Review

cdk16 human (Addgene inc)

Addgene inc is a verified supplier

About

News

Press Release

Team

Advisors

Partners

Contact

Bioz Stars

Bioz vStars

Buy from Supplier

Structured Review

Addgene inc cdk16 human
Cdk16 Human, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/cdk16+human/pmc06081246-16-0-4?v=Addgene+inc
Average 86 stars, based on 1 article reviews

cdk16 human - by Bioz Stars, 2026-07

86/100 stars

Images

Image Search Results

CDK16 is frequently overexpressed in lung cancer and elevated level of CDK16 predicts poor prognosis. A. Western blotting analysis of CDK16 expression in six human lung cancer cell lines and normal lung epithelial BEAS cells. The ratio shows relative CDK16 protein expression normalized for GAPDH (BEAS, set at 1). B. Representative immunohistochemical staining images of CDK16 expression (high or low) in tissue microarrays constructed from lung cancers and paired adjacent lung tissues. Scale bar, 100 μm. C. A summary of immunohistochemical staining results of CDK16 expression in tissue microarrays. D. Kaplan-Meier analysis of overall survival of lung cancer patients stratified by CDK16 expression.

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 is frequently overexpressed in lung cancer and elevated level of CDK16 predicts poor prognosis. A. Western blotting analysis of CDK16 expression in six human lung cancer cell lines and normal lung epithelial BEAS cells. The ratio shows relative CDK16 protein expression normalized for GAPDH (BEAS, set at 1). B. Representative immunohistochemical staining images of CDK16 expression (high or low) in tissue microarrays constructed from lung cancers and paired adjacent lung tissues. Scale bar, 100 μm. C. A summary of immunohistochemical staining results of CDK16 expression in tissue microarrays. D. Kaplan-Meier analysis of overall survival of lung cancer patients stratified by CDK16 expression.

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Western Blot, Expressing, Immunohistochemical staining, Staining, Construct

CDK16 knockdown leads to p53 accumulation. A. A549 cells were transfected with indicated siRNAs for 48 h, and samples were collected and analyzed by Western blotting with the indicated antibodies. The ratio shows relative p53 protein expression normalized for GAPDH (scramble, set at 1). B . A549 cells infected with lenti-CRISPR sgRNAs targeting CDK16 or control were incubated with puromycin for one week, and endogenous CDK16 and p53 were analyzed by western blotting. C. A549 and H292 cells were transfected with constructs encoding SFB-CDK16. Cells were collected 24 h later and cell lysates were analyzed by Western blotting using indicated antibodies. D. H292, U2OS, and MCF-7 cells were transfected with indicated siRNAs for 48 h, and analyzed as described in (A). E. A549 and H292 cells were transfected with scrambled or CDK16 siRNAs for 48 h. The mRNA levels for the indicated genes were determined by Real-time quantitative PCR (n=3). F. HCC827 and H1975 cells were transfected with scrambled or CDK16 siRNAs as indicated for 48 h and analyzed as described in (A).

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 knockdown leads to p53 accumulation. A. A549 cells were transfected with indicated siRNAs for 48 h, and samples were collected and analyzed by Western blotting with the indicated antibodies. The ratio shows relative p53 protein expression normalized for GAPDH (scramble, set at 1). B . A549 cells infected with lenti-CRISPR sgRNAs targeting CDK16 or control were incubated with puromycin for one week, and endogenous CDK16 and p53 were analyzed by western blotting. C. A549 and H292 cells were transfected with constructs encoding SFB-CDK16. Cells were collected 24 h later and cell lysates were analyzed by Western blotting using indicated antibodies. D. H292, U2OS, and MCF-7 cells were transfected with indicated siRNAs for 48 h, and analyzed as described in (A). E. A549 and H292 cells were transfected with scrambled or CDK16 siRNAs for 48 h. The mRNA levels for the indicated genes were determined by Real-time quantitative PCR (n=3). F. HCC827 and H1975 cells were transfected with scrambled or CDK16 siRNAs as indicated for 48 h and analyzed as described in (A).

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Transfection, Western Blot, Expressing, Infection, CRISPR, Incubation, Construct, Real-time Polymerase Chain Reaction

CDK16 directly interacts with p53 in vivo and in vitro . A. HEK293T cells transfected with the indicated constructs were collected 24 h later. Cells were lysed with NETN buffer. Immunoprecipitation (IP) using S-protein agarose were performed and Western blotted with the indicated antibodies. WCL: whole cell lysate. B. Endogenous CDK16 associated with p53 in A549 cells. A549 cells were lysed and subjected to immunoprecipitation using anti-IgG or anti-p53 as indicated, and were analyzed by Western blotting using indicated antibodies. C. Beads coated with GST or GST-p53 fusion proteins were incubated with SFB-CDK16 protein overnight. GST pulldown was immunoblotted with indicated antibodies. D. Schematic description of p53 domains and deletion mutants used in this study. E. HEK293T cells co-transfected with constructs encoding SFB-CDK16 and indicated p53 mutants were collected 24 h later. Cells were lysed, the supernatants were incubated with S-protein agarose beads, and then analyzed by Western blotting using antibodies as indicated.

