Journal: BMC Evolutionary Biology

Article Title: Evolution of motif variants and positional bias of the cyclic-AMP response element

doi: 10.1186/1471-2148-7-S1-S15

Figure Lengend Snippet: Summary of TSS-SRs from 9 species

Article Snippet: Gallus gallus , galGal2 , RefSeq Genes , 1,692.

Techniques:

Journal: The EMBO Journal

Article Title: Parthenolide disrupts mitosis by inhibiting ZNF207/BUGZ-promoted kinetochore-microtubule attachment

doi: 10.1038/s44318-025-00469-2

Figure Lengend Snippet: ( A ) Volcano plot showing quantitative mass-spectrometry results. Dashed horizontal line shows the p -value cut-off ( p < 0.05) and vertical dashed lines indicate the upregulated/downregulated (competed/non-competed by free PTL) proteins. The green transparent region groups all the proteins that satisfy the p -value cut-off and are upregulated (competed) with a SILAC ratio higher than 2. N (number of experiments): 3. Statistical analysis was performed using unpaired t-test. ( B ) Representative spinning disk confocal time-series of mitosis in HeLa parental cells and HeLa stably expressing GFP-BUGZ undergoing indicated treatments. Scale bar: 10 µm. ( C ) Quantification of chromosome congression status and mitotic duration in HeLa parental cells and HeLa stably expressing GFP-BUGZ undergoing indicated treatments. Median is plotted for mitotic duration. N , n ( N = number of cells, n = number of experiments) for congression phenotype: HeLa + DMSO (44, 3), HeLa + 15 µM PTL (41, 3), HeLa GFP-BUGZ + DMSO (85, 3), HeLa GFP-BUGZ + 15 µM PTL (30, 3), HeLa GFP-BUGZ + 30 µM PTL (56, 3), HeLa GFP-BUGZ + 45 µM PTL (58, 3); N , n ( N = number of cells, n = number of experiments) for mitotic duration: HeLa + DMSO (40, 3), HeLa + 15 µM PTL (36, 3), HeLa GFP-BUGZ + DMSO (85, 3), HeLa GFP-BUGZ + 15 µM PTL (30, 3), HeLa GFP-BUGZ + 30 µM PTL (56, 3), HeLa GFP-BUGZ + 45 µM PTL (58, 3). .

Article Snippet: The BUGZ gene (human, NCBI RefSeq: NM_001032293.3 ) encoding wild-type BUGZ and C54A mutant were commercially synthesized (GenScript) as siRNA-resistant sequences into a pGenDONR vector (pGenDONR-BUGZ and pGenDONR-BUGZ C54A).

Techniques: Mass Spectrometry, Multiplex sample analysis, Stable Transfection, Expressing

Journal: The EMBO Journal

Article Title: Parthenolide disrupts mitosis by inhibiting ZNF207/BUGZ-promoted kinetochore-microtubule attachment

doi: 10.1038/s44318-025-00469-2

Figure Lengend Snippet: ( A ) Representative spinning-disk confocal time-series of mitosis in HeLa cells stably expressing GFP-BUGZ and infected with adenovirus to express H2B-RFP. Scale bar: 10 µm. ( B ) Representative spinning-disk confocal time-series of mitosis in control, 15 µM PTL- and siBUGZ-treated U2OS cells stably expressing H2B-GFP/mScarlet-α-tubulin. Scale bar: 10 µm.

Article Snippet: The BUGZ gene (human, NCBI RefSeq: NM_001032293.3 ) encoding wild-type BUGZ and C54A mutant were commercially synthesized (GenScript) as siRNA-resistant sequences into a pGenDONR vector (pGenDONR-BUGZ and pGenDONR-BUGZ C54A).

