Astrocytic response mediated by the CLU risk allele inhibits OPC proliferation and myelination in a human iPSC model.

The C allele of rs11136000 variant in the clusterin (CLU) gene represents the third strongest known genetic risk factor for late-onset Alzheimer's disease. However, whether this single-nucleotide polymorphism (SNP) is functional and what the underlying mechanisms are remain unclear. In this study, the CLU rs11136000 SNP is identified as a functional variant by a small-scale CRISPR-Cas9 screen. Astrocytes derived from isogenic induced pluripotent stem cells (iPSCs) carrying the "C" or "T" allele of the CLU rs11136000 SNP exhibit different CLU expression levels. TAR DNA-binding protein-43 (TDP-43) preferentially binds to the "C" allele to promote CLU expression and exacerbate inflammation. The interferon response and CXCL10 expression are elevated in cytokine-treated C/C astrocytes, leading to inhibition of oligodendrocyte progenitor cell (OPC) proliferation and myelination. Accordingly, elevated CLU and CXCL10 but reduced myelin basic protein (MBP) expression are detected in human brains of C/C carriers. Our study uncovers a mechanism underlying reduced white matter integrity observed in the CLU rs11136000 risk "C" allele carriers.

Long noncoding RNA LEENE promotes angiogenesis and ischemic recovery in diabetes models.

Impaired angiogenesis in diabetes is a key process contributing to ischemic diseases such as peripheral arterial disease. Epigenetic mechanisms, including those mediated by long noncoding RNAs (lncRNAs), are crucial links connecting diabetes and the related chronic tissue ischemia. Here we identify the lncRNA that enhances endothelial nitric oxide synthase (eNOS) expression (LEENE) as a regulator of angiogenesis and ischemic response. LEENE expression was decreased in diabetic conditions in cultured endothelial cells (ECs), mouse hind limb muscles, and human arteries. Inhibition of LEENE in human microvascular ECs reduced their angiogenic capacity with a dysregulated angiogenic gene program. Diabetic mice deficient in Leene demonstrated impaired angiogenesis and perfusion following hind limb ischemia. Importantly, overexpression of human LEENE rescued the impaired ischemic response in Leene-knockout mice at tissue functional and single-cell transcriptomic levels. Mechanistically, LEENE RNA promoted transcription of proangiogenic genes in ECs, such as KDR (encoding VEGFR2) and NOS3 (encoding eNOS), potentially by interacting with LEO1, a key component of the RNA polymerase II-associated factor complex and MYC, a crucial transcription factor for angiogenesis. Taken together, our findings demonstrate an essential role for LEENE in the regulation of angiogenesis and tissue perfusion. Functional enhancement of LEENE to restore angiogenesis for tissue repair and regeneration may represent a potential strategy to tackle ischemic vascular diseases.

Targeting PUS7 suppresses tRNA pseudouridylation and glioblastoma tumorigenesis.

Pseudouridine is the most frequent epitranscriptomic modification. However, its cellular functions remain largely unknown. Here, we show that pseudouridine synthase 7 (PUS7) is highly expressed in glioblastoma versus normal brain tissues, and high PUS7 expression levels are associated with worse survival in patients with glioblastoma. PUS7 expression and catalytic activity are required for glioblastoma stem cell (GSC) tumorigenesis. Mechanistically, we identify PUS7 targets in GSCs through small RNA pseudouridine sequencing and show that pseudouridylation of PUS7-regulated transfer RNA is critical for codon-specific translational control of key regulators of GSCs. Moreover, we identify chemical inhibitors for PUS7 and show that these compounds prevent PUS7-mediated pseudouridine modification, suppress tumorigenesis and extend the life span of tumor-bearing mice. Overall, we identify an epitranscriptomic regulatory mechanism in glioblastoma and provide preclinical evidence of a potential therapeutic strategy for glioblastoma.

Discovery of a bacterial peptide as a modulator of GLP-1 and metabolic disease.

Early work in rodents highlighted the gut microbiota's importance in metabolic disease, including Type II Diabetes Mellitus (T2DM) and obesity. Glucagon-like peptide-1 (GLP-1), an incretin secreted by L-cells lining the gastrointestinal epithelium, has important functions: promoting insulin secretion, insulin sensitivity, and ß-cell mass, while inhibiting gastric emptying and appetite. We set out to identify microbial strains with GLP-1 stimulatory activity as potential metabolic disease therapeutics. Over 1500 human-derived strains were isolated from healthy individuals and screened for GLP-1 modulation by incubating bacterial cell-free supernatants with NCI H716 L-cells. Approximately 45 strains capable of increasing GLP-1 were discovered. All GLP-1 positive strains were identified as Staphylococcus epidermidis by 16S rRNA sequencing. Mass spectrometry analysis identified a 3 kDa peptide, Hld (delta-toxin), present in GLP-1 positive supernatants but absent in GLP-1 neutral supernatants. Studies in NCI-H716 cells and human jejunal enteroids engineered to make more enteroendocrine cells demonstrated that Hld alone is sufficient to enhance GLP-1 secretion. When administered in high-fat-fed mice, Hld-producing S. epidermidis significantly reduced markers associated with obesity and T2DM. Further characterization of Hld suggests GLP-1 stimulatory action of Hld occurs via calcium signaling. The presented results identify a novel host-microbe interaction which may ultimately lead to the development of a microbial peptide-based therapeutic for metabolic disease.

FolC2-mediated folate metabolism contributes to suppression of inflammation by probiotic Lactobacillus reuteri.

Bacterial-derived compounds from the intestinal microbiome modulate host mucosal immunity. Identification and mechanistic studies of these compounds provide insights into host-microbial mutualism. Specific Lactobacillus reuteri strains suppress production of the proinflammatory cytokine, tumor necrosis factor (TNF), and are protective in a mouse model of colitis. Human-derived L. reuteri strain ATCC PTA 6475 suppresses intestinal inflammation and produces 5,10-methenyltetrahydrofolic acid polyglutamates. Insertional mutagenesis identified the bifunctional dihydrofolate synthase/folylpolyglutamate synthase type 2 (folC2) gene as essential for 5,10-methenyltetrahydrofolic acid polyglutamate biosynthesis, as well as for suppression of TNF production by activated human monocytes, and for the anti-inflammatory effect of L. reuteri 6475 in a trinitrobenzene sulfonic acid-induced mouse model of acute colitis. In contrast, folC encodes the enzyme responsible for folate polyglutamylation but does not impact TNF suppression by L. reuteri. Comparative transcriptomics between wild-type and mutant L. reuteri strains revealed additional genes involved in immunomodulation, including previously identified hdc genes involved in histidine to histamine conversion. The folC2 mutant yielded diminished hdc gene cluster expression and diminished histamine production, suggesting a link between folate and histadine/histamine metabolism. The identification of genes and gene networks regulating production of bacterial-derived immunoregulatory molecules may lead to improved anti-inflammatory strategies for digestive diseases.