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 directly interacts with p53 in vivo and in vitro . A. HEK293T cells transfected with the indicated constructs were collected 24 h later. Cells were lysed with NETN buffer. Immunoprecipitation (IP) using S-protein agarose were performed and Western blotted with the indicated antibodies. WCL: whole cell lysate. B. Endogenous CDK16 associated with p53 in A549 cells. A549 cells were lysed and subjected to immunoprecipitation using anti-IgG or anti-p53 as indicated, and were analyzed by Western blotting using indicated antibodies. C. Beads coated with GST or GST-p53 fusion proteins were incubated with SFB-CDK16 protein overnight. GST pulldown was immunoblotted with indicated antibodies. D. Schematic description of p53 domains and deletion mutants used in this study. E. HEK293T cells co-transfected with constructs encoding SFB-CDK16 and indicated p53 mutants were collected 24 h later. Cells were lysed, the supernatants were incubated with S-protein agarose beads, and then analyzed by Western blotting using antibodies as indicated.

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: In Vivo, In Vitro, Transfection, Construct, Immunoprecipitation, Western Blot, Incubation

$CDK16 phosphorylates p53 at Ser315 site and inhibits p53 transcriptional activity. A. A549 cells were transfected with the indicated siRNA for 48 h, then incubated with 10 μM MG132 for 4 h prior to collection. Cell lysates were analyzed by Western blotting using indicated antibodies. The ratio shows relative phosphorylated p53 protein expression normalized for GAPDH (scramble, set at 1). B. Alignment of CDK16 candidate phosphorylation site in p53 from different species. C. CDK16 phosphorylates p53 at Ser315 site in vitro . HA-p53-WT or S315A proteins were incubated in vitro with immunoprecipitates isolated from HEK293T cells transfected with constructs encoding SFB-CDK16 and then analyzed by Western blotting using indicated antibodies. D. CDK16 depletion increases p53 abundance in the nucleus. A549 cells were fractionated into cytoplasmic and nuclear fractions, and the indicated proteins in each compartment were analyzed by Western blotting. α-tubulin and LaminB1 were used respectively as cytoplasmic and nuclear loading controls. E. The mRNA levels of the indicated genes were analyzed by Real time quantitative PCR in A549 cells transfected with scramble or CDK16 siRNAs. *** P < 0.001 (n=3).$

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 phosphorylates p53 at Ser315 site and inhibits p53 transcriptional activity. A. A549 cells were transfected with the indicated siRNA for 48 h, then incubated with 10 μM MG132 for 4 h prior to collection. Cell lysates were analyzed by Western blotting using indicated antibodies. The ratio shows relative phosphorylated p53 protein expression normalized for GAPDH (scramble, set at 1). B. Alignment of CDK16 candidate phosphorylation site in p53 from different species. C. CDK16 phosphorylates p53 at Ser315 site in vitro . HA-p53-WT or S315A proteins were incubated in vitro with immunoprecipitates isolated from HEK293T cells transfected with constructs encoding SFB-CDK16 and then analyzed by Western blotting using indicated antibodies. D. CDK16 depletion increases p53 abundance in the nucleus. A549 cells were fractionated into cytoplasmic and nuclear fractions, and the indicated proteins in each compartment were analyzed by Western blotting. α-tubulin and LaminB1 were used respectively as cytoplasmic and nuclear loading controls. E. The mRNA levels of the indicated genes were analyzed by Real time quantitative PCR in A549 cells transfected with scramble or CDK16 siRNAs. *** P < 0.001 (n=3).

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Activity Assay, Transfection, Incubation, Western Blot, Expressing, In Vitro, Isolation, Construct, Real-time Polymerase Chain Reaction

CDK16 regulates p53 stability via the ubiquitin/proteasome pathway. A. A549 cells were transfected with the indicated siRNA for 48 h, then incubated with 10 μM MG132 for 4 h prior to harvesting. Cell lysates were analyzed by Western blotting using indicated antibodies. The ratio shows relative p53 protein expression normalized for GAPDH (scramble, set at 1). B. Left panel: A549 cells transfected with indicated siRNA for 48 h were treated with CHX (20 mg/mL) for the indicated times and then analyzed by Western blotting. Right panel: quantification of p53 band intensities is shown. C. A549 cells were transfected with indicated constructs after cells were transfected with CDK16 siRNA for 24 h. Cells were harvested after treatment with MG132 (10 μM) for 4 h. The samples were subjected to immunoprecipitation using S-protein agarose beads followed by Western blotting analysis as indicated. D and E. A549 cells were transfected with indicated constructs for 24 h and collected after treatment with MG132 (10 μM) for 4 h. The samples were subjected to immunoprecipitation using S-protein agarose beads and analyzed by Western blotting using indicated antibodies.

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 regulates p53 stability via the ubiquitin/proteasome pathway. A. A549 cells were transfected with the indicated siRNA for 48 h, then incubated with 10 μM MG132 for 4 h prior to harvesting. Cell lysates were analyzed by Western blotting using indicated antibodies. The ratio shows relative p53 protein expression normalized for GAPDH (scramble, set at 1). B. Left panel: A549 cells transfected with indicated siRNA for 48 h were treated with CHX (20 mg/mL) for the indicated times and then analyzed by Western blotting. Right panel: quantification of p53 band intensities is shown. C. A549 cells were transfected with indicated constructs after cells were transfected with CDK16 siRNA for 24 h. Cells were harvested after treatment with MG132 (10 μM) for 4 h. The samples were subjected to immunoprecipitation using S-protein agarose beads followed by Western blotting analysis as indicated. D and E. A549 cells were transfected with indicated constructs for 24 h and collected after treatment with MG132 (10 μM) for 4 h. The samples were subjected to immunoprecipitation using S-protein agarose beads and analyzed by Western blotting using indicated antibodies.

