Nutrient Starvation Research Articles

Impact of context-dependent autophagy states on tumor progression.

Macroautophagy is a cellular quality-control process that degrades proteins, protein aggregates and damaged organelles. Autophagy plays a fundamental role in cancer where, in the presence of stressors (for example, nutrient starvation, hypoxia, mechanical pressure), tumor cells activate it to degrade intracellular substrates and provide energy. Cell-autonomous autophagy in tumor cells and cell-nonautonomous autophagy in the tumor microenvironment and in the host converge on mechanisms that modulate metabolic fitness, DNA integrity and immune escape and, consequently, support tumor growth. In this Review, we will discuss insights into the tumor-modulating roles of autophagy in different contexts and reflect on how future studies using physiological culture systems may help to understand the complexity and open new therapeutic avenues.

The impact of pericytes on the stability of microvascular networks in response to nanoparticles

Recapitulating the normal physiology of the microvasculature is pivotal in the development of more complex in-vitro models and organ-on-chip designs. Pericytes are an important component of the vasculature, promoting vessel stability, inhibiting vascular permeability and maintaining the vascular hierarchical architecture. The use of such co-culture for the testing of therapeutics and nanoparticle safety is increasingly considered for the validation of therapeutic strategies. This report presents the use of a microfluidic model for such applications. Interactions between endothelial cells and pericytes are first explored. We identify basal conditions required to form stable and reproducible endothelial networks. We then investigate interactions between endothelial cells and pericytes via direct co-culture. In our system, pericytes prevented vessel hyperplasia and maintained vessel length in prolonged culture (> 10 days). In addition, these vessels displayed barrier function and expression of junction markers associated with vessel maturation, including VE-cadherin, β-catenin and ZO-1. Furthermore, pericytes maintained vessel integrity following stress (nutrient starvation) and prevented vessel regression, in contrast to the striking dissociation of networks in endothelial monocultures. This response was also observed when endothelial/pericyte co-cultures were exposed to high concentrations of moderately toxic cationic nanoparticles used for gene delivery. This study highlights the importance of pericytes in protecting vascular networks from stress and external agents and their importance to the design of advanced in-vitro models, including for the testing of nanotoxicity, to better recapitulate physiological response and avoid false positives.

ATG8-dependent LMX1B-autophagy crosstalk shapes human midbrain dopaminergic neuronal resilience.

The LIM homeodomain transcription factors LMX1A and LMX1B are essential mediators of midbrain dopaminergic neuronal (mDAN) differentiation and survival. Here we show that LMX1A and LMX1B are autophagy transcription factors that provide cellular stress protection. Their suppression dampens the autophagy response, lowers mitochondrial respiration, and elevates mitochondrial ROS, and their inducible overexpression protects against rotenone toxicity in human iPSC-derived mDANs in vitro. Significantly, we show that LMX1A and LMX1B stability is in part regulated by autophagy, and that these transcription factors bind to multiple ATG8 proteins. Binding is dependent on subcellular localization and nutrient status, with LMX1B interacting with LC3B in the nucleus under basal conditions and associating with both cytosolic and nuclear LC3B during nutrient starvation. Crucially, ATG8 binding stimulates LMX1B-mediated transcription for efficient autophagy and cell stress protection, thereby establishing a novel LMX1B-autophagy regulatory axis that contributes to mDAN maintenance and survival in the adult brain.

