THAP3 recruits SMYD3 to OXPHOS genes and epigenetically promotes mitochondrial respiration in hepatocellular carcinoma.

Mitochondria harbor the oxidative phosphorylation (OXPHOS) system to sustain cellular respiration. However, the transcriptional regulation of OXPHOS remains largely unexplored. Through the cancer genome atlas (TCGA) transcriptome analysis, transcription factor THAP domain-containing 3 (THAP3) was found to be strongly associated with OXPHOS gene expression. Mechanistically, THAP3 recruited the histone methyltransferase SET and MYND domain-containing protein 3 (SMYD3) to upregulate H3K4me3 and promote OXPHOS gene expression. The levels of THAP3 and SMYD3 were altered by metabolic cues. They collaboratively supported liver cancer cell proliferation and colony formation. In clinical human liver cancer, both of them were overexpressed. THAP3 positively correlated with OXPHOS gene expression. Together, THAP3 cooperates with SMYD3 to epigenetically upregulate cellular respiration and liver cancer cell proliferation.

SUCLG1 restricts POLRMT succinylation to enhance mitochondrial biogenesis and leukemia progression.

Mitochondria are cellular powerhouses that generate energy through the electron transport chain (ETC). The mitochondrial genome (mtDNA) encodes essential ETC proteins in a compartmentalized manner, however, the mechanism underlying metabolic regulation of mtDNA function remains unknown. Here, we report that expression of tricarboxylic acid cycle enzyme succinate-CoA ligase SUCLG1 strongly correlates with ETC genes across various TCGA cancer transcriptomes. Mechanistically, SUCLG1 restricts succinyl-CoA levels to suppress the succinylation of mitochondrial RNA polymerase (POLRMT). Lysine 622 succinylation disrupts the interaction of POLRMT with mtDNA and mitochondrial transcription factors. SUCLG1-mediated POLRMT hyposuccinylation maintains mtDNA transcription, mitochondrial biogenesis, and leukemia cell proliferation. Specifically, leukemia-promoting FMS-like tyrosine kinase 3 (FLT3) mutations modulate nuclear transcription and upregulate SUCLG1 expression to reduce succinyl-CoA and POLRMT succinylation, resulting in enhanced mitobiogenesis. In line, genetic depletion of POLRMT or SUCLG1 significantly delays disease progression in mouse and humanized leukemia models. Importantly, succinyl-CoA level and POLRMT succinylation are downregulated in FLT3-mutated clinical leukemia samples, linking enhanced mitobiogenesis to cancer progression. Together, SUCLG1 connects succinyl-CoA with POLRMT succinylation to modulate mitochondrial function and cancer development.

A carrier-free supramolecular nano-twin-drug for overcoming irinotecan-resistance and enhancing efficacy against colorectal cancer.

Irinotecan (Ir) is commonly employed as a first-line chemotherapeutic treatment for colorectal cancer (CRC). However, tremendous impediments remain to be addressed to surmount drug resistance and ameliorate adverse events. Poly-ADP-Ribose Polymerase (PARP) participates in the maintenance of genome stability and the repair of DNA damage, thus playing a critical role in chemotherapy resistance. In this work, we introduce a novel curative strategy that utilizes nanoparticles (NPs) prepared by dynamic supramolecular co-assembly of Ir and a PARP inhibitor (PARPi) niraparib (Nir) through π-π stacking and hydrogen bond interactions. The Ir and Nir self-assembled Nano-Twin-Drug of (Nir-Ir NPs) could enhance the therapeutic effect on CRC by synergistically inhibiting the DNA damage repair pathway and activating the tumor cell apoptosis process without obvious toxicity. In addition, the Nir-Ir NPs could effectively reverse irinotecan-resistance by inhibiting the expression of multiple resistance protein-1 (MRP-1). Overall, our study underscores the distinctive advantages and potential of Nir-Ir NPs as a complementary strategy to chemotherapy by simultaneously overcoming the Ir resistance and improving the anti-tumor efficacy against CRC.

De Novo PACSIN1 Gene Variant Found in Childhood Lupus and a Role for PACSIN1/TRAF4 Complex in Toll-like Receptor 7 Activation.