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Transfection, Incubation, Western Blot, Expressing, Construct, Immunoprecipitation

CDK16 silencing results in enhanced radiosensitivity of lung cancer cells. A. Colony formation ability was significantly reduced in CDK16-depleted A549 cells. *** P < 0.001 compared with controls cells (n=3). B. Apoptosis was remarkably increased in CDK16-depleted A549 cells. A549 cells were transfected with indicated siRNAs. 48 h later, cells were subjected to annexin V-EGFP/propidium iodide staining, and analyzed by flow cytometry (n=3). C. A549 cells were transfected with indicated siRNAs for 48 h, and cells were stained with propidium iodide (PI) and analyzed by flow cytometry (n=3). D. Knockdown of CDK16 promoted cellular ROS production. A549 cells were transfected with indicated siRNAs. 48 h later, cells were collected and ROS levels were measured by flow cytometry (n=3). E. Upper panel: A549 cells were irradiated with 2 Gy of ionizing radiation and harvested at indicated time points. Immunostaining staining was performed to determine γ-H2AX foci formation. Scale bar, 50 μm. Lower panel: quantification result of γ-H2AX foci in A549 cells is shown. * P < 0.05, ** P < 0.01 compared with controls cells (n=3). F. Radiation sensitivity of A549 cells lacking CDK16. A549 cells transfected with indicated siRNAs were irradiated with indicated doses of IR. The percentages of surviving colonies were evaluated two weeks later (n=3).

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: CDK16 silencing results in enhanced radiosensitivity of lung cancer cells. A. Colony formation ability was significantly reduced in CDK16-depleted A549 cells. *** P < 0.001 compared with controls cells (n=3). B. Apoptosis was remarkably increased in CDK16-depleted A549 cells. A549 cells were transfected with indicated siRNAs. 48 h later, cells were subjected to annexin V-EGFP/propidium iodide staining, and analyzed by flow cytometry (n=3). C. A549 cells were transfected with indicated siRNAs for 48 h, and cells were stained with propidium iodide (PI) and analyzed by flow cytometry (n=3). D. Knockdown of CDK16 promoted cellular ROS production. A549 cells were transfected with indicated siRNAs. 48 h later, cells were collected and ROS levels were measured by flow cytometry (n=3). E. Upper panel: A549 cells were irradiated with 2 Gy of ionizing radiation and harvested at indicated time points. Immunostaining staining was performed to determine γ-H2AX foci formation. Scale bar, 50 μm. Lower panel: quantification result of γ-H2AX foci in A549 cells is shown. * P < 0.05, ** P < 0.01 compared with controls cells (n=3). F. Radiation sensitivity of A549 cells lacking CDK16. A549 cells transfected with indicated siRNAs were irradiated with indicated doses of IR. The percentages of surviving colonies were evaluated two weeks later (n=3).

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Transfection, Staining, Flow Cytometry, Irradiation, Immunostaining

p53 mediates the biological effects of CDK16 depletion in lung cancer cells. A. A549 cells were transfected with indicated siRNAs for 48 h. Cell lysates were analyzed by immunoblotting as indicated. B. A549 cells transfected with indicated siRNAs were seeded and cultured for two weeks. The colonies were stained and then counted. Representative pictures are shown. *** P < 0.001 (n=3). C. A549 cells were transfected with indicated siRNAs. 48 h later, cells were subjected to annexin V-EGFP/propidium iodide staining and analyzed by flow cytometry. *** P < 0.001 (n=3). D. A549 cells were transfected with indicated siRNAs and ROS levels were measured by flow cytometry. * P < 0.05, ** P < 0.01 (n=3). E. Left panel: A549 cells were irradiated with 2 Gy of ionizing radiation and harvested at 4 h post IR. Immunostaining was performed to determine γ-H2AX foci formation. Scale bar, 50 μm. Right panel: quantification result of γ-H2AX foci in A549 cells is shown. * P < 0.05, ** P < 0.01 (n=3). F. A549 cells transfected with indicated siRNAs were irradiated with indicated doses. After two weeks, colonies containing more than 50 cells were counted. (n=3).

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: p53 mediates the biological effects of CDK16 depletion in lung cancer cells. A. A549 cells were transfected with indicated siRNAs for 48 h. Cell lysates were analyzed by immunoblotting as indicated. B. A549 cells transfected with indicated siRNAs were seeded and cultured for two weeks. The colonies were stained and then counted. Representative pictures are shown. *** P < 0.001 (n=3). C. A549 cells were transfected with indicated siRNAs. 48 h later, cells were subjected to annexin V-EGFP/propidium iodide staining and analyzed by flow cytometry. *** P < 0.001 (n=3). D. A549 cells were transfected with indicated siRNAs and ROS levels were measured by flow cytometry. * P < 0.05, ** P < 0.01 (n=3). E. Left panel: A549 cells were irradiated with 2 Gy of ionizing radiation and harvested at 4 h post IR. Immunostaining was performed to determine γ-H2AX foci formation. Scale bar, 50 μm. Right panel: quantification result of γ-H2AX foci in A549 cells is shown. * P < 0.05, ** P < 0.01 (n=3). F. A549 cells transfected with indicated siRNAs were irradiated with indicated doses. After two weeks, colonies containing more than 50 cells were counted. (n=3).

