bims-micesi 2024-12-22 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2024–12–22
fifteen papers selected by
Valentina Piano, Uniklinik Köln

Quantitative Analysis of Kinetochore Protein Levels and Inter-Kinetochore Distances in Mammalian Cells During Mitosis.
DeSUMOylating isopeptidase 1 participates in the faithful chromosome segregation and vincristine sensitivity.
An evolutionary perspective on the relationship between kinetochore size and CENP-E dependence for chromosome alignment.
Identification of antimitotic sulfonamides inhibiting chromosome congression.
Selective regulation of kinesin-5 function by β-tubulin carboxy-terminal tails.
Phosphorylation of Ephexin4 at Ser-41 contributes to chromosome alignment via RhoG activation in cell division.
CENcyclopedia: Dynamic Landscape of Kinetochore Architecture Throughout the Cell Cycle.
Isolation and manipulation of meiotic spindles from mouse oocytes reveals migration regulated by pulling force during asymmetric division.
Base Excision Repair in Mitotic Cells and the Role of Apurinic/Apyrimidinic Endonuclease 1 (APE1) in Post-Mitotic Transcriptional Reactivation of Genes.
A novel SUN1-ALLAN complex coordinates segregation of the bipartite MTOC across the nuclear envelope during rapid closed mitosis in Plasmodium.
Binding site maturation modulated by molecular density underlies Ndc80 binding to kinetochore receptor CENP-T.
Molecular mechanism targeting condensin for chromosome condensation.
Anapc5 and Anapc7 as genetic modifiers of KIF18A function in fertility and mitotic progression.
Lgl resets Par complex membrane loading at mitotic exit to enable asymmetric neural stem cell division.
Nuclear Overexpression of SAMHD1 Induces M Phase Stalling in Hepatoma Cells and Suppresses HCC Progression by Interacting with the Cohesin Complex.

Bio Protoc. 2024 Dec 05. 14(23): e5132

Quantitative Analysis of Kinetochore Protein Levels and Inter-Kinetochore Distances in Mammalian Cells During Mitosis.

Neeraj Wasnik, Mahima Singhal, Sukirti Khantwal, Sanghamitra Mylavarapu, Sivaram V S Mylavarapu.

  The mammalian kinetochore is a multi-layered protein complex that forms on the centromeric chromatin. The kinetochore serves as the attachment hub for the plus ends of microtubules emanating from the centrosomes during mitosis. For karyokinesis, bipolar kinetochore-microtubule attachment and subsequent microtubule depolymerization lead to the development of inter-kinetochore tension between the sister chromatids. These events are instrumental in initiating a signaling cascade culminating in the segregation of the sister chromatids equally between the new daughter cells. Of the hundreds of conserved proteins that constitute the mammalian kinetochore, many that reside in the outermost layer are loaded during early mitosis and removed around metaphase-anaphase. Dynamically localized kinetochore proteins include those required for kinetochore-microtubule attachment, spindle assembly checkpoint proteins, various kinases, and molecular motors. The abundance of these kinetochore-localized proteins varies at prometaphase, metaphase, and anaphase, and is thus considered diagnostic of the fidelity of progression through these stages of mitosis. Here, we document detailed, state-of-the-art methodologies based on high-resolution fluorescence confocal microscopy followed by quantification of the levels of kinetochore-localized proteins during mitosis. We also document methods to accurately measure distances between sister kinetochores in mammalian cells, a surrogate readout for inter-kinetochore tension, which is essential for chromosome segregation. Key features • Immunostaining of cultured and suitably fixed adherent mammalian cells growing as monolayers. • Confocal fluorescence imaging for the purpose of fluorescence quantification. • 2D and 3D image reconstruction and analysis of the acquired images using appropriate background correction and normalization. • Quantification of the inter-sister kinetochore distances using 3D image reconstruction. Graphical overview Key steps involved in fluorescence quantification of spindle assembly checkpoint (SAC) protein loading at the kinetochores and in the calculation of inter-sister kinetochore distances. High-resolution confocal image datasets of cells subjected to appropriate treatment as per the experimental need (drugs, inhibitors, gene-specific siRNA, etc.) are used for quantifying the levels of SAC proteins at the kinetochores, measuring inter-sister kinetochore distances, or both. Quantification and analysis of both parameters are performed using image analysis software such as Fiji (open source) or the IMARIS software suite. Figure created with BioRender.com.

