bims-micesi 2022-11-20 papers

Issue of 2022–11–20
five papers selected by
Valentina Piano, Uniklinik Köln

Curr Biol. 2022 Nov 11. pii: S0960-9822(22)01695-5. [Epub ahead of print]

MPS1 localizes to end-on microtubule-attached kinetochores to promote microtubule release.

Daniel Hayward, Emile Roberts, Ulrike Gruneberg.

In eukaryotes, the spindle assembly checkpoint protects genome stability in mitosis by preventing chromosome segregation until incorrect microtubule-kinetochore attachment geometries have been eliminated and chromosome biorientation has been completed. These error correction and checkpoint processes are linked by the conserved Aurora B and MPS1 Ser/Thr kinases.1,2 MPS1-dependent checkpoint signaling is believed to be initiated by kinetochores without end-on microtubule attachments,3,4 including those generated by Aurora B-mediated error correction. The current model posits that MPS1 competes with microtubules for binding sites at the kinetochore.3,4 MPS1 is thought to first recognize kinetochores not blocked by microtubules and then initiate checkpoint signaling. However, MPS1 is also required for chromosome biorientation and correction of microtubule-kinetochore attachment errors.5,6,7,8,9 This latter function, which must require direct interaction with microtubule-attached kinetochores, is not readily explained within the constraints of the current model. Here, we show that MPS1 transiently localizes to end-on attached kinetochores and that this recruitment depends on the relative activities of Aurora B and its counteracting phosphatase PP2A-B56 rather than microtubule-attachment state per se. MPS1 autophosphorylation also regulates MPS1 kinetochore levels but does not determine the response to microtubule attachment. At end-on attached kinetochores, MPS1 actively promotes microtubule release together with Aurora B. Furthermore, in live cells, MPS1 is detected at attached kinetochores before the removal of microtubules. During chromosome alignment, MPS1, therefore, coordinates both the resolution of incorrect microtubule-kinetochore attachments and the initiation of spindle checkpoint signaling.

Keywords: Aurora B; MPS1; PP2A-B56; cell division; chromosome; kinase; kinetochore; mitosis; phosphatase; spindle assembly checkpoint

DOI: https://doi.org/10.1016/j.cub.2022.10.047

Life Sci Alliance. 2023 Jan;pii: e202201540. [Epub ahead of print]6(1):

Endoplasmic reticulum membranes are continuously required to maintain mitotic spindle size and forces.

Margarida Araújo, Alexandra Tavares, Diana V Vieira, Ivo A Telley, Raquel A Oliveira.

Membrane organelle function, localization, and proper partitioning upon cell division depend on interactions with the cytoskeleton. Whether membrane organelles also impact the function of cytoskeletal elements remains less clear. Here, we show that acute disruption of the ER around spindle poles affects mitotic spindle size and function in Drosophila syncytial embryos. Acute ER disruption was achieved through the inhibition of ER membrane fusion by the dominant-negative cytoplasmic domain of atlastin. We reveal that when centrosome-proximal ER membranes are disrupted, specifically at metaphase, mitotic spindles become smaller, despite no significant changes in microtubule dynamics. These smaller spindles are still able to mediate sister chromatid separation, yet with decreased velocity. Furthermore, by inducing mitotic exit, we found that nuclear separation and distribution are affected by ER disruption. Our results suggest that ER integrity around spindle poles is crucial for the maintenance of mitotic spindle shape and pulling forces. In addition, ER integrity also ensures nuclear spacing during syncytial divisions.

DOI: https://doi.org/10.26508/lsa.202201540

Chromosome Res. 2022 Nov 18.

Ellman's reagent prevents dephosphorylation of histones during isolation of mitotic chromosomes.

James R Paulson, Erica R Vander Mause, Elizabeth Dillinger, Megan E Luedeman, Bakhtawar Usman.