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Transfection, Western Blot, Cell Culture, Staining, Flow Cytometry, Irradiation, Immunostaining

A proposed model shows mechanistically how CDK16 participates in radioresistance by phosphorylating and destabilizing p53. A. High level of CDK16 leads to its binding to p53, which subsequently phosphorylates p53 at Ser315 site and promotes p53 degradation via the ubiquitin/proteasome system. B. Loss of CDK16 or CDK16 inhibition prevents p53 phosphorylation, which leads to p53 stabilization and transcriptional activation of p53 that ultimately inhibits radioresistance in lung cancer.

Journal: Theranostics

Article Title: CDK16 Phosphorylates and Degrades p53 to Promote Radioresistance and Predicts Prognosis in Lung Cancer

doi: 10.7150/thno.21963

Figure Lengend Snippet: A proposed model shows mechanistically how CDK16 participates in radioresistance by phosphorylating and destabilizing p53. A. High level of CDK16 leads to its binding to p53, which subsequently phosphorylates p53 at Ser315 site and promotes p53 degradation via the ubiquitin/proteasome system. B. Loss of CDK16 or CDK16 inhibition prevents p53 phosphorylation, which leads to p53 stabilization and transcriptional activation of p53 that ultimately inhibits radioresistance in lung cancer.

Article Snippet: Anti-human CDK16 antibodies were purchased from Sigma and Proteintech.

Techniques: Binding Assay, Inhibition, Activation Assay

Expression of CDK16 in EC and adjacent normal tissues. (A) CDK16 expression was higher in tumour tissues than in adjacent normal tissues in TCGA cohort ( p < 0.001). (B) qRT-PCR revealed that CDK16 expression was higher in tumour tissues than in adjacent normal tissues. (C) Immunohistochemical analysis revealed that CDK16 expression was higher in tumour than in adjacent normal tissues (*, p < 0.005; ** p < 0.01; *** p < 0.001).

Journal: Frontiers in Genetics

Article Title: Endometrial cancer prognosis prediction using correlation models based on CDK family genes

doi: 10.3389/fgene.2022.1021600

Figure Lengend Snippet: Expression of CDK16 in EC and adjacent normal tissues. (A) CDK16 expression was higher in tumour tissues than in adjacent normal tissues in TCGA cohort ( p < 0.001). (B) qRT-PCR revealed that CDK16 expression was higher in tumour tissues than in adjacent normal tissues. (C) Immunohistochemical analysis revealed that CDK16 expression was higher in tumour than in adjacent normal tissues (*, p < 0.005; ** p < 0.01; *** p < 0.001).

Article Snippet: The rabbit anti-human CDK16 antibody was purchased from CUSABIO (CSB-PA017648ESR2HU) (100 μl, Wuhan, China).

Techniques: Expressing, Quantitative RT-PCR, Immunohistochemical staining

Correlation between CDK16 expression and the clinicopathological features of patients with EC: (A) age, (B) weight (C) pathological stage, (D) histological grade. (E) Heatmap demonstrating the correlation between CDK16 expression and the clinicopathological characteristics of patients with EC.

Journal: Frontiers in Genetics

Article Title: Endometrial cancer prognosis prediction using correlation models based on CDK family genes

doi: 10.3389/fgene.2022.1021600

Figure Lengend Snippet: Correlation between CDK16 expression and the clinicopathological features of patients with EC: (A) age, (B) weight (C) pathological stage, (D) histological grade. (E) Heatmap demonstrating the correlation between CDK16 expression and the clinicopathological characteristics of patients with EC.

Article Snippet: The rabbit anti-human CDK16 antibody was purchased from CUSABIO (CSB-PA017648ESR2HU) (100 μl, Wuhan, China).