OBJECTIVE: Increased Toll-like receptor 7 (TLR-7) signaling leading to the production of type I interferon (IFN) is an important contributor to human systemic lupus erythematosus (SLE). Protein kinase C and casein kinase substrate in neurons 1 (PACSIN1), a molecule that regulates synaptic vesicle recycling, has been linked to TLR-7/TLR-9-mediated type I IFN production in humans and mice, but the underlying mechanism is unknown. We undertook this study to explore the pathogenicity and underlying mechanism of a de novo PACSIN1 missense variant identified in a child with SLE. METHODS: PACSIN1 Q59K de novo and null variants were introduced into a human plasmacytoid dendritic cell line and into mice using CRISPR/Cas9 editing. The effects of the variants on TLR-7/TLR-9 signaling in human and mouse cells, as well as PACSIN1 messenger RNA and IFN signature in SLE patients, were assessed using real-time polymerase chain reaction and flow cytometry. Mechanisms were investigated using luciferase reporter assays, RNA interference, coimmunoprecipitation, and immunofluorescence. RESULTS: We established that PACSIN1 forms a trimolecular complex with tumor necrosis factor receptor-associated factor 4 (TRAF4) and TRAF6 that is important for the regulation of type I IFN. The Q59K mutation in PACSIN1 augments binding to neural Wiskott-Aldrich syndrome protein while it decreases binding to TRAF4, leading to unrestrained TRAF6-mediated activation of type I IFN. Intriguingly, PACSIN1 Q59K increased TLR-7 but not TLR-9 signaling in human cells, leading to elevated expression of IFNß and IFN-inducible genes. Untreated SLE patients had high PACSIN1 expression in peripheral blood cells that correlated positively with IFN-related genes. Introduction of the Pacsin1 Q59K mutation into mice caused increased surface TLR-7 and TRAIL expression in B cells. CONCLUSION: PACSIN1 Q59K increases IFNß activity through the impairment of TRAF4-mediated inhibition of TLR-7 signaling, possibly contributing to SLE risk.

P2RY8 variants in lupus patients uncover a role for the receptor in immunological tolerance.

B cell self-tolerance is maintained through multiple checkpoints, including restraints on intracellular signaling and cell trafficking. P2RY8 is a receptor with established roles in germinal center (GC) B cell migration inhibition and growth regulation. Somatic P2RY8 variants are common in GC-derived B cell lymphomas. Here, we identify germline novel or rare P2RY8 missense variants in lupus kindreds or the related antiphospholipid syndrome, including a "de novo" variant in a child with severe nephritis. All variants decreased protein expression, F-actin abundance, and GPCR-RhoA signaling, and those with stronger effects increased AKT and ERK activity and cell migration. Remarkably, P2RY8 was reduced in B cell subsets from some SLE patients lacking P2RY8 gene variants. Low P2RY8 correlated with lupus nephritis and increased age-associated B cells and plasma cells. By contrast, P2RY8 overexpression in cells and mice restrained plasma cell development and reinforced negative selection of DNA-reactive developing B cells. These findings uncover a role of P2RY8 in immunological tolerance and lupus pathogenesis.

An antibody-drug conjugate targeting a GSTA glycosite-signature epitope of MUC1 expressed by non-small cell lung cancer.

Antibodies targeting aberrantly glycosylated proteins are ineffective in treating cancer. Antibody-drug conjugates have emerged as effective alternatives, facilitating tumor-specific drug delivery. Previous studies have assessed the aberrantly glycosylated tandem repeat region of MUC1 glycoprotein as three site-specific glycosylated neoantigen peptide motifs (PDTR, GSTA, and GVTS) for binding with a monoclonal antibody. This study aimed to develop an antibody-drug conjugate for cancer treatment based on monoclonal antibodies against the aforementioned three neoantigen peptide motifs. Internalization of monoclonal antibodies was assessed via immunofluorescence staining and colocalization with lysosomal markers in live cells. Antibody positivity in tumor and peritumoral tissue samples was assessed via immunohistochemistry. The efficacy of anti-MUC1 ADCs was evaluated using various cancer cell lines and a mouse tumor xenograft model. An anti-MUC1 ADC was synthesized by conjugating GSTA neoantigen-specific 16A with monomethyl auristatin E (MMAE), which displayed potent antitumoral efficacy with an IC50 ranging 0.2-49.4 nM toward various cancer cells. In vivo, 16A-MMAE inhibited tumor growth in a dose-dependent manner in a mouse xenograft model established using the NCI-H838 NSCLC cell line, at a minimum effective dose of 1 mg/kg. At 3 mg/kg, 16A-MMAE did not cause significant toxicity in a transgenic mouse expressing human MUC1. The high antitumoral efficacy of 16A-MMAE suggests that aberrant glycosylated MUC1 neoantigen is a potential target for the development of ADCs for treating various cancers. Personalized therapy may be achieved through such glycosite-specific ADCs.

