Pbp1, a yeast ortholog of human ataxin-2, is important for cell growth in the medium containing non-fermentable carbon sources. We had reported that Pbp1 regulates expression of genes related to glycogenesis via transcriptional regulation and genes related to mitochondrial function through mRNA stability control. To further analyze the role of Pbp1 in gene expression, we first examined the time course of gene expression after transfer from YPD medium containing glucose to YPGlyLac medium containing glycerol and lactate. At 12 h after transfer to YPGlyLac medium, the pbp1∆ mutant showed decreased expression of genes related to mitochondrial function but no decrease in expression of glycogenesis-related genes. We also examined a role of the Pbp1-binding factor, Mkt1. The mkt1∆ mutant, like the pbp1∆ mutant, showed slow growth on YPGlyLac plate and reduced expression of genes related to mitochondrial function. Furthermore, we found that mutation of DHH1 gene encoding a decapping activator exacerbated the growth of the pbp1∆ mutant on YPGlyLac plate. The dhh1∆ mutant showed reduced expression of genes related to mitochondrial function. These results indicate that Pbp1 and Mkt1 regulate the expression of genes related to mitochondrial function and that the decapping activator Dhh1 also regulates the expression of those genes.

Abnormalities in spermatogenesis, a fundamental component of male reproductive function, can cause male infertility. Somatic cells constituting the testis microenvironment are essential for controlling normal spermatogenesis. Although testicular somatic cells are thought to sense and respond to germ cells to ensure proper spermatogenesis, the details of this signaling mechanism are unknown. Here, we investigated somatic cell dynamics in testicular tissue lacking spermatogenesis using the mice with deletion of the testis-specific histone H3 variant gene H3t. Testicular tissue sections of H3t mice exhibited an increased interstitial area compared with those of wild-type mice, which was primarily attributed to an increase in Leydig cell numbers. Furthermore, this increase in Leydig cells led to increased testosterone synthesis, which occurred alongside cellular senescence-associated β-galactosidase activity. These findings suggest that Leydig cells monitor the progress of spermatogenesis and possess a mechanism to promote functional germ cell formation.

Control of nutrient homeostasis plays a central role in cell proliferation/survival during embryonic development and tumor growth. Activation of the Notch signaling pathway, a major contributor to cell-cell interactions, is a potential mechanism for cell adaptation to nutrient-poor conditions. Our previous study also demonstrated that during embryogenesis when nutrients such as glutamine and growth factors are potentially maintained at lower levels, Notch signaling suppresses mRNA expression of hexokinase 2 (hk2), which is a glycolysis-associated gene, in the central nervous system. However, whether and how the genetic regulation of HK2 via Notch signaling contributes to cellular adaptability to nutrient-poor environments remains unknown. In this study, we performed gene expression analysis using a U87-MG human glioma cell line and revealed that under conditions where both glutamine and serum were absent, Notch signaling was activated and HK2 expression was downregulated by Notch signaling. We also found that Notch-mediated HK2 suppression was triggered in a Notch ligand-selective manner. Furthermore, HK2 was shown to inhibit cell proliferation of U87-MG gliomas, which might depend on Notch signaling activity. Together, our findings suggest the involvement of Notch-mediated HK2 suppression in an adaptive mechanism of U87-MG glioma cells to nutrient-poor conditions.

In eukaryotes, maintenance of heterochromatin structure that represses gene expression during cell proliferation is essential for guaranteeing cell identity. However, how heterochromatin is maintained and transmitted to the daughter cells remains elusive. In this study, we constructed a reporter system to study the maintenance of heterochromatin in the subtelomeric region of the fission yeast, Schizosaccharomyces pombe. We demonstrated that once subtelomeric heterochromatin was established, it tended to be maintained as a metastable structure through cell proliferation. Using this system, we screened an S. pombe genome-wide gene deletion library for subtelomeric heterochromatin maintenance factors and identified 57 genes related to various cellular processes, in addition to well-characterized heterochromatin factors. We focused on Mrc1, a mediator of DNA replication checkpoint. We found that Mrc1 maintains heterochromatin structure not only at the subtelomeres but also at the pericentromeres and mating-type regions. Furthermore, we showed that Mrc1 is required for the localization of Snf2/Hdac-containing Repressor Complex (SHREC) and the maintenance of hypoacetylation state of histone H3K14. This study complements the recent discoveries that Mrc1 functions as a histone H3-H4 chaperone in heterochromatin maintenance.

