Journal: bioRxiv

Article Title: Suv420 enrichment at the centromere limits Aurora B localization and function

doi: 10.1101/2021.06.14.448441

Figure Lengend Snippet: A) Quantification of anaphase defects showing that cells induced to express cen-Suv39 or cen-Suv420 exhibit an increase in anaphase lagging chromosomes following low nanomolar concentrations of Aurora kinase inhibition while their uninduced counterparts do not. Cells both with and without induction of cen-Suv39-GFP or cen-Suv420-GFP are similarly sensitive to inhibition of the mitotic kinase MPS1. (+/- cen-Suv39-GFP induction: p= 0.00003, 0.0003, and 0.844 for Alisertib, Barasertib, and MPS1IN1, respectively; +/-cen-Suv420-GFP: p = 0.003, 0.004, and 0.724 for Alisertib, Barasertib, and MPS1IN1, respectively) Error bars are +/- SD. B & C) Cen-INCENP-mCherry localizes to centromeres in cen-Suv420-GFP expressing cells. D) Expression of cen-INCENP-mCherry does not limit cen-Suv420-GFP localization to centromeres. GFP-staining intensity between ACA peaks was measured for 3 kinetochore pairs in each of 30 metaphase cells per condition, in each of 3 biological replicates. E) expression of cen-INCENP-mCherry reduces the frequency of lagging chromosomes seen in cen-Suv420-GFP expressing anaphase cells. A minimum of 30 anaphase cells were scored per condition, for each of 3 biological replicates. **: p < 0.01, ***: p < 0.001. F) Quantification of area under the dose response curve (AUC) for each drug in cancer cell lines from the TCGA sorted by top and bottom quartile of either Suv39 or Suv420 expression shows that cells with high expression of either Suv39 or Suv420 exhibit increased sensitivity to Aurora kinase inhibition. Each dot represents an individual cancer cell line tested. Open circles indicate the mean of IC50 AUCs for each condition. *: p < 0.00625, **: p < 0.00125, ***: p <0.000125. Error bars are +/- SEM.

Article Snippet: Expression of centromere-targeted, GFP-tagged cen-GFP, cen-Suv39-GFP and cen-Suv420-GFP fusion proteins, and non-targeted Suv420-GFP was achieved by cloning the respective cDNA into Addgene vector CENP-B DBD INCENP GFP (45237, Addgene) at NheI / BamH1.

Techniques: Inhibition, Expressing, Staining

Journal: Developmental cell

Article Title: CENP-A Ubiquitylation Is Indispensable to Cell Viability

doi: 10.1016/j.devcel.2019.07.015

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse monoclonal anti-CENP-B , Novus Biologicals , H00001059-B01P.

Techniques: Affinity Purification, Recombinant, Modification, Protease Inhibitor, Western Blot, Stripping, Software, Imaging, Laser-Scanning Microscopy, Fluorescence, Microscopy, Membrane

Journal: Current Biology

Article Title: Meiotic Kinetochores Fragment into Multiple Lobes upon Cohesin Loss in Aging Eggs

doi: 10.1016/j.cub.2019.09.006

Figure Lengend Snippet:

Article Snippet: To generate the kinetochore labeling construct, CENPB -mEmerald (Addgene, 54037) was subcloned into pGEMHE vector using the NheI and NotI restrictions sites, while other expression constructs were previously described.

Techniques: Produced, Recombinant, Software