Genomic test ends a long diagnostic odyssey in a patient with resistance to thyroid hormones.

BACKGROUND: Resistance to thyroid hormones is a very rare condition, which is often misdiagnosed and mistreated. The cases where there is a concomitant autoimmune thyroid disorder are ultra-rare and particularly challenging to treat. Diagnostic and research-based genomic testing can sometimes identify pathogenic variants unrelated to the primary reason for testing (incidental findings). CASE PRESENTATION: We present a patient with thyroid resistance associated with hypothyroid Hashimoto thyroiditis. The long diagnostic odyssey spanning over 20-years included repeated misdiagnoses and mistreatments and was concluded by a research-based genomic testing, identifying a "de novo" THRB pathogenic variant. The varying sensitivity of various tissues to thyroid hormones accompanied by hypothyroid Hashimoto thyroiditis continues to pose a significant treatment challenge. CONCLUSIONS: Thyroid hormone resistance continues to be an un(der)- and misdiagnosed thyroid condition whose management is particularly challenging when associated with autoimmune thyroid disease. Whole exome sequencing has the potential to identify THRB pathogenic variants as incidental findings. Reporting such secondary findings from genomic testing may be particularly important in the context of the rarity of the condition and the potential clinical consequences of misdiagnosis and mistreatment.

Quantitative proteomics analysis reveals alterations of lysine acetylation in mouse testis in response to heat shock and X-ray exposure.

Environmental stresses are important factors causing male infertility which attracts broad attention. Protein acetylation is a pivotal post-translational modification and modulates diverse physiological processes including spermatogenesis. In this study, we employed quantitative proteomic techniques and bioinformatics tools to analyze the alterations of acetylome profile of mouse testis after heat shock and X-irradiation. Overall, we identified 1139 lysine acetylation sites in 587 proteins in which 1020 lysine acetylation sites were quantified. The Gene Ontology analysis showed that the major acetylated protein groups were involved in generation of precursor metabolites and metabolic processes, and were localized predominantly in cytosolic and mitochondrial. Compared to the control group, 36 sites of 28 acetylated proteins have changed after heat shock, and 49 sites of 43 acetylated proteins for X-ray exposure. Some of the differentially acetylated proteins have been reported to be associated with the progression of spermatogenesis and male fertility. We observed the up-regulated acetylation level change on testis specific histone 2B and heat shock protein upon heat treatment and a sharp decline of acetylation level on histone H2AX under X-ray treatment, suggesting their roles in male germ cells. Notably, the acetylation level on K279 of histone acetyltransferase (Kat7) was down-regulated in both heat and X-ray treatments, indicating that K279 may be a key acetylated site and affect its functions in spermatogenesis. Our results reveal that protein acetylation might add another layer of complexity to the regulation for spermatogenesis, and further functional studies of these proteins will help us elucidate the mechanisms of abnormal spermatogenesis.

Germline deletion of huntingtin causes male infertility and arrested spermiogenesis in mice.

Human Huntingtin (HTT), a Huntington's disease gene, is highly expressed in the mammalian brain and testis. Simultaneous knockout of mouse Huntingtin (Htt) in brain and testis impairs male fertility, providing evidence for a link between Htt and spermatogenesis; however, the underlying mechanism remains unclear. To understand better the function of Htt in spermatogenesis, we restricted the genetic deletion specifically to the germ cells using the Cre/loxP site-specific recombination strategy and found that the resulting mice manifested smaller testes, azoospermia and complete male infertility. Meiotic chromosome spread experiments showed that the process of meiosis was normal in the absence of Htt. Notably, we found that Htt-deficient round spermatids did not progress beyond step 3 during the post-meiotic phase, when round spermatids differentiate into mature spermatozoa. Using an iTRAQ-based quantitative proteomic assay, we found that knockout of Htt significantly altered the testis protein profile. The differentially expressed proteins exhibited a remarkable enrichment for proteins involved in translation regulation and DNA packaging, suggesting that Htt might play a role in spermatogenesis by regulating translation and DNA packaging in the testis.