Pathogenic microorganisms often target seedlings shortly after germination. If plants exhibit resistance or resilience to pathogens, those exposed to pathogen challenge may grow further and form new unchallenged leaves. The purpose of this study was to examine disease resistance in the newly formed leaves of plants subjected to pathogen challenge. We used Arabidopsis thaliana and the oomycete pathogen Hyaloperonospora arabidopsidis (Hpa) as the model pathosystem. We found that Arabidopsis seedlings primarily challenged with the avirulent isolate Hpa exhibited enhanced disease resistance against the virulent isolate Hpa in newly formed rosette leaves (NFRLs). Our observations indicated that the transcript levels of the transcription factor gene WRKY70, which is essential for full resistance to the virulent isolate HpaNoco2, were elevated and maintained at high levels in the NFRLs. In contrast, the transcript levels of the salicylic acid marker gene PR1 and systemic acquired resistance-related genes did not exhibit sustained elevation. The maintenance of increased transcript levels of WRKY70 operated independently of non-expressor of pathogenesis-related gene 1. These findings suggest that prolonged upregulation of WRKY70 represents a defensive state synchronized with plant development to ensure survival against subsequent infections.

Use of immune checkpoint inhibitors (ICIs) as cancer immunotherapy has advanced rapidly in the clinic; however, ICI initiation can also cause an unexpectedly rapid acceleration of cancer progression in some patients. Here, we used a murine syngeneic melanoma model to conduct mechanistic analysis of cancer-associated fibroblast (CAF) function in cancer progression in the context of immunotherapy. We found that after ICI treatment CAFs acquire inflammatory properties, which can promote tumor progression. Mechanistically, we show that T-cell-derived interferon-γ (IFN-γ) stimulates production of tumor necrosis factor-α (TNF-α) by macrophages, facilitating CAF conversion to inflammatory CAFs. Our findings suggest that CAF/immune cell crosstalk plays an essential role in ICI-associated tumor progression.

The third Asia Pacific Drosophila Neurobiology Conference (APDNC3) was held in the Wako Campus of RIKEN in Tokyo, Japan, from February 27th to March 1st, 2024. While APDNC2 was held in Taiwan in 2019, the global coronavirus pandemic enforced a long hiatus. Hence, APDNC3 was a much-anticipated meeting that attracted ~218 scientists from 18 different countries and regions, 154 from outside Japan. The meeting was divided into 13 scientific, 2 poster, and 3 career development sessions. Two plenary talks were delivered by Professor Daisuke Yamamoto, from NICT and Professor Claude Desplan from NYU. Thirty-seven other speakers were invited to give lectures. Eighty-six poster presenters were selected from submitted abstracts. Talks and posters described how neuronal circuits underlying specific behaviors were identified and how they developed. The presented work also demonstrated circuit-specific cellular and molecular mechanisms in health and disease. It was clear that technological advances, like molecular genetic tools for identifying, manipulating, and imaging individual neurons and the great granularity of the fly brain connectome, were significantly augmenting research. Overall, the meeting highlighted the remarkable biological insights that fly neurobiologists continue to provide.

Completion of DNA replication before chromosome segregation is essential for the stable maintenance of the genome. Under replication stress, DNA synthesis may persist beyond S phase, especially in genomic regions that are difficult to proceed with the replication processes. Incomplete replication in mitosis emerges as non-disjoined segment in mitotic chromosomes leading to anaphase bridges. The resulting chromosome rearrangements are not well characterized, however. Here, we report that incomplete replication due to SMC5/6 deficiency impairs sister chromatid disjunction at difficult-to-replicate regions, including common fragile sites. These non-disjoined regions manifest as cytologically defined symmetric gaps, causing anaphase bridges. These bridges break at the gaps, leading to telomere loss, micronucleation, and fragmentation. Subsequently, fusions between telomere-deficient chromosomes generate complex chromosomal rearrangements, including dicentric chromosomes, suggesting the occurrence of breakage-fusion-bridge cycle. Additionally, chromosomes in micronuclei were pulverized, indicative of chromothripsis. Our findings suggest that incomplete replication facilitates complex chromosomal rearrangements, which may contribute to genomic instability in human cancers.