Abstract 4815: TMEM9 decreases ER stress-induced misfolded proteins via lysosomal protein degradation

Abstract The endoplasmic reticulum (ER) contributes to various protein modifications such as protein folding. Also, the ER-associated degradation (ERAD) complex removes defective proteins, which prevents the accumulation of misfolded proteins to avoid abnormality of physiological activity. Several physiological and pathological stimuli, such as hypoxia and nutrient starvation lead to unfolded protein responses (UPRs). Given that hyper-proliferation causes hypoxia and glucose shortening, cancer cells might induce UPRs resulting in misfolded protein accumulation and aggregation. Although the ERAD complex removes defective proteins, the capacity of the proteasomal degradation pathway is not enough to completely clear defective proteins in hyper-proliferative cancer cells. Thus, it is likely that cancer cells require additional misfolded protein degradation systems. Transmembrane protein 9 (TMEM9) has been identified as a vesicular acidification amplifier, which is upregulated in cancer cells for Wnt/β-catenin signaling activation via lysosomal protein degradation. In this study, TMEM9 downregulated ER stress-induced misfolded proteins that decrease cancer cell proliferation and viability. ERAD inhibitor declined cancer cell proliferation, reverted by TMEM9. Intriguingly, TMEM9 decreased ERAD-induced apoptosis instead of promoted autophagic cell death under glucose shorten conditions. Here we suggest that lysosomal protein degradation is an additional protein degradation system for the removal of defective proteins and contributes to stable cancer cell proliferation via the clearing of garbage proteins. Citation Format: Dong-Han Wi, Su-Rin Jung, Seong-Ju Lee, Ye-seul Jeon, Youn-Sang Jung, Jae-Il Park. TMEM9 decreases ER stress-induced misfolded proteins via lysosomal protein degradation. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4815.

Abstract NG08: Transcriptional and metabolic dynamics of cancer cells under nutrient deprivation

Abstract Cancer is an evolutionary disease driven by molecular alterations in cancer cells and concomitant tumor microenvironments. Unfortunately, cancer cells often evolve into aggressive tumors that ultimately evade treatment. Thus, in order to improve clinical outcomes, there is an urgent need to define mechanisms by which cancer cells evolve. Recent multi-region sequencing studies, including our own, have inferred phylogenetic evolution across distinct lesions collected from patients. While these studies analyzed the extent of intratumoral heterogeneity across tumor types, the molecular determinants of cancer evolution remain unclear. For example, it is challenging to precisely quantify the adaptive dynamics of a cancer cell lineage before, during, and after a selective pressure. Moreover, tumor microenvironments tend to be spatially and temporally heterogeneous, which complicates evolutionary analyses of cancer cells within these microenvironments. To address these challenges in resolving the evolutionary dynamics of cancer cells, our current work combined bioreactor culturing, longitudinal sampling, single cell sequencing, and metabolomics. Cancer cell lines were selected to represent diverse hematological and solid tumor types, including leukemia, lymphoma, myeloma, colorectal, retinoblastoma, and lung cancers. For four weeks, we consistently maintained multiple environmental parameters of each cancer cell population including temperature, pH, oxygen, and agitation. All cancer cell populations were initiated with the same seeding density in identical media. Since we aimed to quantify growth patterns and to define mechanisms by which cancer cells adapt to nutrient starvation, we allowed each cancer cell population to alter cell density as well as metabolite consumption over the course of the experiment. Every 48 hours, cells and media were collected and preserved to establish a “fossil record” for analysis, and cell density and viability were measured. We found that all cancer cell populations demonstrated exponential growth, plateau and death phases over the course of these experiments, and that each cell line exhibited its own characteristic growth pattern and carrying capacity (range 125 - 250 million cancer cells) despite all cell lines having been grown in the same environmental condition. Moreover, these growth patterns were highly concordant among independently maintained populations. Given our environmental controls, these results suggest that the cancer cell population growth patterns we observed reflected cell-intrinsic features. To explore transcriptional dynamics, we used single cell RNA sequencing to analyze longitudinal samples of the cancer cells. We found that transcriptional subclones emerged over the course of the experiment with altered gene expression profiles, including in genes with functions related to cancer cell metabolism such as biosynthesis, stress responses, and nutrient uptake, indicating putative mechanisms by which the cancer cells were adapting to an increasingly stringent environment. To further define environmental constituents, we analyzed longitudinal media samples that were collected at each timepoint with the cancer cells. Multiple metabolites were consumed within the first seven days of culture, including amino acids, vitamins, and nucleotides. Strikingly, we also observed metabolites that were secreted into the media over the course of the experiment, including nucleotides and signaling molecules. These metabolite patterns were consistent with the concomitant gene expression changes of the cancer cells. Overall, our results showed that the cancer cells were simultaneously adapting to and remodeling their environment rather than solely depleting nutrients. Defining such dynamics, especially in the context of fluctuating environmental conditions, will be essential for mechanistic studies of cancer evolution. Citation Format: Alvin P. Makohon-Moore. Transcriptional and metabolic dynamics of cancer cells under nutrient deprivation. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr NG08.

