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Cholesterol metabolism reprogramming plays essential roles in hepatocellular carcinoma (HCC). However, precisely how cholesterol metabolism is dysregulated is not clear. Here, we show that the palmitoyltransferase ZDHHC3 and depalmitoylase ABHD17A regulate HCC cell cholesterol biosynthesis by dynamically S-acylating SREBP cleavage-activating protein (SCAP). SCAP S-acylation by ZDHHC3 at C264 antagonizes HACE1-mediated SCAP ubiquitination. Intriguingly, SREBP2 transcriptionally upregulates ZDHHC3 to form a positive feedback loop, which explains why negative feedback regulation of SCAP/SREBP2 signaling fails in HCC. Increased cholesterol in the tumor microenvironment (TME) restrains CD4 + T cell cytotoxicity. Hence, the cholesterol metabolism reprogramming and cholesterol level alternation in the TME cooperate to promote HCC development. We identified a small-molecule inhibitor of ZDHHC3 that, combined with anti-PD-1 immunotherapy, inhibited diethyl nitrosamine (DEN)/CCl 4 -induced HCC growth in mice. ZDHHC3-mediated SCAP S-acylation reprograms cholesterol metabolism and promotes HCC immune escape. ZDHHC3 is thus identified as a rational chemotherapy target for HCC., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Certain metabolites in the tumor microenvironment (TME) can alter innate immunity. Here, it is shown how phosphomevalonate kinase (PMVK) allows hepatocellular carcinoma (HCC) cells to overcome the anti-tumor immunity mediated by CD8 + T cells. In HCCs, depletion of PMVK is required to facilitate CD8 + T cell activation and their subsequent suppression of tumor growth. Mechanistically, PMVK phosphorylates and stabilizes glutamate decarboxylase 1 (GAD1), thus increasing the synthesis of γ-aminobutyric acid (GABA), which normally functions as a neurotransmitter. However, PMVK also recruits acetyl-CoA acetyltransferase 1 (ACAT1) and allows it to convert GABA, to 4-acetaminobutyric acid (4-Ac-GABA), which is released into the TME. There, 4-Ac-GABA activates the GABAA receptor (GABAAR) on CD8 + T cells, which inhibits AKT1 signaling. This in turn suppresses CD8 + T cell activation, intratumoral infiltration, and the anti-tumor response. Inhibiting PMVK or GABAAR in HCC mouse models overcomes resistance to anti-PD-1 immune checkpoint therapy. These findings reveal non-canonical and cooperative functions among the key metabolic enzymes PMVK, GAD1, and ACAT1 that reprogram glutamine metabolism to synthesize a potent CD8 + T cell inhibitor 4-Ac-GABA. Blocking 4-Ac-GABA signaling in CD8 + T cells, particularly when combined with immune checkpoint inhibition, potentially represents a new and potent form of immunotherapy., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

The "Mlx" and "Myc" transcription factor networks cross-communicate and share many common gene targets. Myc's activity depends upon its heterodimerization with Max, whereas the Mlx Network requires that the Max-like factor Mlx associate with the Myc-like factors MondoA or ChREBP. The current work demonstrates that body-wide Mlx inactivation, like that of Myc, accelerates numerous aging-related phenotypes pertaining to body habitus and metabolism. The deregulation of numerous aging-related Myc target gene sets is also accelerated. Among other functions, these gene sets often regulate ribosomal and mitochondrial structure and function, genomic stability, and aging. Whereas "MycKO" mice have an extended lifespan because of a lower cancer incidence, "MlxKO" mice have normal lifespans and a higher cancer incidence. Like Myc, the expression of Mlx, MondoA, and ChREBP and their control over their target genes deteriorate with age in both mice and humans. Collectively, these findings underscore the importance of lifelong and balanced cross-talk between the two networks to maintain proper function and regulation of the many factors that can affect normal aging., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

