Visualizing Cartwheel Disassembly Process During Mitosis in Fixed and Live Cells by Fluorescence Microscope.

Centrosome is an evolutionarily conserved organelle that comprises two barrel-shaped centrioles surrounded by pericentriolar material (PCM). It functions as the major microtubule-organizing center (MTOC) to regulate cell polarity, motility, intracellular material transport during interphase, and bipolar spindle assembly during mitosis. Cartwheel assembly is considered the first step in the initiation of procentriole biogenesis at early S phase.

STAG2 promotes naive-primed transition via activating Lin28a transcription in mouse embryonic stem cells.

Mouse embryonic stem cells (mESCs) exist in two distinct pluripotent states: the naive and the primed. Mainly by inducing differentiation of mESCs in vitro, conducting RNA sequencing analyses, and specifying expression of the regulatory genes, we explored the regulatory mechanisms underlying the transition between the naive and primed states. We found that, under the defined differentiation-inducing conditions, the naive state of mESCs shifted to the primed state within 2 days of differentiation induction, during which the cell cycle- and differentiation-related proteins changes significantly.

Mitotic ER-mitochondria contact enhances mitochondrial Ca influx to promote cell division.

Cell division is tightly regulated and requires an expanded energy supply. However, how this energy is generated remains unclear. Here, we establish a correlation between two mitochondrial Ca influx events and ATP production during mitosis.

TIP60 acetylation of Bub1 regulates centromeric H2AT120 phosphorylation for faithful chromosome segregation.

Proper function of the centromeres ensures correct attachment of kinetochores to spindle microtubules and faithful chromosome segregation in mitosis. Defects in the integrity and function of centromeres can result in chromosome missegregation and genomic instability. Bub1 is essential for the mitotic centromere dynamics, yet the underlying molecular mechanisms remain largely unclear.

cell-free extracts and their applications in cell biology study.

has proven to be a remarkably versatile model organism in the realm of biological research for numerous years, owing to its straightforward maintenance in laboratory settings and its abundant provision of ample-sized oocytes, eggs, and embryos. The cell cycle of these oocytes, eggs, and early embryos exhibits synchrony, and extracts derived from these cells serve various research purposes. Many fundamental concepts in biochemistry, cell biology, and development have been elucidated through the use of cell-free extracts derived from cells.

NuSAP regulates microtubule flux and Kif2A localization to ensure accurate chromosome congression.

Precise chromosome congression and segregation requires the proper assembly of a steady-state metaphase spindle, which is dynamic and maintained by continuous microtubule flux. NuSAP is a microtubule-stabilizing and -bundling protein that promotes chromosome-dependent spindle assembly. However, its function in spindle dynamics remains unclear.

Cartwheel disassembly regulated by CDK1-cyclin B kinase allows human centriole disengagement and licensing.

Cartwheel assembly is considered the first step in the initiation of procentriole biogenesis; however, the reason for persistence of the assembled human cartwheel structure from S phase to late mitosis remains unclear. Here, we demonstrate mainly using cell synchronization, RNA interference, immunofluorescence and time-lapse-microscopy, biochemical analysis, and methods that the cartwheel persistently assembles and maintains centriole engagement and centrosome integrity during S phase to late G2 phase. Blockade of the continuous accumulation of centriolar Sas-6, a major cartwheel protein, after procentriole formation induces premature centriole disengagement and disrupts pericentriolar matrix integrity.

8 Å structure of the outer rings of the Xenopus laevis nuclear pore complex obtained by cryo-EM and AI.

The nuclear pore complex (NPC), one of the largest protein complexes in eukaryotes, serves as a physical gate to regulate nucleocytoplasmic transport. Here, we determined the 8 Å resolution cryo-electron microscopic (cryo-EM) structure of the outer rings containing nuclear ring (NR) and cytoplasmic ring (CR) from the Xenopus laevis NPC, with local resolutions reaching 4.9 Å.

A design optimized prime editor with expanded scope and capability in plants.

The ability to manipulate the genome in a programmable manner has illuminated biology and shown promise in plant breeding. Prime editing, a versatile gene-editing approach that directly writes new genetic information into a specified DNA site without requiring double-strand DNA breaks, suffers from low efficiency in plants. In this study, N-terminal reverse transcriptase-Cas9 nickase fusion performed better in rice than the commonly applied C-terminal fusion.

NuMA regulates mitotic spindle assembly, structural dynamics and function via phase separation.

A functional mitotic spindle is essential for accurate chromosome congression and segregation during cell proliferation; however, the underlying mechanisms of its assembly remain unclear. Here we show that NuMA regulates this assembly process via phase separation regulated by Aurora A. NuMA undergoes liquid-liquid phase separation during mitotic entry and KifC1 facilitates NuMA condensates concentrating on spindle poles.

SUMO proteases SENP3 and SENP5 spatiotemporally regulate the kinase activity of Aurora A.

Precise chromosome segregation is mediated by a well-assembled mitotic spindle, which requires balance of the kinase activity of Aurora A (AurA, also known as AURKA). However, how this kinase activity is regulated remains largely unclear. Here, using in vivo and in vitro assays, we report that conjugation of SUMO2 with AurA at K258 in early mitosis promotes the kinase activity of AurA and facilitates the binding with its activator Bora.

Sufu negatively regulates both initiations of centrosome duplication and DNA replication.

Centrosome duplication and DNA replication are two pivotal events that higher eukaryotic cells use to initiate proliferation. While DNA replication is initiated through origin licensing, centrosome duplication starts with cartwheel assembly and is partly controlled by CP110. However, the upstream coordinator for both events has been, until now, a mystery.