Abstract 3682: Glutamine addiction of c-MAF overexpressing cells drives dysregulation of purine metabolism

Abstract c-MAF, a bZIP transcription factor, is overexpressed as a consequence of t(14;16) translocation and represents a driver oncogene in this subtype. Clinically, patients with t(14;16) translocation constitute a high-risk group with adverse survival outcomes. Since transcription factors are not classical targets for the development of small molecule inhibitors, elucidating pathways downstream of c-MAF could present as potential therapeutic targets. Critically, gene expression profiling studies of c-MAF showed a class of recurrently deregulated metabolic genes. Hence, this warrants further studies into the underexplored role of metabolism and the deregulated metabolic pathways mediated by c-MAF. First, we confirmed that t(14;16) cells were dependent on c-MAF expression for cell viability. Abrogation of c-MAF deregulated sensitivity to glycolytic inhibitors and influenced metabolite readouts. Importantly, analysis of quantitative metabolic profile of c-MAF driven myeloma using metabolomics identified 41% (48/116) of metabolites to vary significantly. Classification of differential metabolites into metabolic pathways was performed using the software MetaboAnalyst 5.0 to elucidate a role of c-MAF in de novo purine metabolism. Despite an increased source of PRPP derived from the pentose phosphate pathway, we observed significant decrease in output of IMP (fold change=0.6, p=0.028). We postulate that this might result from lowered levels of some non-essential amino acids substrates needed by enzymes in the de novo purine biosynthesis pathway such as glutamine (fold change=0.08, p=0.155) and aspartate (fold change=0.3, p=0.022). Furthermore, GSEA from RNA-seq data performed by c-MAF loss-of-function studies identified ADCY1, ADCY5 and GALR3, suggesting a direct regulation of purine metabolic genes by c-MAF. Notably, to validate whether c-MAF displayed metabolic dependency on glutamine to drive purine biosynthesis, we performed nutrient starvation studies which showed glutamine dependency in c-MAF overexpressed cells. We also systematically knockdown glucose transporters (GLUT8 and GLUT4) or glutamine transporters (ASCT2 and SNAT1) and saw a greater dependency on glutamine transporters for cell viability. As a proof of concept, c-MAF overexpressed cells were more sensitive to purine depletion by LTX suggesting its dependency on purine metabolism. The downstream identification of biological association of c-MAF with cancer metabolism may emerge as a new therapeutic strategy for MM patients. Taken together, our study suggests that metabolic perturbations present as a likely key Achilles heel in MM, and could potentially shift the current treatment paradigm. Citation Format: Julia Sze Lynn Lim, Phyllis Shu Yun Chong, Wee-Joo Chng. Glutamine addiction of c-MAF overexpressing cells drives dysregulation of purine metabolism. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3682.

Evaluation of the Potential of Duckweed as a Human Food, Bioethanol Production Feedstock, and Antileukaemia Drug

This study evaluated the potential of duckweed as a human food, ethanol feedstock, and anticancer drug. First, the nutritional value of wild duckweed was reported for the first time. Its main composition was similar to that of artificially cultivated duckweed, and thus, wild duckweed can serve as a great human food source. In addition, high-starch duckweed induced by nutrient starvation was fermented into bioethanol. A yield of 0.262 g/g, the highest duckweed-ethanol yield reported thus far, was achieved, indicating that duckweed is an excellent feedstock for ethanol production. Finally, the anticancer effects of duckweed flavonoids (DFs) were assessed for the first time using acute myeloid leukaemia (AML) cells as models in vitro and in vivo. The results revealed that DFs possessed antileukaemia activity and were safe and effective for AML therapy. In conclusion, duckweed was demonstrated to be helpful for humans for food security, energy crisis remediation, and tumour treatment.

