Biology subjects

Goldberg, D.

Publications and source records attributed to Goldberg, D..

11 recordsLinked to original sources

bioRxiv2026

SETD6-mediated methylation of PPARγ establishes a transcriptional feedback circuit promoting lipid accumulation in liver-derived cells

Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) is a key transcriptional regulator of genes mediating adipogenesis (fat-cell differentiation) and lipid storage in several cell types like hepatocytes. As such, its regulation is crucial for cell and organismal physiology. Indeed, PPAR{gamma}s activity is regulated by multiple mechanisms, including post-transcriptional modifications, which, when dys-coordinated, may contribute to the pathogenesis of various states, including obesity, insulin resistance, and fatty liver disease. Here, we demonstrate that SETD6 binds to, and methylates PPAR{gamma} at lysine 170 (K170) both in vitro and in liver-derived cells. This methylation event, in turn, is required for PPAR{gamma}-mediated activation of SETD6 transcription via promoter binding, forming a positive feedback regulatory loop. RNA sequencing revealed that both SETD6 and PPAR{gamma} methylation at K170 are required for full induction of lipid metabolism genes expression, manifesting functionally in lipid droplet biogenesis in liver-derived cells. Together, our findings uncover a novel role for lysine methylation of PPAR{gamma} in the regulation of lipid synthesis and lipid droplet biogenesis, thereby identifying putative new therapeutic targets for lipid over-production diseases, including MAFLD (Metabolic dysfunction-associated fatty liver disease) and obesity.

cell biology↗

bioRxiv2025

Quinolines interfere with heme-mediated activation of artemisinins

Artemisinin-based combination therapies (ACTs) remain the mainstay of treatment for Plasmodium falciparum malaria, despite reports of ACT treatment failure. ACTs consist of an artemisinin and a longer-lived partner drug, which is often a quinoline. Given that heme is central to the mechanism of action of artemisinins and some quinolines, we hypothesized that these antimalarials would exhibit strong drug-drug interactions. Previous studies using standard 48 h or 72 h assays identified additive to mildly antagonistic interactions between artemisinins and quinolines. Here, we sought to re-evaluate these interactions using a pulsing assay that better mimics the short in vivo half-life of artemisinins. We found that chloroquine (CQ), piperaquine (PPQ), and amodiaquine substantially antagonize dihydroartemisinin (DHA), the active metabolite of artemisinins. CQ-DHA antagonism was notably exacerbated in CQ-resistant parasites, resulting in a superantagonistic phenotype in isobolograms. Further, we found that CQ co-treatment conferred artemisinin resistance to Kelch 13 wild type parasites in the ring stage survival assay. Using a small molecule probe to measure chemically reactive heme in live parasites, we determined that quinolines block artemisinin activation by rendering cytosolic heme inert. Finally, we probed beyond traditional ACTs, evaluating interactions of the proposed triple ACT, DHA-PPQ-Mefloquine, as well as OZ439-quinoline combinations, which were all found to be antagonistic. Collectively, these in vitro data suggest that peroxide-quinolines may have liabilities as combination therapies.

pharmacology and toxicology↗

bioRxiv2024

The Plasmodium falciparum NCR1 membrane protein is a novel antimalarial target that exports cholesterol to maintain membrane homeostasis

Malaria is an extremely devastating parasitic infection that kills over half a million people each year. It is the leading cause of death in many developing countries, in part, due to a lack of resources and readily available therapeutics. Unfortunately, the most prevalent and deadliest causative agent of malaria, Plasmodium falciparum, has developed resistance to nearly all currently available antimalarial drugs. The P. falciparum Niemann-Pick Type C1-related (PfNCR1) transporter has been identified as a druggable target, as it is required for membrane homeostasis of the parasite. However, the structure and detailed molecular mechanism of this membrane protein are not yet available. Here we present three structures of PfNCR1 both in the absence and presence of the functional inhibitor MMV009108 at resolutions between 2.98 [A] and 3.81 [A] using single-particle cryo-electron microscopy (cryo-EM). The data suggest that PfNCR1 binds cholesterol and forms a cholesterol transport tunnel to modulate the composition of the parasite plasma membrane. Cholesterol efflux assays substantiate this as they show that PfNCR1 is an exporter capable of extruding cholesterol from the membrane. Additionally, the inhibition mechanism of MMV009108 appears to be due to a direct blockage of PfNCR1, preventing this transporter from shuttling cholesterol.