Keywords:  Confocal fluorescence microscopy; Fluorescence quantification; Kinetochore; Metaphase; Mitosis; Prometaphase

DOI:  https://doi.org/10.21769/BioProtoc.5132
FASEB J. 2024 Dec 13. 38(24): e70261

DeSUMOylating isopeptidase 1 participates in the faithful chromosome segregation and vincristine sensitivity.

Yuki Ikeda, Ryuzaburo Yuki, Youhei Saito, Yuji Nakayama.

  SUMOylation, the modification of proteins with a small ubiquitin-like modifier (SUMO), is known to regulate various cellular events, including cell division. This process is dynamic, with its status depending on the balance between SUMOylation and deSUMOylation. While the regulation of cell division by sentrin-specific protease (SENP) family proteins through deSUMOylation has been investigated, the role of another deSUMOylase, deSUMOylating isopeptidase 1 (DESI1), remains unknown. In this study, we explored DESI1's role in cell division. Knockdown of DESI1 accelerated cell division progression, leading to a significant increase in abnormal chromosome segregation. These phenotypes were rescued by re-expression of wild-type DESI1, but not catalytically inactive DESI1. DESI1 knockdown reduced the mitotic arrest caused by nocodazole, suggesting DESI1's involvement in the spindle assembly checkpoint (SAC). Localization of Aurora B, a key SAC regulator, at the metaphase chromosomes was reduced due to decreased Aurora B expression upon DESI1 knockdown. Consistently, DESI1 knockdown reduced transcription of FoxM1 target genes, such as Aurora B, cyclin B1, and CENP-F. The TCGA database showed that both decreased and increased DESI1 expression levels are associated with poor prognosis in patients with certain cancer types. Importantly, we found that DESI1 knockdown reduced sensitivity to vincristine by inducing mitotic slippage. These results suggest that DESI1 is required for faithful chromosome segregation via regulating FoxM1 transcriptional activity and thereby SAC activity in an isopeptidase activity-dependent manner. Our findings identified DESI1 as a novel regulator of cell division and a factor affecting cancer chemotherapy.

Keywords:  Aurora B; DESI1; FoxM1; SUMO; deSUMOylase; spindle assembly checkpoint; vincristine

DOI:  https://doi.org/10.1096/fj.202401560RR
J Cell Sci. 2024 Dec 15. pii: jcs263466. [Epub ahead of print]137(24):

An evolutionary perspective on the relationship between kinetochore size and CENP-E dependence for chromosome alignment.

Ana C Almeida, Helder Rocha, Maximilian W D Raas, Hanh Witte, Ralf J Sommer, Berend Snel, Geert J P L Kops, Reto Gassmann, Helder Maiato.

  Chromosome alignment during mitosis can occur as a consequence of bi-orientation or is assisted by the CENP-E (kinesin-7) motor at kinetochores. We previously found that Indian muntjac chromosomes with larger kinetochores bi-orient more efficiently and are biased to align in a CENP-E-independent manner, suggesting that CENP-E dependence for chromosome alignment negatively correlates with kinetochore size. Here, we used targeted phylogenetic profiling of CENP-E in monocentric (localized centromeres) and holocentric (centromeres spanning the entire chromosome length) clades to test this hypothesis at an evolutionary scale. We found that, despite being present in common ancestors, CENP-E was lost more frequently in taxa with holocentric chromosomes, such as Hemiptera and Nematoda. Functional experiments in two nematodes with holocentric chromosomes in which a CENP-E ortholog is absent (Caenorhabditis elegans) or present (Pristionchus pacificus) revealed that targeted expression of human CENP-E to C. elegans kinetochores partially rescued chromosome alignment defects associated with attenuated polar-ejection forces, whereas CENP-E inactivation in P. pacificus had no detrimental effects on mitosis and viability. These data showcase the dispensability of CENP-E for mitotic chromosome alignment in species with larger kinetochores.

Keywords:  CENP-E; Chromosome; Holocentric; Kinesin; Kinetochore; Mitosis

DOI:  https://doi.org/10.1242/jcs.263466
Biochem Pharmacol. 2024 Dec 17. pii: S0006-2952(24)00719-6. [Epub ahead of print] 116718

Identification of antimitotic sulfonamides inhibiting chromosome congression.