Tardigrades possess the ability to enter an almost completely dehydrated state, anhydrobiosis. The CAHS (cytosolic abundant heat-soluble) protein family has been identified as one of the anhydrobiosis-related proteins. In particular, CAHS3 protein from an anhydrobiotic tardigrade, Ramazzottius varieornatus, shows heat-solubility and reversible condensation and is one of the most highly expressed among the CAHS paralogs. A recently developed tardigrade-specific vector showed tissue-specific expression of RvCAHS3 most pronounced in the epidermis in vivo, contrary to the idea that anhydrobiotic genes are uniformly expressed in all tardigrade cells. In this study, we investigated the regulation of RvCAHS3 gene expression through in vivo expression experiments using tardigrade vectors with a series of truncated upstream regions coupled with in silico analysis to identify the anhydrobiosis-related genes that are expressed under the same regulatory system as RvCAHS3. As a result, the 300-350 bp region upstream of RvCAHS3 is critical for regulating gene expression in tardigrade vector experiments, and three motifs conserved between two species of anhydrobiotic tardigrades were identified within a 500 bp region directly upstream of RvCAHS3 start codon. These motifs, which have also been identified upstream of other CAHS genes, could be associated with the regulatory system of anhydrobiosis-related genes in tardigrades.

Acute liver failure is a serious, life-threatening disease. Although the gut microbiota has been considered to play a role in liver failure, the extent to which it is involved in the pathogenesis of this disease has not been fully elucidated to date. Therefore, we here analyzed the importance of the presence of intestinal microbiota in the pathogenesis of acute liver injury, using D-galactosamine (D-GalN)/lipopolysaccharide (LPS)-treated mice, which is a widely used experimental model of acute liver injury. First, administration of the antibiotic polymyxin B markedly alleviated liver injury. Liver injury was also reduced in germ-free mice, leading to the conclusion that the presence of intestinal microbiota aggravates D-GalN/LPS-induced liver injury. The amount of bacteria and LPS transferred from the gut to the blood was not increased by D-GalN/LPS, suggesting that the worsening of liver injury was not simply owing to the entry of bacteria into the circulation. In conclusion, acute liver injury in polymyxin B-pretreated or germ-free mice was ameliorated by modulation of the gut microbiota. Modification of the gut microbiota using polymyxin B may hence have the potential to alleviate acute liver injury in human patients.

Transcription factor GATA3 is essential for the developmental processes of T cells. Recently, the silencer of a cytokine IFNγ gene was identified, the inhibitory activity of which requires GATA3. GATA3 has 2 Zn fingers and the commonly used GATA3 deficient mice lack both fingers (D2). We have established a mouse line that lacks only one Zn finger close to the C terminus (D1). The D1 mice line developed dermatitis, which was not observed in D2 mice. The expression of S100a8/S100a9 was elevated in D1 to a level higher than in D2, suggesting their roles in dermatitis development. CD8 T cells of both D1 and D2 lines expressed inhibitory receptors associated with the exhausted state. In the absence of MHC class II, the skin inflammation was exacerbated in both lines. The gene expression pattern of CD8 T cells became similar to that of effector T cells. Blocking Ab against LAG3 upregulated the expression of the effector molecules of T cells. These results suggest that the disfunction of GATA3 can lead to the spontaneous activation of CD8 T cells that causes skin inflammation, and that suppressive activity of MHC class II - LAG3 interaction ameliorates dermatitis development.

A single epithelial cell embedded in extracellular matrix (ECM) can proliferate to form an apical lumen-harboring cyst, whose formation is a fundamental step in epithelial organ development. At an early two-cell stage after cell division, the cell doublet typically displays "inverted" polarity, with apical and basolateral proteins being located to the ECM-facing and cell-cell-contacting plasma membranes, respectively. Correct cystogenesis requires polarity reorientation, a process containing apical protein endocytosis from the ECM-abutting periphery and subsequent apical vesicle delivery to a cell-cell contact site for lumen formation. Here, we show that downstream of the ECM-signal-transducer β1-integrin, Rac1, and its effector IQGAP1 promote apical protein endocytosis, contributing to polarity reorientation of mammalian epithelial Madin-Darby canine kidney (MDCK) cells at a later two-cell stage in three-dimensional culture. Rac1-GTP facilitates IQGAP1 interaction with the Rac-specific activator Tiam1, which also contributes to the endocytosis and enhances the effect of IQGAP1. These findings suggest that Tiam1 and IQGAP1 form a positive feedback loop to activate Rac1. With Rac1-GTP, IQGAP1 also binds to AP2α, an adaptor protein subunit for clathrin-mediated endocytosis; depletion of the AP2 complex impairs apical protein endocytosis in MDCK doublets. Thus, Rac1 likely participates in polarity reorientation at the two-cell stage via its interaction with IQGAP1.

