Biology subjects

Imada, S.

Publications and source records attributed to Imada, S..

7 recordsLinked to original sources

bioRxiv2025

The Wnt/APC destruction complex targets SREBP2 in a β-catenin-independent pathway to control cholesterol metabolism

APC, the core scaffold of the Wnt destruction complex, targets the transcriptional co-activator {beta}-catenin for proteolysis. There is no convincing evidence that APC directs degradation of other substrates. Using a reconstituted cytosolic extract-based system and complementary in vivo and cellular assays, we show that SREBP2, the master regulator of cholesterol biosynthesis, is a direct APC-AXIN1 substrate. APC-dependent SREBP2 degradation is conserved in Xenopus embryos, mouse colon, and human colorectal cancer cells and restricts SREBP2 target-gene expression, cholesterol synthesis, and tissue cholesterol levels. Mechanistically, APC and AXIN1 promote SREBP2 degradation via a conserved phosphodegron, which marks SREBP2 for ubiquitination by the E3 enzyme, FBXW7. Like {beta}-catenin, SREBP2 is stabilized by extracellular Wnt ligands; unlike {beta}-catenin, its regulation is independent of GSK3{beta} and CK1 and requires the entire APC mutational cluster region (MCR), whereas {beta}-catenin turnover can operate with only a partial MCR. These findings define a {beta}-catenin-independent branch of Wnt signaling that couples APC to sterol metabolism, providing a mechanistic rationale to target the mevalonate/SREBP2 axis in APC-mutant colorectal cancer.

biochemistry↗

bioRxiv2025

Polyamines buffer labile iron to suppress ferroptosis

Polyamines are essential and evolutionarily conserved metabolites present at millimolar concentrations in mammalian cells. Cells tightly regulate polyamine homeostasis through complex feedback mechanisms, yet the precise role necessitating this regulation remains unclear. Here, we show that polyamines contribute to endogenous buffering of redox-active iron, providing a molecular link between polyamine metabolism and ferroptosis. Using a genome-wide CRISPR screen, we identified a synthetic lethal dependency between polyamine depletion and the key ferroptosis suppressor, GPX4. Mechanistically, we show that polyamine deficiency triggers a redistribution of cellular iron, increasing the labile iron pool and upregulating ferritin. To directly visualize this iron buffering in living cells, we developed a genetically encoded fluorescent reporter for redox-active iron. Live-cell analysis revealed a striking inverse correlation between intracellular polyamine levels and redox-active iron at single-cell resolution. These findings reposition polyamines as key regulators of iron homeostasis, with implications for ferroptosis-linked disease states and cellular redox balance.

cell biology↗

bioRxiv2024

A genetically encoded fluorescent reporter for polyamines

Polyamines are abundant and evolutionarily conserved metabolites that are essential for life. Dietary polyamine supplementation extends life-span and health-span. Dysregulation of polyamine homeostasis is linked to Parkinsons disease and cancer, driving interest in therapeutically targeting this pathway. However, measuring cellular polyamine levels, which vary across cell types and states, remains challenging. We introduce a first-in-class genetically encoded polyamine reporter for real-time measurement of polyamine concentrations in single living cells. This reporter utilizes the polyamine-responsive ribosomal frameshift motif from the OAZ1 gene. We demonstrate broad applicability of this approach and reveal dynamic changes in polyamine levels in response to genetic and pharmacological perturbations. Using this reporter, we conducted a genome-wide CRISPR screen and uncovered an unexpected link between mitochondrial respiration and polyamine import, which are both risk factors for Parkinsons disease. By offering a new lens to examine polyamine biology, this reporter may advance our understanding of these ubiquitous metabolites and accelerate therapy development.

cell biology↗

bioRxiv2024

Polyamines sustain epithelial regeneration in aged intestines by modulating protein homeostasis