Jun-Ichi Sawada, Kenji Matsuno, Naohisa Ogo, Akira Asai.

  The discovery of new small-molecule inhibitors is essential to enhancing our understanding of biological events at the molecular level and driving advancements in drug discovery. Mitotic inhibitors have played a crucial role in development of anticancer drugs. Beyond traditional microtubule inhibitors, various inhibitors targeting specific mitotic factors have been developed. This study aimed to develop novel mitotic inhibitors targeting chromosome alignment. We established a cell-based screening method using Cell Division Cycle Associated 5 (CDCA5) and kinesin-5 as markers, designed to efficiently detect mitotic phenotypes characterized by aberrant bipolar spindles with some misaligned chromosomes. Through this screening, we identified CAIS-1, an aryl sulfonamide with unique antimitotic properties. CAIS-1 exhibits dual functionality by inhibiting chromosome congression at low concentrations and spindle microtubule formation at high concentrations, causing a concentration-dependent mitotic arrest, followed by apoptotic cell death. Mechanistic studies revealed that CAIS-1 directly acts on tubulin at high concentrations, thereby inhibiting tubulin polymerization in vitro. In contrast, at low concentrations, CAIS-1 functions through a mechanism distinct from GSK923295, a conventional chromosome congression inhibitor targeting Centromere-associated protein-E (CENP-E), highlighting its unique mode of action. Moreover, CAIS-2, a structural analog of CAIS-1, selectively inhibits chromosome congression without significantly affecting spindle microtubules. This observation suggests that CAIS-1 and CAIS-2 function as antimitotic sulfonamides with distinct targets beyond tubulin, thus offering additional biological potential of sulfonamide compounds. Together, CAIS-1 and CAIS-2 represent promising tools for providing new molecular insights into kinetochore function during mitosis and for exploring new approaches in anticancer drug development.

Keywords:  Chromosome; Congression; Inhibitor; Mitosis; Screening; Spindle

DOI:  https://doi.org/10.1016/j.bcp.2024.116718
J Cell Biol. 2025 Mar 03. pii: e202405115. [Epub ahead of print]224(3):

Selective regulation of kinesin-5 function by β-tubulin carboxy-terminal tails.

Ezekiel C Thomas, Jeffrey K Moore.

The tubulin code hypothesis predicts that tubulin tails create programs for selective regulation of microtubule-binding proteins, including kinesin motors. However, the molecular mechanisms that determine selective regulation and their relevance in cells are poorly understood. We report selective regulation of budding yeast kinesin-5 motors by the β-tubulin tail. Cin8, but not Kip1, requires the β-tubulin tail for recruitment to the mitotic spindle, creating a balance of both motors in the spindle and efficient mitotic progression. We identify a negatively charged patch in the β-tubulin tail that mediates interaction with Cin8. Using in vitro reconstitution with genetically modified yeast tubulin, we demonstrate that the charged patch of β-tubulin tail increases Cin8 plus-end-directed velocity and processivity. Finally, we determine that the positively charged amino-terminal extension of Cin8 coordinates interactions with the β-tubulin tail. Our work identifies a molecular mechanism underlying selective regulation of closely related kinesin motors by tubulin tails and how this regulation promotes proper function of the mitotic spindle.

DOI: https://doi.org/10.1083/jcb.202405115
J Biol Chem. 2024 Dec 13. pii: S0021-9258(24)02586-9. [Epub ahead of print] 108084

Phosphorylation of Ephexin4 at Ser-41 contributes to chromosome alignment via RhoG activation in cell division.

Ryuji Yasutake, Hiroki Kuwajima, Ryuzaburo Yuki, Junna Tanaka, Youhei Saito, Yuji Nakayama.