Techniques: Expressing

a Schematic representation of the ProtoArray based screen with approximately 9000 human proteins using AMPK (see also Supplementary Data 1). b Details of two sub-arrays incubated with or without AMPK with marked substrates are shown. c GST-CDK16, Cyclin Y-His 6 and GST were incubated in the presence of [γ- 32 P]-ATP with AMPK. Phosphorylation was determined by autoradiography ( 32 P, top). Proteins were visualized by Coomassie blue staining (CB, bottom; n = 2). d HeLa cells were transfected with vectors expressing GFP-CDK16 and Cyclin Y-Flag and treated for 1 h with 0.5 mM AICAR/50 µM A769662 (A769) as indicated. Cyclin Y-Flag was immunoprecipitated with Flag antibodies (IP) and immunoblotted against CDK16 and Cyclin Y or used for in vitro kinase assays with myeloid basic protein (MBP) as substrate. Autoradiographs ( 32 P) and Coomassie blue staining (CB) of MBP are displayed. Whole cell lysates (WCL) were immunoblotted with the indicated antibodies ( n = 3). e Quantification of CDK16 co-immunoprecipitated with Cyclin Y. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01, and *** p < 0.001. All error bars indicate SD ( n = 3; Cyclin Y + AICAR/A769 vs. Cyclin Y/CDK16: t = 8.719, df = 4; Cyclin Y/CDK16 vs. Cyclin Y/CDK16 + AICAR/A769: t = 5.595, df = 4). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a Schematic representation of the ProtoArray based screen with approximately 9000 human proteins using AMPK (see also Supplementary Data 1). b Details of two sub-arrays incubated with or without AMPK with marked substrates are shown. c GST-CDK16, Cyclin Y-His 6 and GST were incubated in the presence of [γ- 32 P]-ATP with AMPK. Phosphorylation was determined by autoradiography ( 32 P, top). Proteins were visualized by Coomassie blue staining (CB, bottom; n = 2). d HeLa cells were transfected with vectors expressing GFP-CDK16 and Cyclin Y-Flag and treated for 1 h with 0.5 mM AICAR/50 µM A769662 (A769) as indicated. Cyclin Y-Flag was immunoprecipitated with Flag antibodies (IP) and immunoblotted against CDK16 and Cyclin Y or used for in vitro kinase assays with myeloid basic protein (MBP) as substrate. Autoradiographs ( 32 P) and Coomassie blue staining (CB) of MBP are displayed. Whole cell lysates (WCL) were immunoblotted with the indicated antibodies ( n = 3). e Quantification of CDK16 co-immunoprecipitated with Cyclin Y. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01, and *** p < 0.001. All error bars indicate SD ( n = 3; Cyclin Y + AICAR/A769 vs. Cyclin Y/CDK16: t = 8.719, df = 4; Cyclin Y/CDK16 vs. Cyclin Y/CDK16 + AICAR/A769: t = 5.595, df = 4). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Article Snippet: YenZym Antibodies generated the rabbit antibodies targeting human CDK16 phospho-S65 (residues 58-SARGPLS-pS-APEIVH-71), phospho-S153 (residues 147-RRLRRV-pS-LSEIGFG-160), and phospho-S155 (residues 149-LRRVSL-pS-EIGFGKL-161).

Techniques: Incubation, Phospho-proteomics, Autoradiography, Staining, Transfection, Expressing, Immunoprecipitation, In Vitro, Two Tailed Test, Standard Deviation

a NIH3T3 cells were transfected with siRNA against Cdk16, Cyclin Y or a control for 72 h and grown in EBSS for 2 h. Proteins were detected as indicated ( n = 3). b Representative confocal images of the NIH3T3 treated as in panel a . Endogenous LC3 was stained with the 4E12 antibody to monitor autophagy. Scale bar: 50 µm. c Quantification of the LC3 dots shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: *** p < 0.001. All error bars indicate SD ( n = 3; 100 cells counted for each replicate; siControl vs. siCdk16: t = 10.09, df = 4; siControl vs. siCyclin Y: t = 9.866, df = 4). d NIH3T3 cells were transfected with siControl or siCyclin Y, treated with 200 nM Bafilomycin A1 (Baf. A1) or 3-MA for 2 h, and grown in EBSS for additional 2 h as indicated. Lysates were immunoblotted with the indicated antibodies ( n = 2). e Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 200 nM Baf. A1 for 4 h prior to growth in EBSS for additional 2 h. Proteins were measured by immunoblotting ( n = 3). f Representative confocal images of the Cdk16 +/+ and Cdk16 −/− MEFs treated as in panel e . Endogenous LC3 was stained (antibody 4E12) to monitor autophagy. Scale bar: 50 µm. g Quantification of the LC3 dots shown in panel f . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: * p < 0.05 and *** p < 0.001. All error bars indicate SD ( n = 3; 100 cells counted for each replicate; EBSS: t = 10.850, df = 4; Baf. A1: t = 4.58, df = 4; EBSS/Baf. A1: t = 3.978, df = 4). h Human CDK16 was stably expressed in immortalized Cdk16 −/− MEFs ( + CDK16). Control cells were infected with an empty virus. Cells were treated with 200 nM Baf. A1 and grown in EBSS. Proteins were detected as indicated ( n = 2). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a NIH3T3 cells were transfected with siRNA against Cdk16, Cyclin Y or a control for 72 h and grown in EBSS for 2 h. Proteins were detected as indicated ( n = 3). b Representative confocal images of the NIH3T3 treated as in panel a . Endogenous LC3 was stained with the 4E12 antibody to monitor autophagy. Scale bar: 50 µm. c Quantification of the LC3 dots shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: *** p < 0.001. All error bars indicate SD ( n = 3; 100 cells counted for each replicate; siControl vs. siCdk16: t = 10.09, df = 4; siControl vs. siCyclin Y: t = 9.866, df = 4). d NIH3T3 cells were transfected with siControl or siCyclin Y, treated with 200 nM Bafilomycin A1 (Baf. A1) or 3-MA for 2 h, and grown in EBSS for additional 2 h as indicated. Lysates were immunoblotted with the indicated antibodies ( n = 2). e Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 200 nM Baf. A1 for 4 h prior to growth in EBSS for additional 2 h. Proteins were measured by immunoblotting ( n = 3). f Representative confocal images of the Cdk16 +/+ and Cdk16 −/− MEFs treated as in panel e . Endogenous LC3 was stained (antibody 4E12) to monitor autophagy. Scale bar: 50 µm. g Quantification of the LC3 dots shown in panel f . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: * p < 0.05 and *** p < 0.001. All error bars indicate SD ( n = 3; 100 cells counted for each replicate; EBSS: t = 10.850, df = 4; Baf. A1: t = 4.58, df = 4; EBSS/Baf. A1: t = 3.978, df = 4). h Human CDK16 was stably expressed in immortalized Cdk16 −/− MEFs ( + CDK16). Control cells were infected with an empty virus. Cells were treated with 200 nM Baf. A1 and grown in EBSS. Proteins were detected as indicated ( n = 2). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Techniques: Transfection, Control, Staining, Two Tailed Test, Western Blot, Stable Transfection, Infection, Virus, Standard Deviation