Phenotypic Switch and Growth of Melanoma Spheroids in the Presence of Mast Cells: Potential Impact of Nutrient-starvation Effects.

Mast cells are abundant in melanoma tumors, and studies suggest that they can be either detrimental or protective for melanoma growth. However, the underlying mechanisms are not fully understood. Here, we adopted an established hanging-drop spheroid system to investigate how mast cells influence melanoma growth and phenotype in a 3-D context. To address the underlying mechanism, we conducted transcriptomic and pathway analyses. In the presence of mast cells or mast cell-conditioned medium, growth of melanoma spheroids was profoundly reduced. Transcriptomic analysis revealed that mast cell-conditioned medium had extensive effects on the gene-expression patterns of melanoma. Pathway analyses revealed profound effects on the expression of genes related to amino acid and protein metabolism. The conditioned medium also induced up-regulation of cancer-related genes, including adhesion molecules implicated in metastatic spreading. In line with this, after transfer to a Matrigel extracellular matrix milieu, spheroids that had been developed in the presence of mast cell-conditioned medium displayed enhanced growth and adhesive properties. However, when assessing the possible impact of nutrient starvation, i.e., reduced nutrient content in mast cell-conditioned medium, we found that the observed effects on growth of melanoma spheroids could potentially be explained by such a scenario. Our findings suggest that the phenotypic alterations of melanoma spheroids grown in the presence of mast cells or mast cell-conditioned media are, at least partly, due to nutrient starvation rather than to the action of factors secreted by mast cells. Our findings may provide insight into the effects on gene-expression events that occur in melanoma tumors under nutrient stress.

Role of the Extracytoplasmic Function Sigma Factor SigE in the Stringent Response of Mycobacterium tuberculosis.

Bacteria respond to nutrient starvation implementing the stringent response, a stress signaling system resulting in metabolic remodeling leading to decreased growth rate and energy requirements. A well-characterized model of stringent response in Mycobacterium tuberculosis is the one induced by growth in low phosphate. The extracytoplasmic function (ECF) sigma factor SigE was previously suggested as having a key role in the activation of stringent response. In this study, we challenge this hypothesis by analyzing the temporal dynamics of the transcriptional response of a sigE mutant and its wild-type parental strain to low phosphate using RNA sequencing. We found that both strains responded to low phosphate with a typical stringent response trait, including the downregulation of genes encoding ribosomal proteins and RNA polymerase. We also observed transcriptional changes that support the occurring of an energetics imbalance, compensated by a reduced activity of the electron transport chain, decreased export of protons, and a remodeling of central metabolism. The most striking difference between the two strains was the induction in the sigE mutant of several stress-related genes, in particular, the genes encoding the ECF sigma factor SigH and the transcriptional regulator WhiB6. Since both proteins respond to redox unbalances, their induction suggests that the sigE mutant is not able to maintain redox homeostasis in response to the energetics imbalance induced by low phosphate. In conclusion, our data suggest that SigE is not directly involved in initiating stringent response but in protecting the cell from stress consequent to the low phosphate exposure and activation of stringent response. IMPORTANCE Mycobacterium tuberculosis can enter a dormant state enabling it to establish latent infections and to become tolerant to antibacterial drugs. Dormant bacteria's physiology and the mechanism(s) used by bacteria to enter dormancy during infection are still unknown due to the lack of reliable animal models. However, several in vitro models, mimicking conditions encountered during infection, can reproduce different aspects of dormancy (growth arrest, metabolic slowdown, drug tolerance). The stringent response, a stress response program enabling bacteria to cope with nutrient starvation, is one of them. In this study, we provide evidence suggesting that the sigma factor SigE is not directly involved in the activation of stringent response as previously hypothesized, but it is important to help the bacteria to handle the metabolic stress related to the adaptation to low phosphate and activation of stringent response, thus giving an important contribution to our understanding of the mechanism behind stringent response development.