  Ephexin proteins are guanine nucleotide exchange factors for the Rho GTPases. We reported that Ephexin4 regulates M-phase progression downstream of phosphorylated EphA2, a receptor-type tyrosine kinase, through RhoG activation; however, the regulation of Ephexin4 during M phase remains unknown. In this study, a novel Ephexin4 phosphorylation site was identified at Ser41, exclusively in M phase. Ephexin4 knockdown prolonged the duration of M phase by activating the spindle assembly checkpoint, at which BubR1 was localized at the kinetochores of the misaligned chromosomes. This delay was alleviated by re-expression of wild-type, but not S41A Ephexin4. The Ephexin4 knockdown caused chromosome misalignment and reduced the RhoG localization to the plasma membrane. These phenotypes were rescued by re-expression of wild-type and phospho-mimic S41E mutant, but not the S41A mutant. Consistently, S41E mutant enhanced active RhoG levels, even in the interphase. Regardless of the Ephexin4 knockdown, active RhoG-G12V was localized at the plasma membrane. Furthermore, Ephexin4 knockdown exacerbated vincristine-induced chromosome misalignment, which was prevented by re-expressing the wild-type but not S41A Ephexin4. Overexpression of wild-type and S41E mutant, but not S41A mutant, resulted in an increased number of Madin-Darby canine kidney (MDCK) cysts with cells inside the lumen, indicating disruption of epithelial morphogenesis by deregulating Ephexin4/RhoG signaling in cell division. Our results suggest that Ephexin4 undergoes phosphorylation at Ser41 in cell division, and the phosphorylation is required for chromosome alignment through RhoG activation. Combined with mitosis-targeting agents, inhibition of Ephexin4 phosphorylation may represent a novel strategy for cancer chemotherapy.

Keywords:  EphA2; Ephexin4; GEF; RhoG; chromosome misalignment; phosphorylation

DOI:  https://doi.org/10.1016/j.jbc.2024.108084
bioRxiv. 2024 Dec 05. pii: 2024.12.05.627000. [Epub ahead of print]

CENcyclopedia: Dynamic Landscape of Kinetochore Architecture Throughout the Cell Cycle.

Yu-Chia Chen, Ece Kilic, Evelyn Wang, Will Rossman, Aussie Suzuki.

The kinetochore, an intricate macromolecular protein complex located on chromosomes, plays a pivotal role in orchestrating chromosome segregation. It functions as a versatile platform for microtubule assembly, diligently monitors microtubule binding fidelity, and acts as a force coupler. Comprising over 100 distinct proteins, many of which exist in multiple copies, the kinetochore's composition dynamically changes throughout the cell cycle, responding to specific timing and conditions. This dynamicity is important for establishing functional kinetochores, yet the regulatory mechanisms of these dynamics have largely remained elusive. In this study, we employed advanced quantitative immunofluorescence techniques to meticulously chart the dynamics of kinetochore protein levels across the cell cycle. These findings offer a comprehensive view of the dynamic landscape of kinetochore architecture, shedding light on the detailed mechanisms of microtubule interaction and the nuanced characteristics of kinetochore proteins. This study significantly advances our understanding of the molecular coordination underlying chromosome segregation.

DOI: https://doi.org/10.1101/2024.12.05.627000
bioRxiv. 2024 Dec 08. pii: 2024.12.06.627260. [Epub ahead of print]

Isolation and manipulation of meiotic spindles from mouse oocytes reveals migration regulated by pulling force during asymmetric division.

Ning Liu, Ryo Kawamura, Wenan Qiang, Ahmed Balboula, John F Marko, Huanyu Qiao.

Spindles are essential for accurate chromosome segregation in all eukaryotic cells. This study presents a novel approach for isolating fresh mammalian spindles from mouse oocytes, establishing it as a valuable in vitro model system for a wide range of possible studies. Our method enables the investigation of the physical properties and migration force of meiotic spindles in oocytes. We found that the spindle length decreases upon isolation from the oocyte. Combining this observation with direct measurements of spindle mechanics, we examined the forces governing spindle migration during oocyte asymmetric division. Our findings suggest that the spindle migration is regulated by a pulling force and a net tensile force of approximately 680 pN is applied to the spindle in vivo during the migration process. This method, unveiling insights into spindle dynamics, holds promise as a robust model for future investigations into spindle formation and chromosome separation. We also found that the same approach could not isolate spindles from somatic cells, indicative of mammalian oocytes having a unique spindle organization amenable to isolation.

DOI: https://doi.org/10.1101/2024.12.06.627260
Int J Mol Sci. 2024 Nov 27. pii: 12735. [Epub ahead of print]25(23):

Base Excision Repair in Mitotic Cells and the Role of Apurinic/Apyrimidinic Endonuclease 1 (APE1) in Post-Mitotic Transcriptional Reactivation of Genes.

