The PITSLRE protein kinases, hereafter referred to as CDK11 because of their association with the cyclin L regulatory partner, belong to large molecular weight protein complexes that contain RNA polymerase II. These CDK11 p110 complexes have been reported to influence transcription as well as interact with the general pre-mRNA-splicing factor RNPS1. Some of these complexes may also play a role in pre-mRNA splicing. Using a two-hybrid interactive screen, the splicing protein 9G8 was identified as an in vivo partner for CDK11 p110 . The identification of several splicing-related factors as CDK11 p110 interactors along with the close relationship between transcription and splicing indicated that CDK11 p110 might influence splicing activity directly. Immunodepletion of CDK11 p110 from splicing extracts greatly reduced the appearance of spliced products using an in vitro assay system. Moreover, the re-addition of these CDK11 p110 immune complexes to the CDK11 p110 -immunodepleted splicing reactions completely restored splicing activity. Similarly, the addition of purified CDK11 p110 amino-terminal domain protein was sufficient to inhibit the splicing reaction. Finally, 9G8 is a phosphoprotein in vivo and is a substrate for CDK11 p110 phosphorylation in vitro. These data are among the first demonstrations showing that a CDK activity is functionally coupled to the regulation of pre-mRNA-splicing events and further support the hypothesis that CDK11 p110 is in a signaling pathway that may help to coordinate transcription and RNA-processing events.The regulation of transcription and RNA-processing events is complex and, in fact, may be coordinated through the action of several protein kinases that belong to the larger cell division control family (i.e. CDKs) 1 (1-7). These CDKs are not only regulated during the cell cycle, they are undoubtedly part of a much larger signal transduction pathway that is modulated by numerous extracellular and intracellular signals (8). A model is emerging in which the phosphorylation status of the RNA polymerase II (RNAP II) carboxyl-terminal domain (CTD) determines which specific transcription, RNA processing, and polyadenylation factors are recruited to the CTD (5, 7-9). The CTD is heavily phosphorylated in vivo, and a number of protein kinases, particularly CDKs, have been identified that modify this domain in vitro. CDK7, CDK8, and CDK9 are all known CTD protein kinases that regulate different aspects of transcription (10 -13). Sequential phosphorylation of specific amino acid residues within the CTD appears to be regulated by this group of CDKs. These phosphorylation events have been linked to the changes in the composition of RNAP II complexes that are associated with transcription from pre-initiation through termination. However, a functional relationship between a specific CDK and the regulation of pre-mRNA-splicing events has only been inferred (14,15).Based upon the findings reported in this study as well as others (14, 15) demonstrating that the PITSLRE p110 protein kinases associ...

Our data indicate that the presence of the CYP2C19*17 allele results in ultra-rapid metabolism of voriconazole after a single oral dose.

Although the PITSLRE protein kinases are members of the cyclin-dependent kinase superfamily, their cellular function is unclear. Previously we demonstrated that the general RNA splicing factor RNPS1 is a specific PITSLRE p110 kinase interactor in vivo. This suggests that the PITSLRE family of protein kinases is involved in some aspect of RNA processing or transcription. Here we identify multiple transcriptional elongation factors, including ELL2, TFIIF 1 , TFIIS, and FACT, as PITSLRE kinase-associated proteins. We demonstrate that PITSLRE p110 protein kinases co-immunoprecipitate and/or co-purify with these elongation factors as well as with RNA polymerase II. Antibody-mediated inhibition of PITSLRE kinase specifically suppressed RNA polymerase II-dependent in vitro transcription initiated at a GC-rich (adenosine deaminase) or TATA box-dependent (Ad2ML) promoter, and this suppression was rescued by readdition of purified PITSLRE p110 kinase. Together, these data strongly suggest that PITSLRE protein kinases participate in a signaling pathway that potentially regulates or links transcription and RNA processing events.Regulation of transcription and RNA processing occurs on many levels, and these two dynamic processes are physically linked within the cell nucleus. It is hypothesized that, in part, regulation of transcription occurs through the active exchange of associated factors with the RNAP II 1 complex during these processes, resulting in transcriptional stimulation or repression (1-3). This hypothesis is based on the identification of numerous positive and negative regulatory transcription factors/complexes, as well as RNA processing enzymes, in association with RNAP II. Transcriptional elongation is facilitated by cellular factors that include FACT and elongator (2, 4). Complexes that repress transcription at some stage in this process include NAT (negative regulator of active transcription), sin3, and NELF (negative elongation factor) (5-7). Many of these complexes exert their effects through direct or indirect association with the RNAP II carboxyl-terminal domain (CTD). The mammalian RNAP II CTD is composed of 52 heptapeptide repeats with the consensus sequence Tyr-Ser-Pro-Thr-Ser-ProSer (YSPTSPS) that are essential for viability (8). The CTD is heavily phosphorylated in vivo, and many protein kinases have been identified that modify this domain. In addition to regulation of transcription initiation and elongation, CTD phosphorylation has been linked to RNA processing events that result in capped, spliced, and tailed mRNA (3, 9 -14). Clearly, phosphorylation plays an important role in regulating the production of mRNA transcripts by affecting the nature of the RNAP II enzyme complex.Several cdks phosphorylate the CTD, and some of these cdks co-purify with RNAP II complexes. These kinases include cdk1, cdk7, cdk8, and cdk9. It is likely that sequential phosphorylation events, as well as phosphorylation of specific residues by specific protein kinases, help regulate transcription from preinitiation through ...