Phosphate Starvation Responses in Plants and Microbe Mediated Phosphorus Recycling in Soil: A Review

Phosphorus (P) is an essential macronutrient, crucial for intensification of agricultural production. Although P is a vital nutrient for all life forms but in natural ecosystem it is least available owing to its high reactive nature. Plants have developed diverse adaptive strategies to improve Pi acquisition and utilization under inadequate condition. They undergo numerous modifications at morphological, physiological, and biochemical level to maintain P homeostasis under deplete conditions of P. In last decades, notable development has been made in utilizing microbes for regulating P supply in soil solution. In this review, we focused on the understanding of P uptake from soil, mobilization within plants, phosphate starvation responses and efficacy of microorganisms in facilitating external P to plants. Moreover, other metabolism in microorganism viz. phosphate accumulation is an important trait of microbes to maintain labile P in soil solution. Application of phosphate accumulating microbes is well known in enhanced biological phosphorus removal (EBPR) system and heavy metal remediation. This review also discussed the phosphate accumulation mechanism in microbes and their importance in regulating P supply in soil. Current knowledge and further exploration of mechanism involved by P accumulating microbes may offer better insight of the regulatory events controlling P homeostasis. It will be beneficial for developing more efficient sustainable technologies to augment P use efficiency in soil for better plant growth promotion and nutrient starvation alleviation.

Helicobacter pylori Dormant States Are Affected by Vitamin C.

Helicobacter pylori colonizes human gastric mucosa, overcoming stressful conditions and entering in a dormant state. This study evaluated: (i) H. pylori's physiological changes from active to viable-but-non-culturable (VBNC) and persister (AP) states, establishing times/conditions; (ii) the ability of vitamin C to interfere with dormancy generation/resuscitation. A dormant state was induced in clinical MDR H. pylori 10A/13 by: nutrient starvation (for VBNC generation), incubating in an unenriched medium (Brucella broth) or saline solution (SS), and (for AP generation) treatment with 10xMIC amoxicillin (AMX). The samples were monitored after 24, 48, and 72 h, 8-14 days by OD600, CFUs/mL, Live/Dead staining, and an MTT viability test. Afterwards, vitamin C was added to the H. pylori suspension before/after the generation of dormant states, and monitoring took place at 24, 48, and 72 h. The VBNC state was generated after 8 days in SS, and the AP state in AMX for 48 h. Vitamin C reduced its entry into a VBNC state. In AP cells, Vitamin C delayed entry, decreasing viable coccal cells and increasing bacillary/U-shaped bacteria. Vitamin C increased resuscitation (60%) in the VBNC state and reduced the aggregates of the AP state. Vitamin C reduced the incidence of dormant states, promoting the resuscitation rate. Pretreatment with Vitamin C could favor the selection of microbial vegetative forms that are more susceptible to H. pylori therapeutical schemes.

Comparative transcriptome analyses under individual and combined nutrient starvations provide insights into N/P/K interactions in rice

