Biology subjects

Naguro, I.

Publications and source records attributed to Naguro, I..

8 recordsLinked to original sources

bioRxiv2023

Contact-Inhibited ERK Signaling is Determined by Cellular-Resolution Western Blotting

Extracellular signal-regulated kinase (ERK) signaling is essential to regulated cell behaviors, including cell proliferation, differentiation, and apoptosis. The influence of cell-cell contacts on ERK signaling is central to epithelial cells, yet few studies have sought to understand the same in cancer cells, particularly with single-cell resolution. To acquire same-cell measurements of both phenotypic (cell-contact state) and targeted-protein profile (ERK phosphorylation), we prepend high-content, whole-cell imaging prior to endpoint cellular-resolution western blot analyses for each of hundreds of individual HeLa cancer cells cultured on that same chip, which we call contactBlot. By indexing the phosphorylation level of ERK in each cell or cell-cluster to the imaged cell-contact state, we compare ERK signaling between isolated and in-contact cells. We observe attenuated ([~]2x) ERK signaling in HeLa cells which are in-contact versus isolated. Attenuation is sustained when the HeLa cells are challenged with hyperosmotic stress. Our findings show the impact of cell-cell contacts on ERK activation with isolated and in-contact cells, while introducing a multi omics tool for control and scrutiny of cell-cell interactions.

bioengineering↗

bioRxiv2023

Zinc controls histone acetyltransferase KAT7 activity to maintain cellular zinc homeostasis

Zinc is an indispensable micronutrient for optimal physiological functions, and zinc deficiency has been implicated in the pathogenesis of various human diseases. One potential mechanism underlying such pathogenic effects is the alteration of gene expression caused by zinc deficiency; however, the details of this process remain largely unexplored. Here, we show that during zinc deficiency, the histone acetyltransferase KAT7 loses its enzymatic activity, leading to the attenuated acetylation of histone H3 at Lys14 (H3K14ac) at enhancer regions. Physiologically, the decrease in H3K14ac leads to the upregulation of the expression of ZIP10, a plasma membrane-localized zinc transporter, thereby facilitating the import of extracellular zinc to maintain cellular zinc homeostasis. Moreover, prolonged zinc deficiency in mice induced by a zinc-deficient diet or high-fat diet, accompanied by decreased H3K14ac levels in the liver, upregulated the expression of genes associated with intracellular lipid droplet formation, leading to the accumulation of lipids within liver tissue. Our findings demonstrate that cells respond to zinc deficiency by converting it into an epigenetic signal that drives physiological or pathophysiological biological processes.

cell biology↗

bioRxiv2023

Proteasomal regulation of ASK family kinases dictates cell fate under hyperosmotic stress

The proteasome has an essential role in proteostasis maintenance and is critical for cell survival under proteotoxic conditions including hyperosmotic stress. However, it is unknown how proteasome activity is linked to cell survival/death under hyperosmotic stress. We have previously reported that apoptosis signal-regulating kinase 3 (ASK3) contributes to cell survival through its inactivation under hyperosmotic stress. 19S regulatory particle subunits of the proteasome were enriched in the ASK3 inactivator candidates identified through our genome-wide small interfering RNA (siRNA) screen. In this study, we demonstrate that the proteasome regulates ASK3 inactivation through its proteolytic activity. Intriguingly, the proteasome inactivates ASK3 via degradation of not ASK3 per se but another ASK family member ASK1 which activates ASK3 in a kinase activity-dependent manner. Furthermore, the elevated ASK1 level under proteasome inhibition sensitizes cells to hyperosmotic stress. These findings suggest that ASK family maintenance links proteasome capacity to cell fate bifurcation under hyperosmotic stress.

molecular biology↗

bioRxiv2022

HES1 contributes to high salt stress response as an enhancer of NFAT5-DNA binding

High salt conditions and subsequent hyperosmolarity are injurious cellular stresses but can activate immune signaling. Nuclear factor of activated T-cells 5 (NFAT5) is an essential transcription factor that induces osmoprotective genes such as aldose reductase (AR) and betaine-GABA transporter 1 (BGT1). High salt stress-mediated NFAT5 activation is also reported to accelerate the inflammatory response and autoimmune diseases. However, the systemic regulation of NFAT5 remains unclear. Here, we performed a genome-wide siRNA screen to comprehensively identify the upstream factors of NFAT5. We monitored NFAT5 nuclear translocation and identified one of the Notch signaling effectors, Hairy and enhancer of split-1 (HES1), as a novel positive regulator of NFAT5. HES1 was induced by high salinity via ERK signaling and facilitated NFAT5 recruitment to its target promoter region, resulting in the proper induction of osmoprotective genes and cytoprotection under high salt stress. These findings suggest that although HES1 is well known as a transcriptional repressor, it positively regulates NFAT5-dependent transcription in the context of a high salinity/hyperosmotic response. One Sentence SummaryHES1 contributes to high salinity/hyperosmotic response through positive regulation of NFAT5-dependent transcription.

