bims-micesi 2023-10-15 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2023‒10‒15
six papers selected by
Valentina Piano, Uniklinik Köln

Nuclear lamin A-associated proteins are required for centromere assembly.
Probing Mitotic Chromosome Mechanics Using Optical Tweezers.
Cytoplasmic division cycles without the nucleus and mitotic CDK/cyclin complexes.
Growing and dividing: how O-GlcNAcylation leads the way.
M phase-specific interaction between SBDS and RNF2 at the mitotic spindles regulates mitotic progression.
SETD2 maintains nuclear lamina stability to safeguard the genome.

bioRxiv. 2023 Sep 26. pii: 2023.09.25.559341. [Epub ahead of print]

Nuclear lamin A-associated proteins are required for centromere assembly.

Adriana Landeros, Destiny A Wallace, Amit Rahi, Christine B Magdongon, Praveen Suraneni, Mohammed A Amin, Manas Chakraborty, Stephen A Adam, Daniel R Foltz, Dileep Varma.

Many Lamin A-associated proteins (LAAP's) that are key constituents of the nuclear envelope (NE), assemble at the "core" domains of chromosomes during NE reformation and mitotic exit. However, the identity and function of the chromosomal core domains remain ill-defined. Here, we show that a distinct section of the core domain overlaps with the centromeres/kinetochores of chromosomes during mitotic telophase. The core domain can thus be demarcated into a kinetochore proximal core (KPC) on one side of the segregated chromosomes and the kinetochore distal core (KDC) on the opposite side, close to the central spindle. We next tested if centromere assembly is connected to NE re-formation. We find that centromere assembly is markedly perturbed after inhibiting the function of LMNA and the core-localized LAAPs, BANF1 and Emerin. We also find that the LAAPs exhibit multiple biochemical interactions with the centromere and inner kinetochore proteins. Consistent with this, normal mitotic progression and chromosome segregation was severely impeded after inhibiting LAAP function. Intriguingly, the inhibition of centromere function also interferes with the assembly of LAAP components at the core domain, suggesting a mutual dependence of LAAP and centromeres for their assembly at the core domains. Finally, we find that the localization of key proteins involved in the centromeric loading of CENP-A, including the Mis18 complex and HJURP were markedly affected in LAAP-inhibited cells. Our evidence points to a model where LAAP assembly at the core domain serves a key function in loading new copies of centromeric proteins during or immediately after mitotic exit.

DOI: https://doi.org/10.1101/2023.09.25.559341
Methods Mol Biol. 2024 ;2694 91-107

Probing Mitotic Chromosome Mechanics Using Optical Tweezers.

Tinka V M Clement, Constantijn van der Smagt, Gijs J L Wuite.

  During mitosis, cells compact their DNA into rodlike shapes, four orders of magnitude shorter than the DNA backbone contour length. We describe an experimental protocol to isolate and study these intricate mitotic chromosomes using optical tweezers. We touch upon the technical details of the required optical tweezers and microfluidics setup, including advanced force calibration procedures to accurately measure the high forces the chromosomes withstand. The procedure used to isolate mitotic chromosomes, including biotinylation of the telomeric ends to facilitate trapping them in optical tweezers, is described in detail. Finally, we provide a protocol for carrying out optical tweezers experiments on the isolated mitotic chromosomes.

Keywords:  Chromosome mechanics; Force calibration for optical traps; Microfluidics; Mitotic chromosome isolation; Mitotic chromosomes; Optical tweezers

DOI:  https://doi.org/10.1007/978-1-0716-3377-9_5
Cell. 2023 Oct 12. pii: S0092-8674(23)01031-0. [Epub ahead of print]186(21): 4694-4709.e16

Cytoplasmic division cycles without the nucleus and mitotic CDK/cyclin complexes.

Anand Bakshi, Fabio Echegaray Iturra, Andrew Alamban, Miquel Rosas-Salvans, Sophie Dumont, Mustafa G Aydogan.

  Cytoplasmic divisions are thought to rely on nuclear divisions and mitotic signals. We demonstrate in Drosophila embryos that cytoplasm can divide repeatedly without nuclei and mitotic CDK/cyclin complexes. Cdk1 normally slows an otherwise faster cytoplasmic division cycle, coupling it with nuclear divisions, and when uncoupled, cytoplasm starts dividing before mitosis. In developing embryos where CDK/cyclin activity can license mitotic microtubule (MT) organizers like the spindle, cytoplasmic divisions can occur without the centrosome, a principal organizer of interphase MTs. However, centrosomes become essential in the absence of CDK/cyclin activity, implying that the cytoplasm can employ either the centrosome-based interphase or CDK/cyclin-dependent mitotic MTs to facilitate its divisions. Finally, we present evidence that autonomous cytoplasmic divisions occur during unperturbed fly embryogenesis and that they may help extrude mitotically stalled nuclei during blastoderm formation. We postulate that cytoplasmic divisions occur in cycles governed by a yet-to-be-uncovered clock mechanism autonomous from CDK/cyclin complexes.

Keywords:  Cdk; Drosophila embryo; autonomous clocks; cell cycle; centrosome; cyclin; cytokinesis; epithelial homeostasis; microtubules; mitosis

DOI:  https://doi.org/10.1016/j.cell.2023.09.010
J Biol Chem. 2023 Oct 09. pii: S0021-9258(23)02358-X. [Epub ahead of print] 105330

Growing and dividing: how O-GlcNAcylation leads the way.