We have previously reported a failure of recovery of synaptic function in the CA1 region of acute hippocampal slices from mice with a conditional neuronal knockout (KO) of GLT-1 (EAAT2, Slc1A2) driven by synapsin-Cre (synGLT-1 KO). The failure of recovery of synaptic function is due to excitotoxic injury. We hypothesized that changes in mitochondrial metabolism contribute to the heightened vulnerability to excitotoxicity in the synGLT-1 KO mice. We found impaired flux of carbon from 13 C-glucose into the tricarboxylic acid cycle in synGLT-1 KO cortical and hippocampal slices compared with wild-type (WT) slices. In addition, we found downregulation of the neuronal glucose transporter GLUT3 in both genotypes. Flux of carbon from [1,2- 13 C]acetate, thought to be astrocyte-specific, was increased in the synGLT-KO hippocampal slices but not cortical slices. Glycogen stores, predominantly localized to astrocytes, are rapidly depleted in slices after cutting, and are replenished during ex vivo incubation. In the synGLT-1 KO, replenishment of glycogen stores during ex vivo incubation was compromised. These results suggest both neuronal and astrocytic metabolic perturbations in the synGLT-1 KO slices. Supplementing incubation medium during recovery with 20 mM D-glucose normalized glycogen replenishment but had no effect on recovery of synaptic function. In contrast, 20 mM non-metabolizable L-glucose substantially improved recovery of synaptic function, suggesting that D-glucose metabolism contributes to the excitotoxic injury in the synGLT-1 KO slices. L-lactate substitution for D-glucose did not promote recovery of synaptic function, implicating mitochondrial metabolism. Consistent with this hypothesis, phosphorylation of pyruvate dehydrogenase, which decreases enzyme activity, was increased in WT slices during the recovery period, but not in synGLT-1 KO slices. Since metabolism of glucose by the mitochondrial electron transport chain is associated with superoxide production, we tested the effect of drugs that scavenge and prevent superoxide production. The superoxide dismutase/catalase mimic EUK-134 conferred complete protection and full recovery of synaptic function. A site-specific inhibitor of complex III superoxide production, S3QEL-2, was also protective, but inhibitors of NADPH oxidase were not. In summary, we find that the failure of recovery of synaptic function in hippocampal slices from the synGLT-1 KO mouse, previously shown to be due to excitotoxic injury, is caused by production of superoxide by mitochondrial metabolism., (© 2024 International Society for Neurochemistry.)

The MYC oncoprotein regulates numerous genes involved in cellular processes such as cell cycle and mitochondrial and ribosomal structure and function. This requires heterodimerization with its partner, MAX, and binding to specific promoter and enhancer elements. Here, we show that MYC and MAX also bind near transcriptional end sites (TESs) of over one-sixth of all annotated genes. These interactions are dose-dependent, evolutionarily conserved, stabilize the normally short-lived MYC protein and regulate expression both in concert with and independent of MYC's binding elsewhere. MYC's TES binding occurs in association with other transcription factors, alters the chromatin landscape, increases nuclease susceptibility and can alter transcriptional read-through, particularly in response to certain stresses. MYC-bound TESs can directly contact promoters and may fine-tune gene expression in response to both physiologic and pathologic stimuli. Collectively, these findings support a previously unrecognized role for MYC in regulating transcription and its read-through via direct intragenic contacts between TESs and promoters.

Tumor cells reprogram their metabolism to produce specialized metabolites that both fuel their own growth and license tumor immune evasion. However, the relationships between these functions remain poorly understood. Here, we report CRISPR screens in a mouse model of colo-rectal cancer (CRC) that implicates the dual specificity phosphatase 18 (DUSP18) in the establishment of tumor-directed immune evasion. Dusp18 inhibition reduces CRC growth rates, which correlate with high levels of CD8 + T cell activation. Mechanistically, DUSP18 dephosphorylates and stabilizes the USF1 bHLH-ZIP transcription factor. In turn, USF1 induces the SREBF2 gene, which allows cells to accumulate the cholesterol biosynthesis intermediate lanosterol and release it into the tumor microenvironment (TME). There, lanosterol uptake by CD8 + T cells suppresses the mevalonate pathway and reduces KRAS protein prenylation and function, which in turn inhibits their activation and establishes a molecular basis for tumor cell immune escape. Finally, the combination of an anti-PD-1 antibody and Lumacaftor, an FDA-approved small molecule inhibitor of DUSP18, inhibits CRC growth in mice and synergistically enhances anti-tumor immunity. Collectively, our findings support the idea that a combination of immune checkpoint and metabolic blockade represents a rationally-designed, mechanistically-based and potential therapy for CRC., (© 2024. The Author(s).)