Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: Expression of CDK11 and CK2 protein complex members in untransformed and malignant breast cells. (A) Immunoblot analysis of cultured breast cell lines, as indicated above the blots. Proteins detected are indicated on the right side of the blots. Actin signal was used as the loading control. (B) Indirect immunofluorescent detection of CDK11, CK2α, and CK2α′ (red color) in breast cell lines. Cell lines are indicated above each set of images and proteins detected are indicated on the left side of the images. Blue, 4′,6-diamidino-2-phenylindole-stained nuclei. Scale bar: 100 μm. (C) Immunohistochemical detection of CDK11 proteins in human normal and malignant breast tissue. Type of breast tissue indicated on the left side of the images. Magnification indicated above the images; dotted ellipse, portion of the 100× image that is shown at 400×. Scale bars: 400 μm for 100× and 100 μm for 400× images. (D) Human microarray tissues stained for CDK11 were scored by two independent observers. The average value was taken and the results plotted for normal ( n = 16) versus triple-negative breast cancer (TNBC; n = 44) tissues. Box, first to third (Q1 to Q3) quartiles; diamond, mean; line inside box, median; whiskers, minimum and maximums of data range. CDK, cyclin-dependent kinase; CK2, casein kinase 2.

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Expressing, Western Blot, Cell Culture, Staining, Immunohistochemical staining, Microarray

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: mRNA expression levels in nontransformed and malignant breast cells a

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Expressing

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: RNA expression levels in normal breast and breast cancer subtypes. Normalized RNAseq read count data for PAM50 breast cancer subtypes and normal breast from The Cancer Genome Atlas were analyzed for CDK11 and CK2 protein complex genes as shown above each plot. Box, first to third (Q1 to Q3) quartiles; line inside box, median; whiskers, 1.5 maximum interquartile range. Normal, n = 95; basal, n = 141; Her2, n = 67; LumA, n = 421; LumB, n = 192. CDK, cyclin-dependent kinase; CK2, casein kinase 2; Her2, human epidermal growth factor receptor 2; LumA, luminal A; LumB, luminal B.

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: RNA Expression

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: Immunoblot analyses following small interfering RNA-mediated downregulation of CDK11 and CK2 in breast cancer cells. Immunoblot analysis of MDA-MB-231 and SUM-149 cell lysates following small interfering RNA (siRNA) transfection. Transfected siRNAs are indicated above the blots, proteins detected are indicated on the right side of the blots. CDK11 p110 , cyclin L1α, cyclin L2α, and CK2αα′β lysates are 72 hours post transfection; caspase 3, Bcl-xL, and survivin lysates are 96 hours post transfection. Actin signal was used as the loading control. CDK, cyclin-dependent kinase; CK2, casein kinase 2.

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Western Blot, Small Interfering RNA, Transfection

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: Protein expression levels following small interfering RNA transfection

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Expressing, Small Interfering RNA

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: mRNA expression levels in small interfering RNA transfected cells

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Expressing, Small Interfering RNA, Transfection

Journal: Breast Cancer Research : BCR

Article Title: Preclinical evaluation of cyclin dependent kinase 11 and casein kinase 2 survival kinases as RNA interference targets for triple negative breast cancer therapy

doi: 10.1186/s13058-015-0524-0

Figure Lengend Snippet: Small interfering RNA-mediated downregulation of CDK11 and CK2 in breast cancer cells decreases cell viability and inhibits clonal survival. (A) Breast cancer cells were transfected with 30 nM single small interfering RNA (siRNA) or 15 nM each of the two siRNAs combined as indicated. After 96 hours, cell viability was determined relative to the untreated cells. Means ± standard errors (SEs) are presented. * P <0.05. ** P <0.01, *** P <0.001 relative to untreated. ^ P = 0.055, # P <0.05, ## P <0.01, ### P <0.001 relative to siCtrl. (B) Triple-negative breast cancer (TNBC) cells were transfected twice with 30 nM single siRNAs or 15 nM each of the two siRNAs combined as indicated and as described in . Seven days after the second transfection, cell colonies were stained and counted. Means ± SE are presented. $ P <0.0001 relative to siCtrl and untreated. (C) Representative crystal violet stained colonies on 35 mm plates 7 days after the second siRNA transfection as described in (B). Cell lines are indicated above the plate images and siRNA transfections are indicated to the left of the plate images. CDK, cyclin-dependent kinase; CK2, casein kinase 2; si, small interfering.

Article Snippet: Antibodies used were: CDK11 (A300-311A), cyclin L1 (A302-058A), CK2α (A300-197A) and CK2α′ (A300-199A) from Bethyl Laboratories; cyclin L2 (600-401-878) from Rockland Immunochemicals (Limerick, PA, USA); CK2β (sc-12739 and sc-46666) and actin (sc-1616) from Santa Cruz Biotechnology; CDK11 (5524), caspase 3 (9661, 9662), lamin A/C (2032), and Bcl-xL (2762) from Cell Signaling (Beverly, MA, USA); and survivin (AF886) from R&D Systems (Minneapolis, MN, USA).

Techniques: Small Interfering RNA, Transfection, Staining

Journal: American journal of physiology. Cell physiology

Article Title: HSP90 modulates the myosin replacement rate in myofibrils.

doi: 10.1152/ajpcell.00245.2017

Figure Lengend Snippet: Fig. 2. Inhibition of heat shock protein 90 (HSP90) activity slows the myosin replacement rate in myofibrils. A–L: sarcomere structures were not disrupted by geldanamycin (GM) treatment. DMSO (A–C and G–I) or 5 g/ml GM (D–F and J–L)-treated cells were stained with indicated antibodies. C, F, I, and L are merged images of A and B, D and E, G and H, and J and K. Arrowheads depict myomesin-positive M-lines in the A-bands in A–F. All images were taken with a confocal microscope. Scale bars 10 m. M–O: relative fluo- rescence intensity after bleaching of green fluorescent protein-tagged myosin heavy chain 3 (GFP-MYH3) was measured in myotubes treated with DMSO or GM. Fluorescence signals were measured at indicated time points (M). Mobile fractions (%) of GFP-MYH3 were 47.0 5.9 in DMSO and 9.6 1.2 in GM (N). Half- lives (hours) of GFP-MYH3 were 6.7 1.2 in DMSO and 3.4 1.0 in GM (O). Values represent the mean SE. *P 0.05, significant difference compared with DMSO control. DMSO control myotubes, n 7; GM-treated myotubes, n 4.

Article Snippet: The antibodies used in this study were mouse anti-myomesin antibody (1:10 dilution; clone B4; Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA) (19), mouse anti-HSP90 antibody (1:400 dilution; clone D7 ; StressMarq Biosciences), and rabbit polyclonal anti-MYH antibody (1:500 dilution; kindly gifted by the late Prof. Howard Holtzer, University of Pennsylvania, Philadelphia).

Techniques: Inhibition, Activity Assay, Staining, Microscopy, Fluorescence, Control