Harmony Saunders, Wagner B Dias, Chad Slawson.

  Cell cycle errors can lead to mutations, chromosomal instability, or death; thus, the precise control of cell cycle progression is essential for viability. The nutrient-sensing post-translational modification, O-GlcNAc, regulates the cell cycle allowing one central control point directing progression of the cell cycle. O-GlcNAc is a single N-acetylglucosamine sugar modification to intracellular proteins that is dynamically added and removed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), respectively. These enzymes act as a rheostat to fine-tune protein function in response to a plethora of stimuli from nutrients to hormones. O-GlcNAc modulates mitogenic growth signaling, senses nutrient flux through the hexosamine biosynthetic pathway, and coordinates with other nutrient-sensing enzymes to progress cells through Gap phase 1 (G1). At the G1/S transition, O-GlcNAc modulates checkpoint control, while in S Phase O-GlcNAcylation coordinates the replication fork. DNA replication errors activate O-GlcNAcylation to control the function of the tumor suppressor p53 at Gap Phase 2 (G2). Finally, in Mitosis (M phase), O-GlcNAc controls M phase progression and the organization of the mitotic spindle and midbody. Critical for M phase control is the interplay between OGT and OGA with mitotic kinases. Importantly, disruptions in OGT and OGA activity induce M phase defects and aneuploidy. These data point to an essential role for the O-GlcNAc rheostat in regulating cell division. In this review, we highlight O-GlcNAc nutrient sensing regulating G1; O-GlcNAc control of DNA replication and repair; and finally, O-GlcNAc organization of mitotic progression and spindle dynamics.

Keywords:  Cyclin; Mini-chromosome complex; Nutrient sensing; O-GlcNAc; O-GlcNAc Transferase; O-GlcNAcase; Spindle; cell cycle; mTOR; p53

DOI:  https://doi.org/10.1016/j.jbc.2023.105330
Biochem Biophys Res Commun. 2023 Oct 04. pii: S0006-291X(23)01157-9. [Epub ahead of print]682 118-123

M phase-specific interaction between SBDS and RNF2 at the mitotic spindles regulates mitotic progression.

Yukihiro Sera, Tsuneo Imanaka, Masafumi Yamaguchi.

  Shwachman-Diamond syndrome (SDS) is an autosomal recessive inherited disorder caused by biallelic mutations in the Shwachman-Bodian-Diamond syndrome (SBDS) gene. SBDS protein is involved in ribosome biogenesis; therefore SDS is classified as a ribosomopathy. SBDS is localized at mitotic spindles and stabilizes microtubules. Previously, we showed that SBDS interacts with ring finger protein 2 (RNF2) and is degraded through RNF2-dependent ubiquitination. In this study, we investigated when and where SBDS interacts with RNF2 and the effects of the interaction on cells. We found that SBDS co-localized with RNF2 on centrosomal microtubules in the mitotic phase (M phase), whereas SBDS and RNF2 localized to the nucleolus and nucleoplasm in the interphase, respectively. The microtubule-binding assay revealed that SBDS interacted directly with microtubules and RNF2 interacted with SBDS bound to microtubules. In addition, SBDS was ubiquitinated and degraded by RNF2 during the M phase. Moreover, RNF2 overexpression accelerated mitotic progression. These findings suggest that SBDS delays mitotic progression, and RNF2 releases cells from suppression through the ubiquitination and subsequent degradation of SBDS. The interaction between SBDS and RNF2 at mitotic spindles might be involved in mitotic progression as a novel regulatory cascade.

Keywords:  Mitotic progression; Mitotic spindle; RNF2; SBDS; SDS

DOI:  https://doi.org/10.1016/j.bbrc.2023.10.013
bioRxiv. 2023 Sep 28. pii: 2023.09.28.560032. [Epub ahead of print]

SETD2 maintains nuclear lamina stability to safeguard the genome.

Abid Khan, James M Metts, Lucas C Collins, C Allie Mills, Kelin Li, Amanda L Brademeyer, Brittany M Bowman, M Ben Major, Jeffrey Aubé, Laura E Herring, Ian J Davis, Brian D Strahl.

Histone methyltransferases play essential roles in the organization and function of chromatin. They are also frequently mutated in human diseases including cancer 1 . One such often mutated methyltransferase, SETD2, associates co-transcriptionally with RNA polymerase II and catalyzes histone H3 lysine 36 trimethylation (H3K36me3) - a modification that contributes to gene transcription, splicing, and DNA repair 2 . While studies on SETD2 have largely focused on the consequences of its catalytic activity, the non-catalytic functions of SETD2 are largely unknown. Here we report a catalysis-independent function of SETD2 in maintaining nuclear lamina stability and genome integrity. We found that SETD2, via its intrinsically disordered N-terminus, associates with nuclear lamina proteins including lamin A/C, lamin B1, and emerin. Depletion of SETD2, or deletion of its N-terminus, resulted in widespread nuclear morphology abnormalities and genome stability defects that were reminiscent of a defective nuclear lamina. Mechanistically, the N-terminus of SETD2 facilitates the association of the mitotic kinase CDK1 with lamins, thereby promoting lamin phosphorylation and depolymerization required for nuclear envelope disassembly during mitosis. Taken together, our findings reveal an unanticipated link between the N-terminus of SETD2 and nuclear lamina organization that may underlie how SETD2 acts as a tumor suppressor.

DOI: https://doi.org/10.1101/2023.09.28.560032