cell biology↗

bioRxiv2022

DNA damage response signaling to mitochondria drives senescence

Cellular senescence is a stress-induced irreversible cell cycle arrest typically accompanied by expression of the cyclin-dependent kinase inhibitor p16INK4a (hereafter referred to as p16) and mitochondrial dysfunction1. Recent studies have indicated that p16-expressing senescent cells accumulate in the body over time and contribute to aging1, 2. Many stresses, such as telomere shortening and oncogene activation, induce senescence by damaging nuclear DNA1. However, the molecular mechanisms linking DNA damage to senescence remain unclear. Here, we show that the outer mitochondrial transmembrane protein BNIP3 drives senescence by triggering a DNA damage response (DDR) of mitochondria. BNIP3 was identified in a genome-wide siRNA screen for genes required for p16 expression upon DNA damage. Mass spectrometric analysis of BNIP3-interacting proteins yielded the DDR kinase ATM and subunits of the mitochondrial contact site and cristae organizing system (MICOS) complex. BNIP3 is an ATM substrate that increases the number of mitochondrial cristae upon DNA damage. This increase enhances the oxidation of fatty acids to acetyl-CoA, an acetyl group donor, thereby promoting histone acetylation and associated p16 expression. Our findings indicate that DDR signaling to mitochondria promotes p16 expression by altering mitochondrial structure and metabolism and highlight the importance of nuclear-mitochondrial communication in senescence induction.

cell biology↗

bioRxiv2022

Mitochondria directly sense osmotic stress to trigger rapid metabolic remodeling via PDH regulation

A high-salt diet significantly impacts various diseases, including cancer and immune diseases. Recent studies suggest that the high-salt/hyperosmotic environment in the body may alter the chronic properties of cancer and immune cells in the disease context. However, little is known about the acute metabolic changes in hyperosmotic stress. Here, we found that hyperosmotic stress for a few minutes induces Warburg-like metabolic remodeling in HeLa and Raw264.7 cells and suppresses fatty acid oxidation. Regarding Warburg-like remodeling, the PDH phosphorylation status was altered bidirectionally (high in hyperosmolarity and low in hypoosmolarity) to osmotic stress in isolated mitochondria, suggesting that mitochondria themselves have an acute osmo-sensing mechanism. Warburg-like remodeling is required for HeLa cells to maintain ATP levels and survive under hyperosmotic conditions. Our findings suggest that cells exhibit acute metabolic remodeling under osmotic stress via the regulation of PDH phosphorylation by direct osmosensing within mitochondria.

cell biology↗

bioRxiv2022

Sodium ion regulates liquidity of biomolecular condensates in hyperosmotic stress response

Biomolecular condensates are membraneless structures formed through phase separation. Recent studies have demonstrated that the material properties of biomolecular condensates are crucial for their biological functions and pathogenicity. However, the phase maintenance of biomolecular condensates in cells remains elusive. Here, we show that sodium ion (Na+) influx regulates the condensate liquidity under hyperosmotic stress. The fluidity of ASK3 condensates increases at the high intracellular Na+ concentration derived from extracellular hyperosmotic solution. Moreover, we identified TRPM4 as a cation channel that allows Na+ influx under hyperosmotic stress. TRPM4 inhibition causes the liquid-to-solid phase transition of ASK3 condensates, leading to impairment of the ASK3 osmoresponse. In addition to ASK3 condensates, intracellular Na+ widely regulates the condensate liquidity and aggregate formation of biomolecules, including DCP1A, TAZ and polyQ-protein, under hyperosmotic stress. Our findings demonstrate that changes in Na+ contribute to the cellular stress response via liquidity maintenance of biomolecular condensates.

cell biology↗

bioRxiv2020

Cells recognize osmotic stress through liquid-liquid phase separation lubricated with poly(ADP-ribose)

Cells are under threat of osmotic perturbation; and cell volume maintenance is critical in cerebral edema, inflammation and aging, in which prominent changes in intracellular or extracellular osmolality emerge. After osmotic stress-enforced cell swelling or shrinkage, the cells regulate intracellular osmolality to recover their volume. However, the mechanisms recognizing osmotic stress remain obscured. We previously clarified that apoptosis signal-regulating kinase 3 (ASK3) bidirectionally responds to osmotic stress and regulates cell volume recovery. Here, we report that macromolecular crowding induces liquid-demixing condensates of ASK3 under hyperosmotic stress, which transduce osmosensing signal into ASK3 inactivation. A genome-wide small interfering RNA (siRNA) screen identified an ASK3 inactivation regulator, nicotinamide phosphoribosyltransferase (NAMPT), related to poly(ADP-ribose) signaling. Furthermore, we clarify that poly(ADP-ribose) keeps ASK3 condensates in the liquid phase and enables ASK3 to become inactivated under hyperosmotic stress. Our findings demonstrate that cells rationally incorporate physicochemical phase separation into their osmosensing systems.

cell biology↗