Suravi Pramanik, Yingling Chen, Kishor K Bhakat.

  Endogenous DNA damage occurs throughout the cell cycle, with cells responding differently at various stages. The base excision repair (BER) pathway predominantly repairs damaged bases in the genome. While extensively studied in interphase cells, it is unknown if BER operates in mitosis and how apurinic/apyrimidinic (AP) sites, intermediates in the BER pathway that inhibit transcriptional elongation, are processed for post-mitotic gene reactivation. In this study, using an alkaline comet assay, we demonstrate that BER is inefficient in mitosis and that AP endonuclease 1 (APE1), a key BER enzyme, is required for the repair of damage post-mitosis. We previously demonstrated that APE1 is acetylated (AcAPE1) in the chromatin. Using high-resolution microscopy, we show that AcAPE1 remains associated with specific regions in the condensed chromatin in each of the phases of mitosis. This association presumably occurs via the binding of APE1 to the G-quadruplex structure, a non-canonical DNA structure predominantly present in the transcribed gene regions. Additionally, using a nascent RNA detection strategy, we demonstrate that the knockdown of APE1 delayed the rapid post-mitotic transcriptional reactivation of genes. Our findings highlight the functional importance of APE1 in the mitotic chromosomes to facilitate faster repair of endogenous damage and rapid post-mitotic gene reactivation in daughter cells.

Keywords:  DNA damage; DNA repair; acetylated APE1; base excision repair; mitosis

DOI:  https://doi.org/10.3390/ijms252312735
bioRxiv. 2024 Dec 05. pii: 2024.12.04.625416. [Epub ahead of print]

A novel SUN1-ALLAN complex coordinates segregation of the bipartite MTOC across the nuclear envelope during rapid closed mitosis in Plasmodium.

Mohammad Zeeshan, Igor Blatov, Ryuji Yanase, David J P Ferguson, Sarah L Pashley, Zeinab Chahine, Yoshiki Yamaryo Botté, Akancha Mishra, Baptiste Marché, Suhani Bhanvadia, Molly Hair, Sagar Batra, Robert Markus, Declan Brady, Andrew Bottrill, Sue Vaughan, Cyrille Y Botté, Karine Le Roch, Anthony A Holder, Eelco C Tromer, Rita Tewari.

Mitosis in eukaryotes involves reorganization of the nuclear envelope (NE) and microtubule-organizing centres (MTOCs). In Plasmodium , the causative agent of malaria, male gametogenesis mitosis is exceptionally rapid and divergent. Within 8 minutes, the haploid male gametocyte genome undergoes three replication cycles (1N to 8N), while maintaining an intact NE. Axonemes assemble in the cytoplasm and connect to a bipartite MTOC-containing nuclear pole and cytoplasmic basal body, producing eight flagellated gametes. The mechanisms coordinating NE remodelling, MTOC dynamics, and flagellum assembly remain poorly understood. Here, we identify the SUN1-ALLAN complex as a novel mediator of NE remodelling and bipartite MTOC coordination during Plasmodium male gametogenesis. SUN1, a conserved NE protein, localizes to dynamic loops and focal points near nuclear spindle poles. ALLAN, a divergent Allantoicase-like protein, has a location like that of SUN1 at nuclear MTOCs. SUN1 and ALLAN form a unique complex, detected by live-cell imaging, ultrastructural expansion microscopy, and interactomics. Deletion of either SUN1 or ALLAN gene disrupts nuclear MTOC organization, leading to basal body mis-segregation, defective spindle assembly, and impaired kinetochore attachment, but axoneme formation remains intact. Ultrastructural analysis revealed nuclear and cytoplasmic MTOC miscoordination, producing aberrant flagellated gametes lacking nuclear material. Sun1 deletion also alters parasite lipid composition, underscoring its role in NE homeostasis. These defects block parasite development in the mosquito and transmission, highlighting the essential functions of this complex. This study reveals a bipartite MTOC and a highly divergent mechanism of NE remodelling during Plasmodium male gametogenesis. The SUN1-ALLAN complex is an unusual adaptation of the LINC complex, in absence of canonical KASH-domain proteins in Plasmodium , providing new insights into the evolution of closed mitosis and highlighting potential targets for blocking malaria transmission.

