SSBD:database v3

Dataset Fig2A_metaphase_5_lsm

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Project

Project
422-Asai-KinetochoreBeads
Title
Dynamics of chromosomes and artificial kinetochore beads in mouse oocytes and embryos
Description
Faithful chromosome segregation requires biorientation, where the pair of kinetochores on the chromosome establish bipolar microtubule attachment. The integrity of the kinetochore, a macromolecular complex built on centromeric DNA, is required for biorientation, but components sufficient for biorientation remain unknown. The authors show that tethering the outer kinetochore heterodimer NDC80-NUF2 to the surface of apolar microbeads establishes their biorientation-like state in mouse cells. NDC80-NUF2 microbeads align at the spindle equator and self-correct alignment errors. The alignment is associated with stable bipolar microtubule attachment and is independent of the outer kinetochore proteins SPC24-SPC25, KNL1, the Mis12 complex, inner kinetochore proteins, and Aurora. Larger microbeads align more rapidly, suggesting a size-dependent biorientation mechanism.
License
CC BY 4.0
Funding
RIKEN intramural grants; RIKEN Pioneering Project “Long-timescale Molecular Chronobiology” (T.S.K.); JSPS KAKENHI 23H04948, 21H02407, and 18H05549 (T.S.K.); Mitsubishi Foundation (T.S.K.).

Dataset

Title
Z-series images of immunocytochemistry stained for NDC80-NUF2 beads, HURP, and DNA during metaphase
Description
Z-series images of immunocytochemistry stained for NDC80-NUF2 beads, HURP (hepatoma-upregulated protein, a kinetochore fiber marker), and DNA during metaphase.NDC80-NUF2 (NDC80-NUF2 heterodimer) beads were formed in the oocyte as follows; The authors microinjected mRNAs encoding C-terminally green fluorescent protein–tagged NDC80 (NDC80-GFP) and NUF2, followed by anti-GFP–conjugated microbeads into the cytoplasm. This .lsm dataset is the processed by Airyscan in the microscope system. The original data is the dataset; Fig2A_metaphase_5_czi.
License
CC BY 4.0
Submitted at
None
Released at
None
Updated at

Biosamples

Description
The fixed mouse oocyte which was microinjected mRNAs encoding C-terminally green fluorescent protein–tagged NDC80 (NDC80-GFP) and NUF2, followed by anti-GFP–conjugated microbeads (1.7 to 2.6 mm in diameter), into the cytoplasm.
Organisms
Mus musculus (NCBI:txid10090)
Strains
B6D2F1
Cells
oocyte (CL_0000023)
Cell lines
Intrinsic variables
Extrinsic variables
Mouse oocyte was microinjected mRNAs encoding C-terminally green fluorescent protein–tagged NDC80 (NDC80-GFP) and NUF2, followed by anti-GFP–conjugated microbeads (1.7 to 2.6 mm in diameter).

Ontologies

UBERON
oocyte (CL_0000023)
Anatomical entities
MeSH: Oocytes
Gene ontology: Biological processes
female meiosis chromosome segregation (GO:0016321) kinetochore assembly (GO:0051382)
Gene ontology: Cellular components
kinetochore (GO:0000776) chromosome (GO:0005694)
Gene ontology: Molecular functions

Genetics

Gene names
Protein names
Kinetochore protein NDC80 homolog (Q9D0F1) Kinetochore protein Nuf2 (Q99P69) Disks large-associated protein 5 (Q8K4R9)
Genetic methods
injection
Protein tags
Alexa Fluor 555 (FBbi:00000443)
Probes
Hoechst 33342 (FBbi:00000052)
Oligo Primer
Genotype

Treatments

Reagent or Compound
Concentrations
FoldDilution

Imaging Methods

Dimensions
1000x1000x113x3x1
X scales
0.03 micrometer
Y scales
0.03 micrometer
Z scales
0.101 micrometer
T scales
Channels
3 channel
Microscopy types
confocal microscopy (FBbi:00000251)
Detection methods
Visualization methods
primary antibody plus labeled secondary antibody Hoechst 33342
Illumination methods
Sources of contrast
Contrast enhancing methods
Resolution enhancing methods
Airyscan
Image parameters
Sample preparation methods
fixation method

Instruments

Body
Zeiss LSM880
Model
Light source
Detector
Objective
Zeiss 40×C-Apochromat 1.2NA
Filter set
Dichroic