Zinc finger E-box binding homeobox 1 (ZEB1) has been identified as a key factor in cancer cell differentiation and metastasis, and has been well studied in the field of cancer cell biology. ZEB2 has a highly similar conformation to ZEB1, but its role in head and neck squamous cell carcinoma (HNSCC) cells is not fully understood. Here, we separately overexpressed ZEB1 and ZEB2 in C57BL/6 mouse oral cancer (MOC) cells and investigated their cellular characteristics, including E-cadherin levels, motile properties, chemoresistance, and metastatic ability in immunocompetent mice. Both ZEB1 and ZEB2 overexpression reduced epithelial traits and converted cells to an aggressive phenotype. Surprisingly, ZEB1 overexpression increased the endogenous level of ZEB2 in MOC cells, and vice versa. The molecular mechanisms underlying these findings remain unclear. However, the in vitro anchorage-independent growth of MOC cells overexpressing ZEB2 was considerably greater than that of MOC cells overexpressing ZEB1. These findings suggest that ZEB2, like ZEB1, has the ability to induce the differentiation of cancer cells into those with highly aggressive traits.

The asymmetric cell division determines cell diversity and distinct sibling cell fates by mechanisms linked to mitosis. Many adult stem cells divide asymmetrically to balance self-renewal and differentiation. The process of asymmetric cell division involves an axis of polarity and, second, the localization of cell fate determinants at the cell poles. Asymmetric division of stem cells is achieved by intrinsic and extrinsic fate determinants such as signaling molecules, epigenetics factors, molecules regulating gene expression, and polarized organelles. At least some stem cells perform asymmetric and symmetric cell divisions during development. Asymmetric division ensures that the number of stem cells remains constant throughout life. The asymmetric division of stem cells plays an important role in biological events such as embryogenesis, tissue regeneration and carcinogenesis. This review summarizes recent advances in the regulation of asymmetric stem cell division in model organisms.

The mammalian p57 protein is a member of the CIP/KIP family of cyclin-dependent kinase inhibitors and plays an essential role in the development of multiple tissues during embryogenesis as well as in the maintenance of tissue stem cells in adults. Although several transcription factors have been implicated in regulating the p57 gene, cis-elements such as enhancers that regulate its expression have remained ill-defined. Here we identify a candidate enhancer for the mouse p57 gene (Cdkn1c) within an intron of the Kcnq1 locus by 4C-seq analysis in mouse embryonic stem cells (mESCs). Deletion of this putative enhancer region with the CRISPR-Cas9 system or its suppression by CRISPR interference resulted in a marked attenuation of Cdkn1c expression in differentiating mESCs. Our results thus suggest that this region may serve as an enhancer for the p57 gene during early mouse embryogenesis.

Alport syndrome (AS) is a hereditary disease caused by mutations in the COL4A5 gene and leads to chronic kidney disease. Currently, no specific treatment has been developed. However, a recent study using AS-model mice demonstrated that the exon skipping method could partially rescue the symptoms. In this study, we evaluated the effects of the exon skipping method using kidney organoids generated from AS-patient-derived induced pluripotent stem cells (AS-iPSCs). We generated kidney organoids from AS-iPSCs, which exhibited nephron structures. As expected, the C-terminus of COL4A5 was not expressed in AS-organoids. Interestingly, anti-sense oligonucleotides restored the expression of the C-terminus of COL4A5 in vitro. Next, we transplanted AS-organoids into mice and evaluated glomerular basement membrane formation in vivo. We found that AS-organoids formed a lower slit diaphragm ratio compared to control organoids. Finally, we assessed the effects of exon skipping on transplanted organoids but observed minimum effects. These studies suggest that AS-iPSCs can generate kidney organoids lacking the C-terminus of COL4A5, and that exon skipping can induce its expression in vitro.

Regular exercise is believed to suppress cancer progression. However, the precise molecular mechanisms by which exercise prevents cancer development remain unclear. In this study, using a steatosis-associated liver cancer mouse model, we found that regular exercise at a speed of 18 m/min for 20 min daily suppressed liver cancer development. To explore the underlying mechanisms, we examined the gene expression profiles in the livers of the exercise and non-exercise groups. The expressions of circadian genes, such as Per1 and Cry2, were upregulated in the exercise group. As circadian rhythm disruption is known to cause various diseases, including cancer, improving circadian rhythm through exercise could contribute to cancer prevention. We further found that the expression of a series of E2F1 and c-Myc target genes that directly affect the proliferation of cancer cells was downregulated in the exercise group. However, the expression of E2F1 and c-Myc was transcriptionally unchanged but degraded at the post-translational level by exercise. Cry2, which is regulated by the Skp1-Cul1-FBXL3 (SCF) ubiquitin ligase complex by binding to FBXL3, can form a complex with E2F1 and c-Myc, which we think is the mechanism to degrade them. Our study revealed a previously unknown mechanism by which exercise prevents cancer development.