DOI: https://doi.org/10.1101/2024.12.04.625416
Proc Natl Acad Sci U S A. 2024 Dec 24. 121(52): e2401344121

Binding site maturation modulated by molecular density underlies Ndc80 binding to kinetochore receptor CENP-T.

Ekaterina V Tarasovetc, Gunter B Sissoko, Aleksandr Maiorov, Anna S Mukhina, Fazoil I Ataullakhanov, Iain M Cheeseman, Ekaterina L Grishchuk.

  Macromolecular assembly depends on tightly regulated pairwise binding interactions that are selectively favored at assembly sites while being disfavored in the soluble phase. This selective control can arise due to molecular density-enhanced binding, as recently found for the kinetochore scaffold protein CENP-T. When clustered, CENP-T recruits markedly more Ndc80 complexes than its monomeric counterpart, but the underlying molecular basis remains elusive. Here, we use quantitative in vitro assays to reveal two distinct mechanisms driving this behavior. First, Ndc80 binding to CENP-T is a two-step process: initially, Ndc80 molecules rapidly associate and dissociate from disordered N-terminal binding sites on CENP-T. Over time, these sites undergo maturation, resulting in stronger Ndc80 retention. Second, we find that this maturation transition is regulated by a kinetic barrier that is sensitive to the molecular environment. In the soluble phase, binding site maturation is slow, but within CENP-T clusters, this process is markedly accelerated. Notably, the two Ndc80 binding sites in human CENP-T exhibit distinct maturation rates and environmental sensitivities, which correlate with their different amino acid content and predicted binding conformations. This clustering-induced maturation is evident in dividing human cells, suggesting a distinct regulatory entry point for controlling kinetochore assembly. We propose that the tunable acceleration of binding site maturation by molecular crowding may represent a general mechanism for promoting the formation of macromolecular structures.

Keywords:  binding site maturation; kinetic barrier; kinetochore; molecular density; single-molecule binding assay

DOI:  https://doi.org/10.1073/pnas.2401344121
EMBO J. 2024 Dec 17.

Molecular mechanism targeting condensin for chromosome condensation.

Menglu Wang, Daniel Robertson, Juan Zou, Christos Spanos, Juri Rappsilber, Adele L Marston.

  Genomes are organised into DNA loops by the Structural Maintenance of Chromosomes (SMC) proteins. SMCs establish functional chromosomal sub-domains for DNA repair, gene expression and chromosome segregation, but how SMC activity is specifically targeted is unclear. Here, we define the molecular mechanism targeting the condensin SMC complex to specific chromosomal regions in budding yeast. A conserved pocket on the condensin HAWK subunit Ycg1 binds to chromosomal receptors carrying a related motif, CR1. In early mitosis, CR1 motifs in receptors Sgo1 and Lrs4 recruit condensin to pericentromeres and rDNA, to facilitate sister kinetochore biorientation and rDNA condensation, respectively. We additionally find that chromosome arm condensation begins as sister kinetochores come under tension, in a manner dependent on the Ycg1 pocket. We propose that multiple CR1-containing proteins recruit condensin to chromosomes and identify several additional candidates based on their sequence. Overall, we uncover the molecular mechanism that targets condensin to functionalise chromosomal domains to achieve accurate chromosome segregation during mitosis.

Keywords:  Condensin; Lrs4; Pericentromeres; Shugoshin; rDNA

DOI:  https://doi.org/10.1038/s44318-024-00336-6
bioRxiv. 2024 Dec 04. pii: 2024.12.03.626395. [Epub ahead of print]

Anapc5 and Anapc7 as genetic modifiers of KIF18A function in fertility and mitotic progression.

Carleigh Nesbit, Whitney Martin, Anne Czechanski, Candice Byers, Narayanan Raghupathy, Ardian Ferraj, Jason Stumpff, Laura Reinholdt.