Techniques: Stable Transfection, Expressing, Infection, Control

Journal: The EMBO Journal

Article Title: Parthenolide disrupts mitosis by inhibiting ZNF207/BUGZ-promoted kinetochore-microtubule attachment

doi: 10.1038/s44318-025-00469-2

Figure Lengend Snippet: ( A ) Fluorencence and Coomassie staining of immunoprecipitated FLAG-BUGZ from HEK 293T cells treated either with DMSO or 15 µM alkyne-PTL. ( B ) Representative spinning-disk confocal maximum projections of click-based imaging of 15 µM fluor-488-alkyne-PTL in nocodazole arrested U2OS cells co-stained with α-tubulin and CENP-C, as markers for microtubules and kinetochores, respectively. Scale bar: 10 µm. ( C ) Quantification of the relative levels of 15 µM fluor-488-alkyne-PTL at kinetochores. N , n (number of cells, number of experiments): siNT (38, 4) siBUGZ (40, 4). *** p ≤0.001. ( D ) Representative confocal maximum projections of BUB1 immunostainings in nocodazole arrested U2OS cells treated with DMSO or 15 µM PTL. Scale bar: 10 µm. ( E ) Quantification of the relative levels of BUB1 at kinetochores. N , n ( N = number of cells, n = number of experiments): DMSO (40, 4), 15 µM PTL (40, 4). ( F ) Extracted ion chromatograms for peptides with and without PTL modification at Cys54 (red, and green, respectively). ( G ) Extent of PTL-modification at Cys54 from 3 independent experiments. ( H ) AlphaFold 3 model of BUGZ Zinc Finger domains and Cys54 positioning showing the catalytic mechanism leading to PTL selectivity. Replicates are color-coded for all quantifications. Data in ( C ) and ( E ) are presented as mean ± SD values, while data in ( G ) are presented in mean ± SEM values. Statistical analysis was performed by unpaired t-test with Welch’s correction in ( C ) and Mann-Whitney test in ( E ). .

Article Snippet: The BUGZ gene (human, NCBI RefSeq: NM_001032293.3 ) encoding wild-type BUGZ and C54A mutant were commercially synthesized (GenScript) as siRNA-resistant sequences into a pGenDONR vector (pGenDONR-BUGZ and pGenDONR-BUGZ C54A).

Techniques: Staining, Immunoprecipitation, Imaging, Modification, MANN-WHITNEY

Journal: The EMBO Journal

Article Title: Parthenolide disrupts mitosis by inhibiting ZNF207/BUGZ-promoted kinetochore-microtubule attachment

doi: 10.1038/s44318-025-00469-2

Figure Lengend Snippet: ( A ) Fluorencence and Comassie staining of immunoprecipitated FLAG empty and FLAG-BUGZ from HEK 293T cells treated alkyne-PTL. ( B ) Representative confocal images of click-based imaging of 5 µM fluor-Cy5-alkyne-PTL in HeLa GFP-BUGZ cells under the indicated conditions. Scale bar: 10 µm. ( C ) Scatter plot showing the intensity of BUGZ (x-axis) and 5 µM fluor-Cy5-alkyne-PTL (y-axis) at individual kinetochores from the indicated conditions in ( B ). Each dot represents a single kinetochore. A Pearson correlation line is shown for the correlation between BUGZ and fluor-Cy5-alkyne-PTL levels. ( D ) Quantification of 5 µM fluor-Cy5-alkyne-PTL intensity at kinetochores normalized to CENP-C intensity for the conditions indicated in ( B ). N , n (number of cells, number of experiments): siNT (19, 3) siBUGZ (21, 3). **** p ≤ 0.0001. Replicates are color coded. Data are presented as mean ± SD values from three independent replicates. Statistical analysis was performed using unpaired t-test. ( E ) Immunoblot for BUGZ depletion efficiency in HeLa GFP-BUGZ cells. ( F ) Illustration of domain architecture of BUGZ. ( G ) Western-blot with anti-BUGZ antibody of in cellulo GFP-Trap pulldown sample from HeLa cells stably expressing GFP-BUGZ treated with 50 µM PTL. ( H ) Extracted ion chromatograms and MS-MS spectra for peptides with and without PTL modification at Cys54 (red and green, respectively) from the GFP-Trap sample shown in ( G ).

Article Snippet: The BUGZ gene (human, NCBI RefSeq: NM_001032293.3 ) encoding wild-type BUGZ and C54A mutant were commercially synthesized (GenScript) as siRNA-resistant sequences into a pGenDONR vector (pGenDONR-BUGZ and pGenDONR-BUGZ C54A).

Techniques: Staining, Immunoprecipitation, Imaging, Western Blot, Stable Transfection, Expressing, Tandem Mass Spectroscopy, Modification