Aging hampers the regenerative potential of intestinal epithelium across species including humans, yet the underlying causes remain elusive. Here, using proteomic and metabolomic profiling of intestinal tissues together with functional assays, we characterized the temporal dynamics of regeneration following injury induced by 5-fluorouracil, a commonly used chemotherapeutic agent. Comparison of regeneration dynamics in mice of different ages revealed the emergence of a proteostasis stress signature and increased levels of polyamines following injury exclusively in old epithelia. Mechanistically, we show that delayed regeneration is an intrinsic feature of aged epithelial cells that display reduced protein synthesis and accumulation of ubiquitylated proteins. Notably, dietary restriction followed by re-feeding prior to injury increases polyamine pathway activation, enhances protein synthesis, and restores the regenerative capacity of aged intestines. Our findings highlight promising epithelial targets for interventions aimed at tackling the decline in tissue repair mechanisms associated with aging.

molecular biology↗

bioRxiv2022

Integrative single cell and spatial transcriptomics of colorectal cancer reveals multicellular functional units that support tumor progression

While advances in single cell genomics have helped to chart the cellular components of tumor ecosystems, it has been more challenging to characterize their specific spatial organization and functional interactions. Here, we combine single cell RNA-seq, spatial transcriptomics by Slide-seq, and in situ multiplex RNA analysis, to create a detailed spatial map of healthy and dysplastic colon cellular ecosystems and their association with disease progression. We profiled inducible genetic CRC mouse models that recapitulate key features of human CRC, assigned cell types and epithelial expression programs to spatial tissue locations in tumors, and computationally used them to identify the regional features spanning different cells in the same spatial niche. We find that tumors were organized in cellular neighborhoods, each with a distinct composition of cell subtypes, expression programs, and local cellular interactions. Comparing to scRNA-seq and Slide-seq data from human CRC, we find that both cell composition and layout features were conserved between the species, with mouse neighborhoods correlating with malignancy and clinical outcome in human patient tumors, highlighting the relevance of our findings to human disease. Our work offers a comprehensive framework that is applicable across various tissues, tumors, and disease conditions, with tools for the extrapolation of findings from experimental mouse models to human diseases.

cancer biology↗

bioRxiv2022

Lymphatics constitute a novel component of the intestinal stem cell niche

Intestinal stem cells (ISCs) depend on niche factors for their proper function. However, the source of these ISC niche factors and how they support ISCs remain controversial due to their redundant expression patterns. Here, we report that the maintenance of ISCs depends on both lymphatic endothelial cells (LECs) and Rspo3+Grem1+ fibroblasts (RGFs). We found that LECs are surrounded by RGFs and located in close proximity to Lgr5+ ISCs. RSPO3 production is restricted to LECs and RGFs and they can partially compensate for each other; however, RSPO3 loss in both of LECs and RGFs drastically compromises ISC numbers, villi length, and repair after irradiation-induced injury. Mechanistically, irradiation-induced damage expands LEC and RGF numbers and enhances the latters generation of RSPO3 through IL-1 receptor activation. We propose that LECs represent a novel component of the ISC niche, which together with RGFs, provide essential RSPO3 to sustain ISCs in homeostasis and regeneration.

cell biology↗

bioRxiv2021

Stiffness Regulates Intestinal Stem Cell Fate

Does fibrotic gut stiffening caused by inflammatory bowel diseases (IBD) direct the fate of intestinal stem cells (ISCs)? To address this question we first developed a novel long-term culture of quasi-3D gut organoids plated on hydrogel matrix of varying stiffness. Stiffening from 0.6kPa to 9.6kPa significantly reduces Lgr5high ISCs and Ki67+ progenitor cells while promoting their differentiation towards goblet cells. These stiffness-driven events are attributable to YAP nuclear translocation. Matrix stiffening also extends the expression of the stemness marker Olfactomedin 4 (Olfm4) into villus-like regions, mediated by cytoplasmic YAP. We next used single-cell RNA sequencing to generate for the first time the stiffness-regulated transcriptional signatures of ISCs and their differentiated counterparts. These signatures confirm the impact of stiffening on ISC fate and additionally suggest a stiffening-induced switch in metabolic phenotype, from oxidative phosphorylation to glycolysis. Finally, we used colon samples from IBD patients as well as chronic colitis murine models to confirm the in vivo stiffening-induced epithelial deterioration similar to that observed in vitro. Together, these results demonstrate stiffness-dependent ISC reprograming wherein YAP nuclear translocation diminishes ISCs and Ki67+ progenitors and drives their differentiation towards goblet cells, suggesting stiffening as potential target to mitigate gut epithelial deterioration during IBD.

cell biology↗