Crops often suffer from simultaneous limitations of multiple nutrients in soils, including nitrogen (N), phosphorus (P) and potassium (K), which are three major macronutrients essential for ensuring growth and yield. Although plant responses to individual N, P, and K deficiency have been well documented, our understanding of the responses to combined nutrient deficiencies and the crosstalk between nutrient starvation responses is still limited. Here, we compared the physiological responses in rice under seven kinds of single and multiple low nutrient stress of N, P and K, and used RNA sequencing approaches to compare their transcriptome changes. A total of 13,000 genes were found to be differentially expressed under all these single and multiple low N/P/K stresses, and 66 and 174 of them were shared by all these stresses in roots and shoots, respectively. Functional enrichment analyses of the DEGs showed that a group of biological and metabolic processes were shared by these low N/P/K stresses. Comparative analyses indicated that DEGs under multiple low nutrient stress was not the simple summation of single nutrient stress. N was found to be the predominant factor affecting the transcriptome under combined nutrient stress. N, P, or K availability exhibited massive influences on the transcriptomic responses to starvation of other nutrients. Many genes involved in nutrient transport, hormone signaling, and transcriptional regulation were commonly responsive to low N/P/K stresses. Some transcription factors were predicted to regulate the expression of genes that are commonly responsive to N, P, and K starvations. These results revealed the interactions between N, P, and K starvation responses, and will be helpful for further elucidation of the molecular mechanisms underlying nutrient interactions.

The ecl family gene ecl3+ is induced by phosphate starvation and contributes to sexual differentiation in fission yeast.

In Schizosaccharomyces pombe, ecl family genes are induced by several signals, such as starvation of various nutrients, including sulfur, amino acids and Mg2+, and environmental stress, including heat or oxidative stress. These genes mediate appropriate cellular responses and contribute to the maintenance of cell viability and induction of sexual differentiation. Although this yeast has three ecl family genes with overlapping functions, any environmental conditions that induce ecl3+ remain unidentified. We demonstrate that ecl3+ is induced by phosphate starvation, similar to its chromosomally neighboring genes, pho1+ and pho84+, which respectively encode an extracellular acid phosphatase and an inorganic phosphate transporter. ecl3+ expression was induced by the transcription factor Pho7 and affected by the cyclin-dependent kinase (CDK)-activating kinase Csk1. Phosphate starvation induced G1 arrest and sexual differentiation via ecl family genes. Biochemical analyses suggested that this G1 arrest was mediated by the stabilization of the CDK inhibitor Rum1, which was dependent on ecl family genes. This study shows that ecl family genes are required for appropriate responses to phosphate starvation and provides novel insights into the diversity and similarity of starvation responses.

Functional genome annotation and transcriptome analysis of Pseudozyma hubeiensis BOT-O, an oleaginous yeast that utilizes glucose and xylose at equal rates

Pseudozyma hubeiensis is a basidiomycete yeast that has the highly desirable traits for lignocellulose valorisation of being equally efficient at utilization of glucose and xylose, and capable of their co-utilization. The species has previously mainly been studied for its capacity to produce secreted biosurfactants in the form of mannosylerythritol lipids, but it is also an oleaginous species capable of accumulating high levels of triacylglycerol storage lipids during nutrient starvation. In this study, we aimed to further characterize the oleaginous nature of P. hubeiensis by evaluating metabolism and gene expression responses during storage lipid formation conditions with glucose or xylose as a carbon source.The genome of the recently isolated P. hubeiensis BOT-O strain was sequenced using MinION long-read sequencing and resulted in the most contiguous P. hubeiensis assembly to date with 18.95 Mb in 31 contigs. Using transcriptome data as experimental support, we generated the first mRNA-supported P. hubeiensis genome annotation and identified 6540 genes. 80% of the predicted genes were assigned functional annotations based on protein homology to other yeasts. Based on the annotation, key metabolic pathways in BOT-O were reconstructed, including pathways for storage lipids, mannosylerythritol lipids and xylose assimilation. BOT-O was confirmed to consume glucose and xylose at equal rates, but during mixed glucose-xylose cultivation glucose was found to be taken up faster. Differential expression analysis revealed that only a total of 122 genes were significantly differentially expressed at a cut-off of |log2 fold change| ≥ 2 when comparing cultivation on xylose with glucose, during exponential growth and during nitrogen-starvation. Of these 122 genes, a core-set of 24 genes was identified that were differentially expressed at all time points. Nitrogen-starvation resulted in a larger transcriptional effect, with a total of 1179 genes with significant expression changes at the designated fold change cut-off compared with exponential growth on either glucose or xylose.