a NIH3T3 cells stably expressing mCherry-GFP-LC3 were transfected with HA-CDK16 and Cyclin Y-Flag as indicated or treated for 2 h with EBSS or for 4 h with 200 nM Bafilomycin A1 (Baf. A1) and lysates were immunoblotted as indicated. KR kinase-deficient CDK16 mutant, AA CDK16 binding deficient Cyclin Y mutant ( n = 3). b Representative confocal images of the NIH3T3-mCherry-GFP-LC3 cells treated as in panel a . Staining of the HA-CDK16 in purple identified transfected cells. Autophagosomes (yellow dots) and autolysosomes (red dots) were detected by an overlay of the GFP and mCherry fluorescent signals. Scale bar: 20 µm. c Quantification of autophagosomes (yellow dots) and autolysosomes (red dots) of cells shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01. All error bars indicate SD ( n = 3; 50 cells counted for each replicate; wt vs. KR: t = 5.707, df = 4; wt vs. AA: t = 5.557, df = 4). d GFP-tagged CDK14, CDK15 or CDK16 were expressed with or without Cyclin Y-Flag in HeLa cells. Cyclin Y was immunoprecipitated with a Flag antibody (IP). Lysates were immunoblotted with the indicated antibodies (WCL). ( n = 4) e Representative confocal images of HeLa cells treated as in panel d . Fluorescent GFP signals identified CDK expressing cells. Endogenous LC3 (red) was used to measure autophagy with the 4E12 antibody. Scale bar: 50 µm. f Quantification of the LC3 dots shown in panel e . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 100 cells were counted for each treatment; control vs. Cyclin Y/CDK16: t = 6.771, df = 14; Cyclin Y/CDK14 vs. Cyclin Y/CDK16: t = 6.855, df = 14; Cyclin Y/CDK15 vs. Cyclin Y/CDK16: t = 7.139, df = 15). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a NIH3T3 cells stably expressing mCherry-GFP-LC3 were transfected with HA-CDK16 and Cyclin Y-Flag as indicated or treated for 2 h with EBSS or for 4 h with 200 nM Bafilomycin A1 (Baf. A1) and lysates were immunoblotted as indicated. KR kinase-deficient CDK16 mutant, AA CDK16 binding deficient Cyclin Y mutant ( n = 3). b Representative confocal images of the NIH3T3-mCherry-GFP-LC3 cells treated as in panel a . Staining of the HA-CDK16 in purple identified transfected cells. Autophagosomes (yellow dots) and autolysosomes (red dots) were detected by an overlay of the GFP and mCherry fluorescent signals. Scale bar: 20 µm. c Quantification of autophagosomes (yellow dots) and autolysosomes (red dots) of cells shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01. All error bars indicate SD ( n = 3; 50 cells counted for each replicate; wt vs. KR: t = 5.707, df = 4; wt vs. AA: t = 5.557, df = 4). d GFP-tagged CDK14, CDK15 or CDK16 were expressed with or without Cyclin Y-Flag in HeLa cells. Cyclin Y was immunoprecipitated with a Flag antibody (IP). Lysates were immunoblotted with the indicated antibodies (WCL). ( n = 4) e Representative confocal images of HeLa cells treated as in panel d . Fluorescent GFP signals identified CDK expressing cells. Endogenous LC3 (red) was used to measure autophagy with the 4E12 antibody. Scale bar: 50 µm. f Quantification of the LC3 dots shown in panel e . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 100 cells were counted for each treatment; control vs. Cyclin Y/CDK16: t = 6.771, df = 14; Cyclin Y/CDK14 vs. Cyclin Y/CDK16: t = 6.855, df = 14; Cyclin Y/CDK15 vs. Cyclin Y/CDK16: t = 7.139, df = 15). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Techniques: Stable Transfection, Expressing, Transfection, Mutagenesis, Binding Assay, Staining, Two Tailed Test, Immunoprecipitation, Control, Standard Deviation