The kinesin family member 18A ( KIF18A ) is an essential regulator of microtubule dynamics and chromosome alignment during mitosis. Functional dependency on KIF18A varies by cell type and genetic context but the heritable factors that influence this dependency remain unknown. To address this, we took advantage of the variable penetrance observed in different mouse strain backgrounds to screen for loci that modulate germ cell depletion in the absence of KIF18A. We found a significant association at a Chr5 locus where anaphase promoting complex subunits 5 ( Anapc5 ) and 7 ( Anapc7 ) were the top candidate genes. We found that both genes were differentially expressed in a sensitive strain background when compared to resistant strain background at key timepoints in gonadal development. We also identified a novel retroviral insertion in Anapc7 that may in part explain the observed expression differences. In cell line models, we found that depletion of KIF18A induced mitotic arrest, which was partially rescued by co-depletion of ANAPC7 (APC7) and exacerbated by co-depletion of ANAPC5 (APC5). These findings suggest that differential expression and activity of Anapc5 and Anapc7 may influence sensitivity to KIF18A depletion in germ cells and CIN cells, with potential implications for optimizing antineoplastic therapies.

DOI: https://doi.org/10.1101/2024.12.03.626395
bioRxiv. 2024 Dec 02. pii: 2024.09.29.615680. [Epub ahead of print]

Lgl resets Par complex membrane loading at mitotic exit to enable asymmetric neural stem cell division.

Bryce LaFoya, Sarah E Welch, Kenneth E Prehoda.

The Par complex regulates cell polarity in diverse animal cells 1-4 , but how its localization is restricted to a specific membrane domain remains unclear. We investigated how the tumor suppressor Lethal giant larvae (Lgl) polarizes the Par complex in Drosophila neural stem cells (NSCs or neuroblasts). In contrast to epithelial cells, where Lgl and the Par complex occupy mutually exclusive membrane domains, Lgl is cytoplasmic when the Par complex is apically polarized in NSCs5. Importantly, we found that Lgl's key function is not in directly regulating metaphase Par polarity, but rather in removing the Par complex from the membrane at the end of mitosis, creating a "polarity reset" for the next cell cycle. Without this Lgl-mediated reset, we found that residual Par complex remains on the basal membrane during subsequent divisions, disrupting fate determinant polarization and proper asymmetric cell division. These findings reveal a novel mechanism of polarity regulation by Lgl and highlight the importance of the prepolarized state in Par-mediated polarity.

DOI: https://doi.org/10.1101/2024.09.29.615680
Adv Sci (Weinh). 2024 Dec 16. e2411988

Nuclear Overexpression of SAMHD1 Induces M Phase Stalling in Hepatoma Cells and Suppresses HCC Progression by Interacting with the Cohesin Complex.

Juntang Shao, Wei Wang, Shiyu Li, Guangfa Yin, Lili Han, Xinyu Wang, Meng Cai, Tao Yang, Ying Wang, Wenyan Qu, Yanhong Jiao, Peng Wang, Hanyang Xu, Xu Zhu, Songcheng Ying, Sa Xu, Qiang Sheng, Jian Fang, Tongcui Jiang, Chuansheng Wei, Yujun Shen, Yuxian Shen.

  Emerging evidence suggests that the sterile alpha-motif (SAM) and histidine-aspartate (HD) domain-containing protein 1 (SAMHD1) is implicated in various cancers, including hepatocellular carcinoma (HCC). However, its precise role in tumor cells and the underlying mechanisms remain unclear. This study aimed to investigate the expression patterns, prognostic values, and functional role of SAMHD1 in HCC progression. We constructed liver tissue microarrays using tumor and paired paratumor tissue specimens from 187 patients with primary HCC. Our findings indicate that nuclear SAMHD1 protein levels are increased in tumors compared to paratumor tissues. Moreover, nuclear SAMHD1 levels decline in advanced tumor stages, with higher SAMHD1 nuclear staining correlating with favorable prognostic outcomes. Hepatocyte-specific SAMHD1 knockout mice, generated by crossing SAMHD1fl/fl mice with Alb-cre mice, showed accelerated tumor progression in a diethylnitrosamine (DEN)-induced HCC model. In hepatoma cell lines, nuclear overexpression of SAMHD1 inhibited cell proliferation by stalling mitosis, independent of its deoxynucleotide triphosphohydrolase (dNTPase) function. Mechanistically, SAMHD1 interacts with the cohesin complex in nucleus, enhancing sister chromatid cohesion during cell division, which delays metaphase progression. Our findings suggest that nuclear SAMHD1 plays a critical role in slowing HCC progression by regulating mitosis, highlighting its potential as a therapeutic target by manipulating cohesin dynamics.

Keywords:  HCC; SAMHD1; cell cycle; cohesin complex

DOI:  https://doi.org/10.1002/advs.202411988