Biology subjects

Gross, S. S.

Publications and source records attributed to Gross, S. S..

9 recordsLinked to original sources

bioRxiv2025

NAD⁺ Replenishment Mitigates Cardiomyocyte Senescence and Corrects Heart Failure with Preserved Ejection Fraction in Aged Mice

Cardiomyocyte senescence, characterized by elevated cell cycle inhibitor expression, persistent DNA damage response, and mitochondrial dysfunction, contributes to myocardial stiffness and the progression of heart failure with preserved ejection fraction (HFpEF), the most common form of heart failure affecting individuals over 65. In this study, we investigated the role of NAD metabolism in cardiomyocyte senescence and cardiac function. Aged mice exhibited reduced cardiac NAD levels, impaired NAD biosynthesis and mobilization, and increased consumption, leading to suppressed SIRT1/6 activity and accumulation of senescent cardiomyocytes. This was accompanied by diastolic dysfunction consistent with HFpEF. In senescent AC16 cardiomyocytes, NAD depletion promoted senescence, which was reversed by the NAD precursors nicotinamide riboside (NR) and dihydronicotinamide riboside (NRH). In aged mice, two months of NR or NRH treatment improved diastolic function and reduced cardiomyocyte senescence. While NR primarily activated SIRT1 to suppress cell cycle arrest markers, NRH more robustly activated both SIRT1 and SIRT6, enhancing DNA damage repair. Acetylated H2AX, a SIRT6 substrate elevated in aged hearts and senescent cells, was selectively deacetylated by NRH. These findings identify NAD availability as a critical regulator of cardiac senescence and support NAD precursors, particularly NRH, as promising senescence-reducing therapies for treating aging-associated HFpEF.

pharmacology and toxicology↗

bioRxiv2024

Epigenetic regulation by TET1 in gene-environmental interactions influencing susceptibility to congenital malformations

The etiology of neural tube defects (NTDs) involves complex gene-environmental interactions. Folic acid (FA) prevents NTDs, but the mechanisms remain poorly understood and at least 30% of human NTDs resist the beneficial effects of FA supplementation. Here, we identify the DNA demethylase TET1 as a nexus of folate-dependent one-carbon metabolism and genetic risk factors post-neural tube closure. We determine that cranial NTDs in Tet1-/- embryos occur at two to three times higher penetrance in genetically heterogeneous than in homogeneous genetic backgrounds, suggesting a strong impact of genetic modifiers on phenotypic expression. Quantitative trait locus mapping identified a strong NTD risk locus in the 129S6 strain, which harbors missense and modifier variants at genes implicated in intracellular endocytic trafficking and developmental signaling. NTDs across Tet1-/- strains are resistant to FA supplementation. However, both excess and depleted maternal FA diets modify the impact of Tet1 loss on offspring DNA methylation primarily at neurodevelopmental loci. FA deficiency reveals susceptibility to NTD and other structural brain defects due to haploinsufficiency of Tet1. In contrast, excess FA in Tet1-/- embryos drives promoter DNA hypermethylation and reduced expression of multiple membrane solute transporters, including a FA transporter, accompanied by loss of phospholipid metabolites. Overall, our study unravels interactions between modified maternal FA status, Tet1 gene dosage and genetic backgrounds that impact neurotransmitter functions, cellular methylation and individual susceptibilities to congenital malformations, further implicating that epigenetic dysregulation may underlie NTDs resistant to FA supplementation.

developmental biology↗

bioRxiv2023

Progesterone induces meiosis through two obligate co-receptors with PLA2 activity

The steroid hormone progesterone (P4) regulates multiple aspects of reproductive and metabolic physiology. Classical P4 signaling operates through nuclear receptors that regulate transcription. In addition, P4 signals through membrane P4 receptors (mPRs) in a rapid nongenomic modality. Despite the established physiological importance of P4 nongenomic signaling, the details of its signal transduction cascade remain elusive. Here, using Xenopus oocyte maturation as a well- established physiological readout of nongenomic P4 signaling, we identify the lipid hydrolase ABHD2 (/{beta} hydrolase domain-containing protein 2) as an essential mPR{beta} co-receptor to trigger meiosis. We show using functional assays coupled to unbiased and targeted cell-based lipidomics that ABHD2 possesses a phospholipase A2 (PLA2) activity that requires mPR{beta}. This PLA2 activity bifurcates P4 signaling by inducing clathrin-dependent endocytosis of mPR{beta}, resulting in the production of lipid messengers that are G-protein coupled receptors agonists. Therefore, P4 drives meiosis by inducing an ABHD2 PLA2 activity that requires both mPR{beta} and ABHD2 as obligate co-receptors. Significance StatementNongenomic progesterone signaling is important for many physiological functions yet the details of its signaling remain elusive. Here we define the early signaling steps downstream of membrane progesterone receptor {beta} (mPR{beta}) during Xenopus oocyte meiosis. We show that progesterone requires two cell membrane receptors to work in unison to signal. The co-receptor complex possesses lipase activity that produces lipid messenger and induces receptor endocytosis to trigger meiosis progression. Our findings have broad physiological implications because nongenomic progesterone signaling operates in many tissues and regulates reproduction and metabolism.