a HeLa cells were co-transfected with siRNA against ULK1, Beclin1 or a control and vectors expressing GFP-CDK16 and Cyclin Y-Flag for 72 h. Lysates were immunoblotted with the indicated antibodies. ( n = 3). b Representative confocal images of HeLa cells treated as in panel a . GFP-CDK16 identified transfected cells. Endogenous LC3 staining (red) with the 4E12 antibody monitored autophagy. Scale bar: 50 µm. c Quantification of the LC3 dots shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 250 cells were analyzed for each treatment; siControl vs. siControl Cyclin Y/CDK16: t = 13.49, df = 8; siControl Cyclin Y/CDK16 vs. siULK1 Cyclin Y/CDK16: t = 13.53, df = 8; siControl Cyclin Y/CDK16 vs. siBeclin1 Cyclin Y/CDK16: t = 13.15, df = 8). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a HeLa cells were co-transfected with siRNA against ULK1, Beclin1 or a control and vectors expressing GFP-CDK16 and Cyclin Y-Flag for 72 h. Lysates were immunoblotted with the indicated antibodies. ( n = 3). b Representative confocal images of HeLa cells treated as in panel a . GFP-CDK16 identified transfected cells. Endogenous LC3 staining (red) with the 4E12 antibody monitored autophagy. Scale bar: 50 µm. c Quantification of the LC3 dots shown in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 250 cells were analyzed for each treatment; siControl vs. siControl Cyclin Y/CDK16: t = 13.49, df = 8; siControl Cyclin Y/CDK16 vs. siULK1 Cyclin Y/CDK16: t = 13.53, df = 8; siControl Cyclin Y/CDK16 vs. siBeclin1 Cyclin Y/CDK16: t = 13.15, df = 8). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Techniques: Transfection, Control, Expressing, Staining, Two Tailed Test, Standard Deviation

a Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 1 mM AICAR for 1 or 16 h. Lysates were immunoblotted with the indicated antibodies ( n = 3). b Representative confocal images of the Cdk16 +/+ and Cdk16 −/− MEFs treated as in panel a . Endogenous LC3 Staining with the 4E12 antibody monitored autophagy. Scale bar: 50 µm. c LC3 dots of experiments as shown in panel b were quantified. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01. All error bars indicate SD. ( n = 3; 100 cells counted for each replicate; Cdk16 +/+ + AICAR vs. Cdk16 −/− + AICAR: t = 4.964; df = 4). d Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 1 mM AICAR for 1 h and/or with 200 nM Bafilomycin A1 (Baf. A1) for 6 h as indicated. For the combination, AICAR was added during the last hour of Baf. A1 treatment. Lysates were immunoblotted with the indicated antibodies ( n = 1). e NIH3T3 cells were transfected with siRNA against Cyclin Y or control siRNA and stimulated with 0.5 mM AICAR/50 µM A769662 (A769) or with 200 nM Baf. A1 or a combination. Proteins were analyzed as indicated. ( n = 3). f Representative confocal images of the NIH3T3 cells treated as in panel e . Endogenous LC3 was stained with the 4E12 antibody to depict autophagy. Scale bar: 50 µm. g LC3 dots as shown in panel f were quantified. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01, *** p < 0.001. All error bars indicate SD. ( n = 3; 100 cells counted for each replicate; AICAR/A769: t = 9.494, df = 6; Baf. A1: t = 6.549, df = 6; AICAR/A769 + Baf. A1: t = 9.484, df = 4). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 1 mM AICAR for 1 or 16 h. Lysates were immunoblotted with the indicated antibodies ( n = 3). b Representative confocal images of the Cdk16 +/+ and Cdk16 −/− MEFs treated as in panel a . Endogenous LC3 Staining with the 4E12 antibody monitored autophagy. Scale bar: 50 µm. c LC3 dots of experiments as shown in panel b were quantified. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01. All error bars indicate SD. ( n = 3; 100 cells counted for each replicate; Cdk16 +/+ + AICAR vs. Cdk16 −/− + AICAR: t = 4.964; df = 4). d Immortalized Cdk16 +/+ and Cdk16 −/− MEFs were treated with 1 mM AICAR for 1 h and/or with 200 nM Bafilomycin A1 (Baf. A1) for 6 h as indicated. For the combination, AICAR was added during the last hour of Baf. A1 treatment. Lysates were immunoblotted with the indicated antibodies ( n = 1). e NIH3T3 cells were transfected with siRNA against Cyclin Y or control siRNA and stimulated with 0.5 mM AICAR/50 µM A769662 (A769) or with 200 nM Baf. A1 or a combination. Proteins were analyzed as indicated. ( n = 3). f Representative confocal images of the NIH3T3 cells treated as in panel e . Endogenous LC3 was stained with the 4E12 antibody to depict autophagy. Scale bar: 50 µm. g LC3 dots as shown in panel f were quantified. Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01, *** p < 0.001. All error bars indicate SD. ( n = 3; 100 cells counted for each replicate; AICAR/A769: t = 9.494, df = 6; Baf. A1: t = 6.549, df = 6; AICAR/A769 + Baf. A1: t = 9.484, df = 4). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Techniques: Staining, Two Tailed Test, Transfection, Control, Standard Deviation