The PITSLRE protein kinases, hereafter referred to as cyclin-dependent kinase 11 (CDK11) due to their association with cyclin L, are part of large molecular weight protein complexes that contain RNA polymerase II (RNAP II) as well as numerous transcription and RNA processing factors. Data presented here demonstrate that the influence of CDK11 p110 on transcription and splicing does not involve phosphorylation of the RNAP II carboxyl-terminal domain by CDK11 p110 . We have isolated a DRB-and heparin-sensitive protein kinase activity that co-purifies with CDK11 p110 after ion exchange and affinity purification chromatography. This protein kinase was identified as casein kinase 2 (CK2) by immunoblot and mass spectrometry analyses. In addition to the RNAP II carboxyl-terminal domain, CK2 phosphorylates the CDK11 p110 amino-terminal domain. These data suggest that CDK11 p110 isoforms participate in signaling pathways that include CK2 and that its function may help to coordinate the regulation of RNA transcription and processing events. Future experiments will determine how phosphorylation of CDK11 p110 by CK2 specifically affects RNA transcription and/or processing events.The complex biochemical events of transcription and RNA processing, resulting in the production of mature RNA transcripts, are now understood to be highly integrated and co-dependent processes (1). It is hypothesized that regulation of these events occurs through the active exchange of associated factors with the RNAP II 1 complex (2-4). This hypothesis is based upon identification of numerous positive and negative regulatory factors/complexes, influencing both transcription and RNA processing enzymes, in physical association with RNAP II. Many of these complexes exert their effects directly or indirectly through association with the RNAP II CTD. In mammals, the RNAP II CTD is composed of 52 heptapeptide repeats with the consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser (YSPTSPS), which are essential for viability (5).The RNAP II CTD is heavily phosphorylated in vivo, and it is likely that sequential phosphorylation events, as well as phosphorylation of specific residues by specific protein kinases, help regulate transcript production. This model appears to fit much of the data coming from numerous laboratories and was recently proposed as the most likely means of coordinating the various steps of transcription, RNA processing, and mRNA export (6). Many protein kinases modify the RNAP II CTD. Several of the CTD kinases identified thus far are from the cyclin-dependent kinase family (CDKs) and include CDK1, CDK7, CDK8, and CDK9. In addition, another regulator of cell cycle events, casein kinase 2 (CK2), is known to phosphorylate a number of transcriptional proteins, including the RNAP II CTD and the RAP74 subunit of TFIIF (7, 8).Data from this laboratory and others demonstrate that the CDK11 p110 (PITSLRE) protein kinases associate with the cyclin L regulatory protein, bind directly to various splicing factors, and play a role in pre-mRNA splicing (9 -1...

Our findings provide insight into the epigenetic regulation of Myc via histone demethylation and proof-of-concept for inhibition of histone demethylases to target Myc signaling in cancers such as neuroblastoma.

Summary CDK11p58 is required for the maintenance of sister chromatid cohesion

No abstract

Histone lysine demethylases facilitate the activity of oncogenic transcription factors including possibly MYC. Here we show that multiple histone demethylases influence the viability and poor prognosis of neuroblastoma cells where MYC is often overexpressed. We also identified the approved small molecule antifungal agent ciclopirox as a novel pan-histone demethylase inhibitor. Ciclopirox targeted several histone demethylases including KDM4B implicated in MYC function. Accordingly, ciclopirox inhibited Myc signaling in parallel with mitochondrial oxidative phosphorylation, resulting in suppression of neuroblastoma cell viability and inhibition of tumor growth associated with an induction of differentiation. Our findings provide new insights into epigenetic regulation of MYC function and suggest a novel pharmacologic basis to target histone demethylases as an indirect MYC targeting approach for cancer therapy.