biochemistry↗

bioRxiv2024

Repeated fasting events sensitize enhancers, transcription factor activity and gene expression to support augmented ketogenesis

Mammals withstand frequent and prolonged fasting periods due to hepatic production of ketone bodies. Because the fasting response is transcriptionally-regulated, we asked whether enhancer dynamics impose a transcriptional program during recurrent fasting and whether this generates effects distinct from a single fasting bout. We found that mice undergoing alternate-day fasting (ADF) respond profoundly differently to a following fasting bout compared to mice first experiencing fasting. Hundreds of genes enabling ketogenesis are sensitized (induced more strongly by fasting following ADF). Liver enhancers regulating these genes are also sensitized and harbor increased binding of PPAR, the main ketogenic transcription factor. ADF leads to augmented ketogenesis compared to a single fasting bout in wild-type, but not hepatocyte-specific PPAR-deficient mice. Thus, we found that past fasting events are remembered in hepatocytes, sensitizing their enhancers to the next fasting bout and augment ketogenesis. Our findings shed light on transcriptional regulation mediating adaptation to repeated signals. O_FIG O_LINKSMALLFIG WIDTH=159 HEIGHT=200 SRC="FIGDIR/small/592891v1_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@13c84faorg.highwire.dtl.DTLVardef@3e42f3org.highwire.dtl.DTLVardef@19d9603org.highwire.dtl.DTLVardef@e3ae25_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Transcriptional cascades during fasting amplify gluconeogenesis and instigate a secondary wave of ketogenic gene transcription

Background & AimsDuring fasting, bodily homeostasis is maintained due to hepatic production of glucose (gluconeogenesis) and ketone bodies (ketogenesis). The main hormones governing hepatic fuel production are glucagon and glucocorticoids that initiate transcriptional programs aimed at supporting gluconeogenesis and ketogenesis. MethodsUsing primary mouse hepatocytes as an ex vivo model, we employed transcriptomic analysis (RNA-seq), genome-wide profiling of enhancer dynamics (ChIP-seq), perturbation experiments (inhibitors, shRNA), hepatic glucose production measurements and computational analyses. ResultsWe found that in addition to the known metabolic genes transcriptionally induced by glucagon and glucocorticoids, these hormones induce a set of genes encoding transcription factors (TFs) thereby initiating transcriptional cascades. Upon activation by glucocorticoids, the glucocorticoid receptor (GR) induced the genes encoding two TFs: CCAAT/enhancer-binding protein beta (C/EBP{beta}) and peroxisome proliferator-activated receptor alpha (PPAR). We found that C/EBP{beta} mainly serves as an amplifier of hormone-induced gene programs in hepatocytes. C/EBP{beta} augmented gluconeogenic gene expression and hepatic glucose production. Conversely, the GR-PPAR cascade initiated a secondary transcriptional wave of genes supporting ketogenesis. The cascade led to synergistic induction of ketogenic genes which is dependent on protein synthesis. Genome-wide analysis of enhancer dynamics revealed numerous enhancers activated by the GR-PPAR cascade. These enhancers were proximal to ketogenic genes, enriched for the PPAR response element and showed increased PPAR binding. ConclusionThis study reveals abundant transcriptional cascades occurring during fasting. These cascades serve two separated purposes: the amplification of the primary gluconeogenic transcriptional program and the induction of a secondary gene program aimed at enhancing ketogenesis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/588039v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@e02207org.highwire.dtl.DTLVardef@da0c7aorg.highwire.dtl.DTLVardef@1695a8borg.highwire.dtl.DTLVardef@1b78fb0_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2023

