-type Mps1 transgene but not by a catalytically inactive mutant. An analogue-sensitive allele, Mps1(M602A), also rescues the RNAi-induced defect, but not when inhibited by the adenosine triphosphate analogue 1-NM-PP1. Thus, Mps1 activity does restrain anaphase during an unperturbed mitosis. Furthermore, although catalytically inactive Mps1 can restore kinetochore localization of Mad1, only the active kinase restores Mad2 localization.