cell biology↗

bioRxiv2022

Selective detection of m6A derived from mRNA using the Phospho-tag m6A assay

N6-methyladenosine (m6A) is a modified nucleotide found in mRNA, ribosome RNA (rRNA) and small nuclear RNA (snRNA). m6A in mRNA has important roles in regulating mRNA stability, splicing, and other processes. Numerous studies have described m6A as a dynamic modification using mass spectrometry-based quantification of m6A in mRNA samples prepared from different cellular conditions. However, these results have been questioned based on the finding that the mRNA purification protocols often result in varying levels of rRNA contamination. Additionally, mRNA purification protocols disproportionately enrich for the 3 ends of mRNA, a region that is enriched in m6A. To address these problems, we developed the Phospho-tag m6A assay, a highly efficient method for quantifying m6A specifically from mRNA. In this assay, a series of selective RNase digestion steps is performed, which results in m6A from rRNA and snRNA being liberated as m6A monophosphate, while m6A from mRNA is mostly liberated as m6A nucleoside. m6A levels are normalized to transcript levels, using m7G monophosphate liberated by yDcpS decapping enzyme as a surrogate for mRNA levels. Notably, this approach uses total cellular RNA, rather than purified mRNA, which simplifies the steps for m6A detection and overcomes the 3-end biases associated with mRNA purification. Overall, the Phospho-tag m6A provides a simple and efficient method for quantification of mRNA-derived m6A from total RNA samples.

molecular biology↗

bioRxiv2021

Non-conserved metabolic regulation by LKB1 distinguishes human and mouse lung adenocarcinoma

KRAS is the most frequently mutated oncogene in human lung adenocarcinomas (hLUAD) and activating mutations in KRAS frequently co-occur with loss-of-function mutations in the tumor suppressor genes, TP53 or STK11/LKB1. However, mutation of all three genes is rarely observed in hLUAD, even though engineered mutations of all three genes produces a highly aggressive lung adenocarcinoma in mice (mLUAD). Here we provide an explanation of this difference between hLUAD and mLUAD by uncovering an evolutionary divergence in regulation of the glycolytic enzyme triosephosphate isomerase (TPI1). Using KRAS/TP53 mutant hLUAD cell lines, we show that TPI1 enzymatic activity can be altered via phosphorylation at Ser21 by the Salt Inducible Kinases (SIKs) in an LKB1-dependent manner; this allows modulation of glycolytic flux between completion of glycolysis and production of glycerol lipids. This metabolic flexibility appears to be critical in rapidly growing cells with KRAS and TP53 mutations, explaining why loss of LKB1 creates a metabolic liability in these tumors. In mice, the amino acid at position 21 of TPI1 is a Cys residue which can be oxidized to alter TPI1 activity, allowing regulation of glycolytic flux balance without a need for SIK kinases or LKB1. Our findings reveal an unexpected role for TPI1 in metabolic reprogramming and suggest that LKB1 and SIK family kinases are potential targets for treating KRAS/TP53 mutant hLUAD. Our data also provide a cautionary example of the limits of genetically engineered murine models as tools to study human diseases such as cancers.

cancer biology↗

bioRxiv2019

Accelerated trans-sulfuration metabolically defines a discrete subclass of ALS patients

Amyotrophic lateral sclerosis (ALS) is a disease characterized by progressive paralysis and death. Most ALS cases are sporadic (sALS) and patient heterogeneity poses a formidable challenge for the development of viable biomarkers and effective therapies. Applying untargeted metabolite profiling on 77 sALS patient-derived primary dermal fibroblast lines and 45 sex/age matched controls, we found that [~]25% of cell lines (termed sALS-1) are characterized by upregulated trans-sulfuration, where methionine-derived homocysteine is channeled into cysteine and glutathione synthesis. sALS-1 fibroblasts exhibit a growth defect when grown under oxidative conditions, that can be fully-rescued by N-acetylcysteine. [U-13C]-glucose tracing shows that activation of the trans-sulfuration pathway is associated with accelerated glucose flux into the TCA cycle. Based on four metabolites, we developed a support vector machine model capable of distinguishing sALS-1 with 97.5% accuracy. Importantly, plasma metabolite profiling identifies a systemic perturbation of cysteine metabolism as a hallmark of sALS-1. These results indicate that sALS patients can be stratified into distinct metabotypes, differently sensitive to metabolic stress, and provides new insights into metabolic biomarkers for personalized sALS therapy.