Live-cell fluorescence imaging of ciliary dynamics.

The cilium was one of the first organelles observed through a microscope. Motile cilia appear as oscillating cell appendages and have long been recognized to function in cell motility. In contrast, the far more widespread non-motile cilia, termed primary cilia, were thought to be vestigial and largely ignored following their initial description over a century ago.

PLK4-phosphorylated NEDD1 facilitates cartwheel assembly and centriole biogenesis initiations.

Centrosome duplication occurs under strict spatiotemporal regulation once per cell cycle, and it begins with cartwheel assembly and daughter centriole biogenesis at the lateral sites of the mother centrioles. However, although much of this process is understood, how centrosome duplication is initiated remains unclear. Here, we show that cartwheel assembly followed by daughter centriole biogenesis is initiated on the NEDD1-containing layer of the pericentriolar material (PCM) by the recruitment of SAS-6 to the mother centriole under the regulation of PLK4.

Aurora B regulates PP1γ-Repo-Man interactions to maintain the chromosome condensation state.

The mitotic kinase Aurora B regulates the condensation of chromatin into chromosomes by phosphorylating chromatin proteins during early mitosis, whereas the phosphatase PP1γ performs the opposite function. The roles of Aurora B and PP1γ must be tightly coordinated to maintain chromosomes at a high phosphorylation state, but the precise mechanisms regulating their function remain largely unclear. Here, mainly through immunofluorescence microscopy and co-immunoprecipitation assays, we find that dissociation of PP1γ from chromosomes is essential for maintaining chromosome phosphorylation.

K6-linked SUMOylation of BAF regulates nuclear integrity and DNA replication in mammalian cells.

Barrier-to-autointegration factor (BAF) is a highly conserved protein in metazoans that has multiple functions during the cell cycle. We found that BAF is SUMOylated at K6, and that this modification is essential for its nuclear localization and function, including nuclear integrity maintenance and DNA replication. K6-linked SUMOylation of BAF promotes binding and interaction with lamin A/C to regulate nuclear integrity.

Homozygous mutations in can induce asthenoteratospermia with severe MMAF.

Background: Asthenoteratospermia, one of the most common causes for male infertility, often presents with defective sperm heads and/or flagella. Multiple morphological abnormalities of the sperm flagella (MMAF) is one of the common clinical manifestations of asthenoteratospermia. Variants in several genes including , , and are involved in the genetic pathogenesis of asthenoteratospermia.

Phosphorylation of importin-α1 by CDK1-cyclin B1 controls mitotic spindle assembly.

Importin-α serves as an adaptor linking importin-β to proteins carrying a nuclear localization sequence (NLS). During interphase, this interaction enables nuclear protein import, while in mitosis it regulates spindle assembly factors (SAFs) and controls microtubule nucleation, stabilization and spindle function. Here, we show that human importin-α1 is regulated during the cell cycle and is phosphorylated at two sites (threonine 9 and serine 62) during mitosis by the major mitotic protein kinase CDK1-cyclin B.

Postmitotic annulate lamellae assembly contributes to nuclear envelope reconstitution in daughter cells.

In higher eukaryotic cells, the nuclear envelope (NE) is composed of double nuclear membranes studded with nuclear pore complexes (NPCs) and undergoes dynamic disassembly and reassembly during the cell cycle. However, how the NE and NPC reassemble remains largely unclear. Here, using HeLa, HEK293, and cells, along with immunofluorescence microscopy and transmission EM methods, we found that postmitotic annulate lamellae (AL) assembly contributes to NE and NPC assembly.

The microtubule-associated protein EML3 regulates mitotic spindle assembly by recruiting the Augmin complex to spindle microtubules.

In all eukaryotes, a functional mitotic spindle is essential for distributing duplicated chromosomes into daughter cells. Mitotic spindle assembly involves highly ordered arrangement of microtubules (MTs). The Augmin protein complex recruits γ-tubulin ring complex (γ-TuRC) to MTs and thereby promotes MT-based MT nucleation and mitotic spindle assembly.

Patched1-ArhGAP36-PKA-Inversin axis determines the ciliary translocation of Smoothened for Sonic Hedgehog pathway activation.

The Sonic Hedgehog (Shh) pathway conducts primarily in the primary cilium and plays important roles in cell proliferation, individual development, and tumorigenesis. Shh ligand binding with its ciliary membrane-localized transmembrane receptor Patched1 results in the removal of Patched1 from and the translocation of the transmembrane oncoprotein Smoothened into the cilium, leading to Shh signaling activation. However, how these processes are coupled remains unknown.

The Plk1 kinase negatively regulates the Hedgehog signaling pathway by phosphorylating Gli1.

Hedgehog (Hh) signaling is a highly conserved cell signaling pathway important for cell life, development and tumorigenesis. Increasing evidence suggests that the Hh signaling pathway functions in certain phases of the cell cycle. However, the coordination between Hh signaling and cell cycle control remains poorly understood.

DNA replication licensing factor Cdc6 and Plk4 kinase antagonistically regulate centrosome duplication via Sas-6.

Centrosome number is tightly controlled during the cell cycle to ensure proper spindle assembly and cell division. However, the underlying mechanism that controls centrosome number remains largely unclear. We show herein that the DNA replication licensing factor Cdc6 is recruited to the proximal side of the centrioles via cyclin A to negatively regulate centrosome duplication by binding and inhibiting the cartwheel protein Sas-6 from forming a stable complex with another centriole duplication core protein, STIL.