The cell cycle is driven by cyclin-dependent kinases (Cdks). The decision whether the cell cycle proceeds is made during G1 phase, when Cdk4/6 functions. Cyclin-dependent kinase inhibitor 2 (Cdkn2) is a specific inhibitor of Cdk4/6, and their interaction depends on D84 in Cdkn2 and R24/31 in Cdk4/6. This knowledge is based mainly on studies in mammalian cells. Here, we comprehensively analyzed Cdk4/6 and Cdkn2 in invertebrates and found that Cdk4/6 was present in most of the investigated phyla, but the distribution of Cdkn2 was rather uneven among and within the phyla. The positive charge of R24/R31 in Cdk4/6 was conserved in all analyzed species in phyla with Cdkn2. The presence of Cdkn2 and the conservation of the positive charge were statistically correlated. We also found that Cdkn2 has been tightly linked to Fas associated factor 1 (Faf1) during evolution. We discuss potential interactions between Cdkn2 and Cdk4/6 in evolution and the possible cause of the strong conservation of the microsynteny.

Missing an entire chromosome or chromosome arm in normal diploid cells has a deleterious impact on cell viability, which may contribute to the development of specific birth defects. Nevertheless, the effects of chromosome loss in human cells have remained unexplored due to the lack of suitable model systems. Here, we developed an efficient, selection-free approach to generate partial monosomy in human induced pluripotent stem cells (iPSCs). The introduction of Cas9 proteins and a pair of gRNAs induces over megabase-sized interstitial chromosomal deletions. Using human chromosome 21 (HSA21) as a model, partial monosomy 21q (PM21q) iPSC lines with deletions ranging from 4.5 to 27.9 Mb were isolated. A 33.6 Mb deletion, encompassing all protein-coding genes on 21q, was also achieved, establishing the first 21q monosomy human iPSC line. Transcriptome and proteome analyses revealed that the abundances of mRNA and protein encoded by the majority of genes in the monosomic regions are half of the diploid expression level, indicating an absence of dosage compensation. The ability to generate customized partial monosomy cell lines on an isogenic, karyotypically normal background should facilitate the gain of novel insights into the impact of chromosome loss on cellular fitness.

Protogyny, being capable of changing from female to male during their lifetime, is prevalent in 20 families of teleosts but is believed to have evolved within specific evolutionary lineages. Therefore, shared regulatory factors governing the sex change process are expected to be conserved across protogynous fishes. However, a comprehensive understanding of this mechanism remains elusive. To identify these factors, we conducted a meta-analysis using gonadal transcriptome data from seven species. We curated data pairs of ovarian tissue and transitional gonad, and employed ratios of expression level as a unified criterion for differential expression, enabling a meta-analysis across species. Our approach revealed that classical sex change-related genes exhibited differential expression levels between the ovary and transitional gonads, consistent with previous reports. These results validate our methodology's robustness. Additionally, we identified novel genes not previously linked to gonadal sex change in fish. Notably, changes in the expression levels of acetoacetyl-CoA synthetase and apolipoprotein Eb, which are involved in cholesterol synthesis and transport, respectively, suggest that the levels of cholesterol, a precursor of steroid hormones crucial for sex change, are decreased upon sex change onset in the gonads. This implies a potential universal influence of cholesterol dynamics on gonadal transformation in protogyny.

Schizophyllum commune, a common wood-decay mushroom known for its extremely high genetic variation and as a rare cause of human respiratory diseases, could be a promising model fungus contributing to both biology and medicine. To better understand its phenotypic variation, we developed an image analysis system that quantifies morphological and physiological traits of mycelial colonies in Petri dishes. This study evaluated growth of six wild and one clinical isolates of Japanese S. commune, subjected to different temperatures and glucose concentrations, including a condition mimicking the human respiratory environment. Our analysis revealed that combinations of two growth indices, area and whiteness, and profiling by clustering algorithms highlighted strain-specific responses. For example, the clinical isolate was the whitest under the respiratory-like condition. We also found that the growth rate was strongly determined by glucose concentration, while the effects of temperature on growth varied among the strains, suggesting that while glucose preference is common in this species, responses to temperature differ between strains. This system showed sufficient sensitivity to detect variation in mycelial growth. Our study provides a key to unraveling morphological and physiological traits behind the high polymorphisms in S. commune, including the ability to colonize the human respiratory tract.