a HeLa cells were transfected with GFP-CDK16, Cyclin Y-Flag or phosphosite mutants as indicated. The interaction of CDK16 and Cyclin Y was analyzed in GFP-specific immunoprecipitations (IP). Proteins were analyzed by immunoblotting as specified (WCL). ( n = 2). b Representative confocal images of HeLa cells from panel a . GFP-CDK16 identified transfected cells and staining for endogenous LC3 (red, antibody 4E12) monitored autophagy. Scale bar: 50 µm. c Quantification of LC3 dots per cell treated as displayed in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01; *** p < 0.001; **** p < 0.0001. All error bars indicate SD. ( n = 1; 100 cells were analyzed for each treatment; control vs. Cyclin Y/CDK16: t = 7.274, df = 11; control vs. Cyclin Y S324A/CDK16: t = 4.356, df = 8; Cyclin Y/CDK16 vs. Cyclin Y S100A/CDK16: t = 8.644, df = 13; Cyclin Y/CDK16 vs. Cyclin Y S326A/CDK16: t = 7.765, df = 13; Cyclin Y S100A/CDK16 vs. Cyclin Y S324A/CDK16: t = 5.214, df = 10; Cyclin Y S324A/CDK16 vs. Cyclin Y S326A/CDK16: t = 4.734, df = 10). d HeLa cells were transfected with GFP-CDK16 and Cyclin Y-Flag-wt or the S326A mutant and siRNA against AMPK-α1/2 or control. The interaction of CDK16 and Cyclin Y was analyzed in GFP-specific immunoprecipitations. Proteins were analyzed by immunoblotting as specified. ( n = 2). e Representative confocal images of the HeLa cells from panel d . GFP-CDK16 identified the transfected cells and staining for endogenous LC3 (red, antibody 4E12) measured autophagy. Scale bar: 50 µm. f Quantification of the LC3 dots shown in panel e . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 250 cells were analyzed for each treatment; siControl vs. siControl Cyclin Y/CDK16: t = 8.113, df = 8; siControl Cyclin Y/CDK16 vs. siAMPK-α1/2 Cyclin Y/CDK16: t = 8.290, df = 8). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: a HeLa cells were transfected with GFP-CDK16, Cyclin Y-Flag or phosphosite mutants as indicated. The interaction of CDK16 and Cyclin Y was analyzed in GFP-specific immunoprecipitations (IP). Proteins were analyzed by immunoblotting as specified (WCL). ( n = 2). b Representative confocal images of HeLa cells from panel a . GFP-CDK16 identified transfected cells and staining for endogenous LC3 (red, antibody 4E12) monitored autophagy. Scale bar: 50 µm. c Quantification of LC3 dots per cell treated as displayed in panel b . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: ** p < 0.01; *** p < 0.001; **** p < 0.0001. All error bars indicate SD. ( n = 1; 100 cells were analyzed for each treatment; control vs. Cyclin Y/CDK16: t = 7.274, df = 11; control vs. Cyclin Y S324A/CDK16: t = 4.356, df = 8; Cyclin Y/CDK16 vs. Cyclin Y S100A/CDK16: t = 8.644, df = 13; Cyclin Y/CDK16 vs. Cyclin Y S326A/CDK16: t = 7.765, df = 13; Cyclin Y S100A/CDK16 vs. Cyclin Y S324A/CDK16: t = 5.214, df = 10; Cyclin Y S324A/CDK16 vs. Cyclin Y S326A/CDK16: t = 4.734, df = 10). d HeLa cells were transfected with GFP-CDK16 and Cyclin Y-Flag-wt or the S326A mutant and siRNA against AMPK-α1/2 or control. The interaction of CDK16 and Cyclin Y was analyzed in GFP-specific immunoprecipitations. Proteins were analyzed by immunoblotting as specified. ( n = 2). e Representative confocal images of the HeLa cells from panel d . GFP-CDK16 identified the transfected cells and staining for endogenous LC3 (red, antibody 4E12) measured autophagy. Scale bar: 50 µm. f Quantification of the LC3 dots shown in panel e . Statistical significance was measured via unpaired and two-tailed Student’s t -tests and is presented as follows: **** p < 0.0001. All error bars indicate SD. ( n = 1; 250 cells were analyzed for each treatment; siControl vs. siControl Cyclin Y/CDK16: t = 8.113, df = 8; siControl Cyclin Y/CDK16 vs. siAMPK-α1/2 Cyclin Y/CDK16: t = 8.290, df = 8). n biological independent replicate. SD standard deviation. Source data are provided as a Source Data file.

Techniques: Transfection, Phospho-proteomics, Western Blot, Staining, Two Tailed Test, Control, Mutagenesis, Standard Deviation

AMPK is activated under energy stress situations (- amino acids; - glucose) or by allosteric activators (AICAR/A769) and phosphorylates Cyclin Y at S326. This phosphorylation allows the interaction of Cyclin Y with CDK16 and stimulates the kinase activity of CDK16, as illustrated by the phosphorylation of Cyclin Y at S336. The CDK16 activity downstream of AMPK is required for the efficient induction of autophagy.

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: AMPK is activated under energy stress situations (- amino acids; - glucose) or by allosteric activators (AICAR/A769) and phosphorylates Cyclin Y at S326. This phosphorylation allows the interaction of Cyclin Y with CDK16 and stimulates the kinase activity of CDK16, as illustrated by the phosphorylation of Cyclin Y at S336. The CDK16 activity downstream of AMPK is required for the efficient induction of autophagy.

Techniques: Phospho-proteomics, Activity Assay

Journal: Nature Communications

Article Title: AMPK-dependent activation of the Cyclin Y/CDK16 complex controls autophagy

doi: 10.1038/s41467-020-14812-0

Figure Lengend Snippet: Antibodies used for western blots.

Techniques: Western Blot

Review

Structured Review

Images

Similar Products

Image Search Results