Enzymes, Signaling Proteins, Antibodies

Errors show statistical significance for the inhibition of transport following PCTAIRE-1 expression. A specific interaction was seen between Sec23Ap and the region of PCTAIRE-1 corresponding to that of PCTAIRE-3 identified in the initial screen (exons 4-8). Again, an interaction with Sec24Dp was also detectable (Fig. 1C). 1D in which the PSTAIRE kinase CDK2 is shown not to interact with any of the COPII components.
The structure of this complex was solved using CDK5 as a phasing model and refined at a resolution of 2.4 Å with one CDK16 monomer in the asymmetric unit. The electron density maps for the structure were generally of high quality. The CDK16 chain was modelled between residues Met162–Glu473, with the exception of regions of poor electron density at the αC helix, PCTAIRE Antibodies Glu199–Ala204, as well as the activation segment Ser312–Val323. The C-terminus between Ala474–His485, including the affinity tag, could also not be built. There was clear density for the indirubin E804 inhibitor in the ATP-binding pocket. The figure shows a schematic of the mechanism by which APP or Aβ may be inducing neuropathology development in AD.

In vivo lung metastasis was continuously monitored for another week after rebastinib withdrawal. To test the effect of rebastinib on tumor growth of TNBC, mice bearing MDA-MB-231 tumors or PDX tumors were randomly grouped when the average tumor size reached 50 mm3. Rebastinib (80 mg/kg) formulated in 0.5% hydroxypropyl methyl  cellulose or vehicle were orally administrated once a day for over 10 days. The guaranteed shelf life from date of receipt for bioscience kits is listed on the product information sheet. Some kits have an expiration date printed on the kit box label, this is the guaranteed shelf life date calculated from the day that the product shipped from our facility.
The newly proposed atypical CDK members PFTAIRE1–2 (CDK14–15) and PCTAIRE1–3 (CDK16–18) kinases share CCNY as an activator and are co-expressed in various tissues with the highest expression in brain and testis . The functions and regulatory networks of these atypical CDKs are largely unknown. Knocking down individual atypical CDKs failed to cause any phenotype, suggesting possible functional redundancy of these kinases . However, a functional study of CDK16 using CDK16 conditional knockout mouse model has validated its essential role in spermatogenesis . Here, our study demonstrated that CDK16 phosphorylates PRC1 and Rb, thereby regulating spindle formation and the Rb-E2F pathway, providing new insights into the function of the atypical CDK subfamily.

At Proteintech, we pride ourselves on our antibody quality, customer service and transparency. As such, we are comparing our antibodies with other vendors, enabling easy identification and comparisons of key data to help you choose the suitable antibody for your needs. Check out our latest Cell Science at a Glance article and accompanying poster for an overview of the mechanism of biogenesis and biological functions of ciliary extracellular vesicles. We interviewed Guillaume Jacquemet, who established his own lab in 2019 at Åbo Akademi University, Turku.
The kinase was found to be homogeneously distributed throughout the cell (Fig. 7). In some instances, PCTAIRE-1 staining was apparently more intense in the perinuclear region (Fig. 6). Time course expression experiments demonstrated that this was not a function of protein accumulation in the cell . The subcellular distribution of PCTAIRE-1 prompted us to examine its possible colocalization with the cytoskeletal network.

The amount of protein transported to the plasma membrane was then measured 60 minutes after temperature shift using immunofluorescence with an antibody directed against the extracellular domain of ts045-G-YFP. The proportion of ts045-G-YFP transported to the plasma membrane in cells expressing GFP, GFP-PCTAIRE-1 or GFP-PCTAIRE-1 was determined (Fig. 6). An 18% reduction in transport was seen in cells coexpressing the kinase-dead mutant. In contrast, cells expressing an active S153A mutant of PCTAIRE-1 showed an equivalent enhanced amount of transport of ts045-G-YFP.
The Proteintech guarantee covers Proteintech antibodies in any species and any application, including those not listed on the datasheet. If the antibody doesn’t perform, you can receive a hassle-free refund or credit note. We then examined the effect of RNAi-mediated depletion of PCTAIRE-1 on the secretory pathway. Two different siRNA duplexes were designed to target the PCTAIRE-1 coding sequence. These were shown to be effective in depletion of the PCTAIRE-1 protein level by immunoblotting (Fig. 4A). Lamin was used as a control for these experiments (Fig. 4B).
The third member of this family, PCTK3, has been the least studied. Although exogenously expressed PCTK1 and PCTK2 phosphorylate myelin basic protein and histone H1 in vitro, PCTK3 kinase activity has not been detected. To characterize PCTK3 activity and elucidate its regulatory mechanism, it is necessary to identify activators for PCTK3. PCTAIRE protein kinases interact directly with the COPII complex and modulate secretory cargo transport.

Initial phases were calculated in PHASER using the first CDK16 structure as a model for molecular replacement . After obtaining the initial electron density maps by rigid body refinement in REFMAC5 , a structural model was generated by automated building in Buccaneer . Several rounds of non-crystallographic symmetry restrained refinement were then performed in REFMAC5 combined with manual modelling with COOT .
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Transcriptomic analysis was performed to reveal the potential molecular mechanisms involved in the function of CDK16. Pharmacological inhibition of CDK16 was achieved by the small molecular inhibitor rebastinib to further assess the anti-tumor utility of targeting CDK16. Consistent with other work (Graeser et al., 2002), expression was detectable in a number of transformed cell lines. PCTAIRE-1 was expressed in human cervical epithelial carcinoma cells and human embryonic kidney 293 (HEK-293) cells with lower levels in Madin-Darby canine kidney cells and trace levels of expression in A549 lung epithelial cells (Fig. 3A).
An in vitro kinase assay using retinoblastoma protein as the substrate showed that PCTK3 was specifically activated by cyclin A2 but not by cyclin E1, although its activity was lower than that of CDK2. Furthermore, immunocytochemistry analysis showed that PCTK3 colocalized with cyclin A2 in the cytoplasm and regulated cyclin A2 stability. Amino acid sequence analysis revealed that PCTK3 contained four putative PKA phosphorylation sites. In vitro and in vivo kinase assays showed that PCTK3 was phosphorylated by PKA at Ser12, Ser66, and Ser109 and that PCTK3 activity significantly increased via phosphorylation at Ser12 by PKA even in the absence of cyclin A2.
Data acquisition was set to stop after 9 min or after a minimum of 10,000 events had been collected in the single events region. List mode files contained all events detected above a threshold of DNA content ≥ 10% of the G0/G1 content. The G0/G1 peak for an untreated cycling control was set at channel 235, which yielded at least 200 channels for S-phase resolution. MBP phosphorylation was analyzed by SDS-PAGE on a 15% Laemmli gel. Labeled bands were visualized by autoradiography or with a PhosphorImager . For in situ detection of kinase activity, 0.5 mg/ml MBP was added to the gel just before polymerization, which was performed as described previously , and kinase activity was measured as described previously .