Histones H1 and H3 are highly phosphorylated in mitotic HeLa cells but are rapidly dephosphorylated by endogenous protein phosphatases during the isolation of metaphase chromosomes. We show that this dephosphorylation can be prevented by including the sulfhydryl reagent 5,5'-dithiobis-(2-nitrobenzoate) (Ellman's reagent, or DTNB) in the isolation buffer. The minimal amount of DTNB required is approximately stoichiometric with the number of sulfhydryl groups in the lysate. Inhibition of the protein phosphatases can subsequently be reversed by treatment with dithiothreitol or 2-mercaptoethanol. DTNB is compatible with the isolation of either metaphase chromosome clusters or individual metaphase chromosomes. It should be useful in investigations of the structure and biochemistry of chromatin and chromosomes and in the study of possible functions for mitotic histone phosphorylation.

Keywords: Chromosome structure; Histone H1; Mitosis; Phosphatases; Protein phosphorylation

DOI: https://doi.org/10.1007/s10577-022-09709-1

J Biol Chem. 2022 Nov 14. pii: S0021-9258(22)01146-2. [Epub ahead of print] 102703

The SCF-FBXW7 E3 ubiquitin ligase triggers degradation of Histone 3 Lysine 4 methyltransferase complex component WDR5 to prevent mitotic slippage.

Simon Hänle-Kreidler, Kai T Richter, Ingrid Hoffmann.

During prolonged mitotic arrest induced by anti-microtubule drugs, cell fate decision is determined by two alternative pathways, one leading to cell death, the other inducing premature escape from mitosis by mitotic slippage. FBWX7, a member of the F-box family of proteins and substrate-targeting subunit of the SCF (SKP1-CUL1-F-Box) E3 ubiquitin ligase complex promotes mitotic cell death and prevents mitotic slippage, but molecular details underlying these roles for FBWX7 are unclear. In this study, we report that WDR5, a component of the mixed lineage leukemia (MLL) complex of Histone 3 Lysine 4 (H3K4) methyltransferases, is a substrate of FBXW7. We determined by co-immunoprecipitation experiments and in vitro binding assays that WDR5 interacts with FBXW7 in vivo and in vitro. SCF-FBXW7 mediates ubiquitination of WDR5 and targets it for proteasomal degradation. Furthermore, we find that WDR5 depletion counteracts FBXW7 loss-of-function by reducing mitotic slippage and polyploidization. In conclusion, our data elucidate a new mechanism in mitotic cell fate regulation which might contribute to prevent chemotherapy resistance in patients after anti-microtubule drug treatment.

Keywords: FBXW7; WDR5; chemoresistance; mitotic slippage; ubiquitylation (ubiquitination)

DOI: https://doi.org/10.1016/j.jbc.2022.102703

Int J Biochem Cell Biol. 2022 Nov 15. pii: S1357-2725(22)00178-9. [Epub ahead of print] 106333

APC/C CDH1 Ubiquitinates STAT3 in Mitosis.

Debanjan Bhattacharjee, Sreeram Kaveti, Nishant Jain.

STAT3, an oncogene drives tumor growth and is associated with poor prognosis. However, small molecule-based STAT3 inhibitors were unsuccessful in clinics. Recently, STAT3 degraders that ubiquitinate STAT3 were found to elicit long-lasting anti-tumor responses. Thus, triggering STAT3 ubiquitination in cancers is a better strategy than STAT3 inhibition. However, not much is known about the identity of E3-ligases that ubiquitinate STAT3 in cancers. Therefore, to design better therapies to degrade STAT3, we sought to identify E3-ligases that ubiquitinate STAT3 in cancer cells. To answer this question, we determined the cell cycle-dependent ubiquitination of STAT3 in HEK293T cells and examined the link between STAT3 dephosphorylation and ubiquitination. We found that STAT3 is more strongly ubiquitinated in mitosis than in other phases of the cell cycle. We observed that APC/C CDH1 binds and ubiquitinates STAT3 in mitosis. Further, we also found that inhibiting phosphatases decreases STAT3 ubiquitination. We conclude that APC/C CDH1 ubiquitinates STAT3 in mitosis. We suggest that mitosis can be a potential therapeutic window for treating STAT3-activated cancers.

Keywords: CDH1; STAT3; Ubiquitination; mitosis; phosphorylation

DOI: https://doi.org/10.1016/j.biocel.2022.106333