Metal-Responsive Transcription Factors Co-Regulate Anti-Sigma Factor (Rsd) and Ribosome Dimerization Factor Expression.

Bacteria exposed to stress survive by regulating the expression of several genes at the transcriptional and translational levels. For instance, in Escherichia coli, when growth is arrested in response to stress, such as nutrient starvation, the anti-sigma factor Rsd is expressed to inactivate the global regulator RpoD and activate the sigma factor RpoS. However, ribosome modulation factor (RMF) expressed in response to growth arrest binds to 70S ribosomes to form inactive 100S ribosomes and inhibit translational activity. Moreover, stress due to fluctuations in the concentration of metal ions essential for various intracellular pathways is regulated by a homeostatic mechanism involving metal-responsive transcription factors (TFs). Therefore, in this study, we examined the binding of a few metal-responsive TFs to the promoter regions of rsd and rmf through promoter-specific TF screening and studied the effects of these TFs on the expression of rsd and rmf in each TF gene-deficient E. coli strain through quantitative PCR, Western blot imaging, and 100S ribosome formation analysis. Our results suggest that several metal-responsive TFs (CueR, Fur, KdpE, MntR, NhaR, PhoP, ZntR, and ZraR) and metal ions (Cu2+, Fe2+, K+, Mn2+, Na+, Mg2+, and Zn2+) influence rsd and rmf gene expression while regulating transcriptional and translational activities.

Elucidation of Drug Transport Mechanism by Serum Protein and Development for Pancreatic Cancer Treatment

Human serum albumin (HSA) and α1-acid glycoprotein (AGP) are the major drug-binding proteins in the blood and regulate the tissue transfer of bound drugs. We succeeded in clarifying the three-dimensional structure of AGP for the first time in the world from X-ray crystal structure analysis. Using a site-directed mutagenesis method by constructing yeast expression systems as well as the three-dimensional structure, we elucidated the properties of drug binding sites of AGP. We also found that structural change due to the interaction between AGP and cell membranes causes the release of bound drugs and reported an "AGP-mediated drug transport process." Pancreatic cancer has an extremely low response rate to anticancer drugs compared to other cancers and is resistant to starvation of nutrients including fatty acids. We clarified that glutamine metabolism is involved in this tolerance. Furthermore, aiming at efficient drug delivery and effective treatment for pancreatic cancer, we focused on nitric oxide (NO) which increases pancreatic blood flow and has a cell-killing effect on tumors and surrounding stromal tissues. We successfully synthesized nitrated phenylbutyrate (NPB), which binds to HSA and has an antitumor effect in vitro and vivo. The binding of NPB to HSA is considered to be useful for delivery to tumors through the enhanced permeability and retention (EPR) effect and HSA receptors.

The phospholipid flippase ATP9A enhances macropinocytosis to promote nutrient starvation tolerance in hepatocellular carcinoma.

Macropinocytosis is an effective strategy to mitigate nutrient starvation. It can fuel cancer cell growth in nutrient-limited conditions. However, whether and how macropinocytosis contributes to the rapid proliferation of hepatocellular carcinoma cells, which frequently experience an inadequate nutrient supply, remains unclear. Here, we demonstrated that nutrient starvation strongly induced macropinocytosis in some hepatocellular carcinoma cells. It allowed the cells to acquire extracellular nutrients and supported their energy supply to maintain rapid proliferation. Furthermore, we found that the phospholipid flippase ATP9A was critical for regulating macropinocytosis in hepatocellular carcinoma cells and that high ATP9A levels predicted a poor outcome for patients with hepatocellular carcinoma. ATP9A interacted with ATP6V1A and facilitated its transport to the plasma membrane, which promoted plasma membrane cholesterol accumulation and drove RAC1-dependent macropinocytosis. Macropinocytosis inhibitors significantly suppressed the energy supply and proliferation of hepatocellular carcinoma cells characterised by high ATP9A expression under nutrient-limited conditions. These results have revealed a novel mechanism that overcomes nutrient starvation in hepatocellular carcinoma cells and have identified the key regulator of macropinocytosis in hepatocellular carcinoma. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Non-muscle MYH10/myosin IIB recruits ESCRT-III to participate in autophagosome closure to maintain neuronal homeostasis