Inversely regulated immune-related processes mediate anxiety-obesity links in zebrafish

Anxiety disorders often associate with metabolic impairments, but the underlying developmental and molecular mechanisms are yet unknown. To seek RNAs that may link anxiety and obesity, we subjected RNA from zebrafish larvae of a caffeine-induced anxiety model and a high fat diet (HFD)-induced obesity model to RNA-sequencing. We found differentially expressed genes in the larval anxiety and obesity models, including long noncoding RNAs and transfer fragment RNAs. Surprisingly, they were inversely regulated and comprised overrepresentation of immune system pathways, e.g., interleukin signaling and inflammation. Similarly, inverse regulation persisted in adulthood, but with different overrepresented immune system processes, e.g., T cell activation, leukocyte cell-cell adhesion and antigen processing and presentation. Furthermore, unlike the known link in adult zebrafish, obesity in zebrafish larvae was not accompanied by anxiety-like behavior. These results may reflect an antagonistic pleiotropic phenomenon involving re-adjusted modulation of the anxiety-metabolic links with the immune system. Furthermore, the HFD potential to normalize the anxiety-upregulated immune-related genes may explain previously reported protective roles of high fat diet in rodent anxiety and Alzheimers disease models.

neuroscience↗

bioRxiv2023

Activation of the Plasmodium egress effector subtilisin-like protease 1 is achieved by plasmepsin X destruction of the propiece.

Following each round of replication, daughter merozoites of the malaria parasite Plasmodium falciparum escape (egress) from the infected host red blood cell (RBC) by rupturing the parasitophorous vacuole membrane (PVM) and the RBC membrane (RBCM). A proteolytic cascade orchestrated by the parasites serine protease, subtilisin-like protease 1 (SUB1) regulates the membrane breakdown. SUB1 activation involves primary auto-processing of the 82 kDa zymogen to a 54 kDa (p54) intermediate that remains bound to its inhibitory propiece (p31) post cleavage. A second processing step converts p54 to the terminal 47 kDa (p47) form of SUB1. Although the aspartic protease plasmepsin X (PM X) has been implicated in the activation of SUB1, the mechanism remains unknown. Here, we show that upon knockdown of PM X the inhibitory p31/p54 complex of SUB1 accumulates in the parasites. Using recombinant PM X and SUB1, we show that PM X can directly cleave both p31 and p54. We have mapped the cleavage sites on recombinant p31. Furthermore, we demonstrate that the conversion of p54 to p47 can be effected by cleavage at either a SUB1 or PM X cleavage site that are adjacent to one another. Importantly once the p31 is removed, p54 is fully functional inside the parasites suggesting that the conversion to p47 is dispensable for SUB1 activity. Relief of propiece inhibition via a heterologous protease is a novel mechanism for subtilisin activation. Significance StatementMalaria parasites replicate inside a parasitophorous vacuole within the host red blood cells. Exit of mature progeny from the infected host cells is essential for further dissemination. Parasite exit is a highly regulated, explosive process that involves membrane breakdown. To do this, the parasite utilizes a serine protease, called the subtilisin-like protease 1 or SUB1 that proteolytically activates various effector proteins. SUB1 activity is dependent on an upstream protease, called plasmepsin X (PM X), although the mechanism was unknown. Here we describe the molecular basis for PM X mediated SUB1 activation. PM X proteolytically degrades the inhibitory segment of SUB1, thereby activating it. Involvement of a heterologous protease is a novel mechanism for subtilisin activation.

microbiology↗

bioRxiv2021

Native structure of the RhopH complex, a key determinant of malaria parasite nutrient acquisition

The RhopH complex is implicated in malaria parasites ability to invade and create new permeability pathways in host erythrocytes, but its mechanisms remain poorly understood. Here we enrich the endogenous RhopH complex in a native soluble form, comprising RhopH2, CLAG3.1 and RhopH3, directly from parasite cell lysates and determine its atomic structure using cryo electron microscopy, mass spectrometry, and the cryoID program. This first direct observation of an exported P. falciparum transmembrane protein--in a soluble, trafficking state and with atomic details of buried putative membrane-insertion helices--offers insights into assembly and trafficking of RhopH and other parasite-derived complexes to the erythrocyte membrane. Our study demonstrates the potential endogenous structural proteomics approach holds for elucidating the molecular mechanisms of hard-to-isolate complexes in their native, functional forms.