neuroscience↗

bioRxiv2018

APOE4 is Associated with Differential Regional Vulnerability to Bioenergetic Deficits in Aged APOE Mice

The {varepsilon}4 allele of apolipoprotein E (APOE) is the dominant genetic risk factor for late-onset Alzheimers disease (AD). However, the reason for the association between APOE4 and AD remains unclear. While much of the research has focused on the ability of the apoE4 protein to increase the aggregation and decrease the clearance of A{beta}, there is also an abundance of data showing that APOE4 negatively impacts many additional processes in the brain, including bioenergetics. In order to gain a more comprehensive understanding of the APOE4s role in AD pathogenesis, we performed a multi-omic analysis of APOE4 vs. APOE3 expression in the entorhinal cortex (EC) and primary visual cortex (PVC) of aged APOE mice. These studies revealed region-specific alterations in several bioenergetic pathways, including oxidative phosphorylation (OxPhos), the TCA-cycle and fatty acid metabolism. Follow-up analysis utilizing the Seahorse platform revealed decreased mitochondrial respiration in the hippocampus and cortex of aged APOE4 vs. APOE3 mice, but not in the EC of these mice. Additional studies, as well as the original multi-omic data suggest that bioernergetic pathways in the EC of aged APOE mice may be differentially regulated by APOE4 expression. Given the importance of the EC as one of the first regions to be affected by AD pathology in humans, this differential bionenergetic regulation observed in the EC vs. other brain regions of aged APOE4 mice may play an important role in the pathogenesis of AD, particularly among APOE4 carriers.

neuroscience↗

bioRxiv2018

The RNA demethylase FTO targets m6Am in snRNA to establish distinct methyl isoforms that influence splicing

Small nuclear RNAs (snRNAs) are core spliceosome components and mediate pre-mRNA splicing. During their biogenesis, snRNAs acquire several constitutive nucleotide modifications. Here we show that snRNAs also contain a regulated and reversible nucleotide modification causing them to exist as two different methyl isoforms, m1 and m2, reflecting the methylation state of the adenosine adjacent to the snRNA cap. We find that snRNA biogenesis involves the formation of an initial m1-isoform with a single-methylated adenosine (2-O-methyladenosine, Am), which is then converted to a dimethylated m2-isoform (N6,2-O-dimethyladenosine, m6Am). The relative m1- and m2-isoform levels are determined by the RNA demethylase FTO, which selectively demethylates the m2-isoform. We show FTO is inhibited by endogenous metabolites, resulting in increased m2-snRNA levels. Furthermore, cells that exhibit high m2-snRNA levels show altered patterns of alternative splicing. Together, these data reveal that FTO has a central role in snRNA biogenesis and controls a previously unknown step of snRNA processing involving reversible methylation, thereby providing a potential link between reversible RNA modifications and mRNA splicing.

molecular biology↗

bioRxiv2018

Accelerated lipid catabolism and autophagy are cancer survival mechanisms under inhibited glutaminolysis

Suppressing glutaminolysis does not always induce cancer cell death in glutamine-dependent tumors because cells may switch to alternative energy sources. To reveal compensatory metabolic pathways, we investigated the metabolome-wide cellular response to inhibited glutaminolysis. We conducted metabolic profiling in the triple-negative breast cancer cell line MB-MDA-231, treated with different dosages of glutaminase inhibitor C.968 at multiple time points. We found that multiple molecules involved in lipid catabolism responded directly to glutamate deficiency as a presumed compensation for energy deficit. Accelerated lipid catabolism, together with oxidative stress induced by glutaminolysis inhibition, triggered autophagy. We therefore simultaneously inhibited glutaminolysis and autophagy, which induced cancer cell death. Our study emphasizes the potential of non-targeted metabolomics to characterize and identify metabolic escape mechanisms contributing to cancer cell survival under treatment. Our findings add to the increasing evidence that combined inhibition of glutaminolysis and autophagy may be effective in glutamine-addicted cancers.

cancer biology↗