ABSTRACT Dysfunction of the endosomal sorting complex required for transport (ESCRT) has been linked to frontotemporal dementia (FTD) due in part to the accumulation of unsealed autophagosomes. However, the mechanisms of ESCRT-mediated membrane closure events on phagophores remain largely unknown. In this study, we found that partial knockdown of non-muscle MYH10/myosin IIB/zip rescues neurodegeneration in both Drosophila and human iPSC-derived cortical neurons expressing FTD-associated mutant CHMP2B, a subunit of ESCRT-III. We also found that MYH10 binds and recruits several autophagy receptor proteins during autophagosome formation induced by mutant CHMP2B or nutrient starvation. Moreover, MYH10 interacted with ESCRT-III to regulate phagophore closure by recruiting ESCRT-III to damaged mitochondria during PRKN/parkin-mediated mitophagy. Evidently, MYH10 is involved in the initiation of induced but not basal autophagy and also links ESCRT-III to mitophagosome sealing, revealing novel roles of MYH10 in the autophagy pathway and in ESCRT-related FTD pathogenesis. Abbreviations: ALS: amyotrophic lateral sclerosis; AP: autophagosome; Atg: autophagy-related; ESCRT: endosomal sorting complex required for transport; FTD: frontotemporal dementia

The adenosinergic machinery in cancer: In-tandem insights from basic mechanisms to therapy.

Extracellular adenosine (eADO) signaling has emerged as an increasingly important regulator of immune responses, including tumor immunity. eADO is mainly produced from extracellular ATP (eATP) hydrolysis. eATP is rapidly accumulated in the extracellular space following cell death or cellular stress triggered by hypoxia, nutrient starvation, or inflammation. eATP plays a pro-inflammatory role by binding and activating the P2 purinergic receptors (P2X and P2Y), while eADO has been reported in many studies to mediate immunosuppression by activating the P1 purinergic receptors (A1, A2A, A2B, and A3) in diverse immune cells. Consequently, the hydrolysis of eATP to eADO alters the immunosurveillance in the tumor microenvironment (TME) not only by reducing eATP levels but also by enhancing adenosine receptor signaling. The effects of both P1 and P2 purinergic receptors are not restricted to immune cells. Here we review the most up-to-date understanding of the tumor adenosinergic system in all cell types, including immune cells, tumor cells, and stromal cells in TME. The potential novel directions of future adenosinergic therapies in immuno-oncology will be discussed.

Mitochondrial dynamics in macrophages: divide to conquer or unite to survive?

Mitochondria have long been appreciated as the metabolic hub of cells. Emerging evidence also posits these organelles as hubs for innate immune signalling and activation, particularly in macrophages. Macrophages are front-line cellular defenders against endogenous and exogenous threats in mammals. These cells use an array of receptors and downstream signalling molecules to respond to a diverse range of stimuli, with mitochondrial biology implicated in many of these responses. Mitochondria have the capacity to both divide through mitochondrial fission and coalesce through mitochondrial fusion. Mitochondrial dynamics, the balance between fission and fusion, regulate many cellular functions, including innate immune pathways in macrophages. In these cells, mitochondrial fission has primarily been associated with pro-inflammatory responses and metabolic adaptation, so can be considered as a combative strategy utilised by immune cells. In contrast, mitochondrial fusion has a more protective role in limiting cell death under conditions of nutrient starvation. Hence, fusion can be viewed as a cellular survival strategy. Here we broadly review the role of mitochondria in macrophage functions, with a focus on how regulated mitochondrial dynamics control different functional responses in these cells.