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Image Search Results
Journal: The Journal of Cell Biology
Article Title: Aurora A kinase phosphorylates Hec1 to regulate metaphase kinetochore–microtubule dynamics
doi: 10.1083/jcb.201707160
Figure Lengend Snippet: Hec1 S69 is phosphorylated throughout mitosis. (A) Amino acid sequences of the human and PtK1 cell Hec1 N-terminal tail domain. Shown in yellow is the human peptide sequence that was used to generate the S69 phosphospecific antibody. The arrow points to S69 in the human sequence and the corresponding serine residue in the PtK1 sequence. Asterisks indicate all other mapped Aurora B kinase sites in the human Hec1 tail domain ( ; ). (B) Immunofluorescence images of HeLa cells stained with phosphospecific antibodies to Hec1 pS69. Depletion of Hec1 (bottom) results in loss of pS69 staining at kinetochores. Cells are also immunostained with antibody 9G3 (pan-Hec1 antibody) and an anticentromere antibody (ACA) derived from human calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasia (CREST) patient serum. (C) Immunofluorescence images of HeLa cells demonstrating kinetochore localization of pS69 during mitosis. Quantification is shown on the right. For each phase shown, ≥400 kinetochores from ≥30 cells were measured. (D) Immunofluorescence images of a HeLa cell stained with antibodies to pS69 and Mad2. For the cell shown, most chromosomes are aligned at the spindle equator, and one chromosome remains near a spindle pole (arrows). A schematic illustrating examples of pole-proximal chromosomes is shown on the left. (E) Immunofluorescence images of HeLa cells depleted of CENP-E to increase the number of pole-proximal chromosomes (arrows) and stained with Hec1 phosphospecific antibodies. Quantification is shown on the right from one representative experiment. n values are as follows: pS69, 20 polar kinetochores and 40 aligned kinetochores; pS55, 17 polar kinetochores and 57 aligned kinetochores; and pS44, 13 polar kinetochores and 29 aligned kinetochores. Error bars indicate SD. Bars: (B, C, and E) 10 µm; (D) 3 µm.
Article Snippet: Two rabbits were immunized with the
Techniques: Sequencing, Residue, Immunofluorescence, Staining, Derivative Assay