biophysics↗

bioRxiv2020

Total mercury concentrations in invasive lionfish (Pterois volitans/miles) from the Atlantic coast of Florida

ABSTRACTInvasive lionfish (Pterois volitans/miles) pose a serious threat to marine ecosystems throughout the western Atlantic Ocean and Caribbean Sea. The development of a fishery for lionfish has been proposed as a strategy for controlling populations; however, there is concern about consumption of this species by humans due to its high trophic position and potential for bioaccumulation of mercury. We analyzed total mercury (THg) in tissues of lionfish from two locations on the east coast of Florida. THg in lionfish increased with size and differed by location and sex. THg was highest in muscle tissue and was strongly positively correlated among tissues. THg in lionfish was lower than other commonly consumed marine fishes, and falls into Florida’s least restrictive advisory level. Consumption of lionfish poses a low risk and concerns over mercury bioaccumulation should not present a significant barrier to lionfish harvest.View Full Text

pharmacology and toxicology↗

bioRxiv2020

The evolution of human SIRT1 for multi-specificity is enabled by active-site adaptation

Many enzymes that catalyze protein post-translational modifications (PTM) can specifically modify multiple target proteins. However, little is known regarding the molecular basis and evolution of multi-specificity in these enzymes. Here, we used bioinformatics and experimental approaches to investigate the molecular basis and evolution of multi-specificity in the sirtuin-1 (SIRT1) deacetylase. Guided by bioinformatics analysis of SIRT1 orthologues and substrates, we identified and examined key mutations that have occurred during the evolution of human SIRT1. We found that while these mutants maintain high activity toward conserved histone substrates they exhibit a dramatic loss of activity toward acetylated p53 protein that appeared only in Metazoa. These results demonstrate that active site substitutions in SIRT1 are essential to enable the inclusion of p53 substrate. Our results are consistent with a model in which promiscuous ancestral activity can provide an evolutionary starting point from which multispecificity toward a number of cellular substrates can develop.

biochemistry↗

bioRxiv2019

Dispersal limitation and stage-specific local filtering processes vary with climate across a network of alpine grasslands

Dispersal dynamics and local filtering interactively generate regional vegetation patterns, but empirical evidence of their combined influence in nature is scarce, representing a missing link between our theoretical understanding of community assembly and real-world observation. Here, we compare seed and adult plant communities at twelve grassland sites with different climates in southern Norway to explore the degree to which community membership is shaped by dispersal limitation and local niche-based filtering, and how this varies with climate. To do this, we first divide species at each site into two groups: \"locally-transient\" species, which occur as seeds but are rare or absent as adults (i.e., they arrive but are filtered out), or \"locally-persistent\" species, which occur consistently as adults in annual vegetation surveys. We then ask questions to reveal where, when, why, and how locally-transient species are systematically disfavored during community assembly. Our results led to four main conclusions: (1) the strength of local filtering on community membership increased with temperature, (2) surprisingly, local filtering was stronger during seedling emergence than during seedling establishment, (3) climate-based niche differences drove differential performance among species, especially for seeds dispersing outside of their realized climate niches into more stressful (colder and drier) climates, and (4) locally-transient species had traits that may made them better dispersers (smaller seeds) but poorer competitors for light (shorter statures, less persistent clonal connections) than locally-persistent species, providing a potential explanation for why they arrived to new sites but failed to establish persistent adult populations. Our study is one of the first to combine seed, seedling, and adult survey data across multiple sites with different climates to provide a rigorous empirical evaluation of the combined influence of dispersal limitation and local filtering on the generation and maintenance of climate-associated vegetation patterns.

ecology↗