The role of N-terminal phosphorylation of DGK-θ.

Diacylglycerol kinases (DGKs) are lipid kinases that mediate the phosphorylation of diacylglycerol (DAG) leading to the production of phosphatidic acid (PtdOH). To examine the role of phosphorylation on DGK-θ, we first identified the phosphorylated sites on endogenous DGK-θ from mouse brain and found four sites: S15, S17, which we refer to phosphomotif-1 sites, and S22 and S26 which we refer to as phosphomotif-2 sites. This study focused on the role of these phosphorylated sites on enzyme activity, membrane binding, thermal stability, and cellular half-life of DGK-θ.

Identification of Synaptic DGKθ Interactors That Stimulate DGKθ Activity.

Lipids and their metabolic enzymes are a critical point of regulation for the membrane curvature required to induce membrane fusion during synaptic vesicle recycling. One such enzyme is diacylglycerol kinase θ (DGKθ), which produces phosphatidic acid (PtdOH) that generates negative membrane curvature. Synapses lacking DGKθ have significantly slower rates of endocytosis, implicating DGKθ as an endocytic regulator.

Multiubiquitination of TRPV4 reduces channel activity independent of surface localization.

Ubiquitin (Ub)-mediated regulation of plasmalemmal ion channel activity canonically occurs via stimulation of endocytosis. Whether ubiquitination can modulate channel activity by alternative mechanisms remains unknown. Here, we show that the transient receptor potential vanilloid 4 (TRPV4) cation channel is multiubiquitinated within its cytosolic N-terminal and C-terminal intrinsically disordered regions (IDRs).

Human Breast Milk Enhances Intestinal Mucosal Barrier Function and Innate Immunity in a Healthy Pediatric Human Enteroid Model.

Breastfeeding has been associated with long lasting health benefits. Nutrients and bioactive components of human breast milk promote cell growth, immune development, and shield the infant gut from insults and microbial threats. The molecular and cellular events involved in these processes are ill defined.

Diverse mitochondrial abnormalities in a new cellular model of TAFFAZZIN deficiency are remediated by cardiolipin-interacting small molecules.

Barth syndrome (BTHS) is an X-linked disorder of mitochondrial phospholipid metabolism caused by pathogenic variants in TAFFAZIN, which results in abnormal cardiolipin (CL) content in the inner mitochondrial membrane. To identify unappreciated pathways of mitochondrial dysfunction in BTHS, we utilized an unbiased proteomics strategy and identified that complex I (CI) of the mitochondrial respiratory chain and the mitochondrial quality control protease presenilin-associated rhomboid-like protein (PARL) are altered in a new HEK293-based tafazzin-deficiency model. Follow-up studies confirmed decreased steady state levels of specific CI subunits and an assembly factor in the absence of tafazzin; this decrease is in part based on decreased transcription and results in reduced CI assembly and function.

Immortalized striatal precursor neurons from Huntington's disease patient-derived iPS cells as a platform for target identification and screening for experimental therapeutics.

We have previously established induced pluripotent stem cell (iPSC) models of Huntington's disease (HD), demonstrating CAG-repeat-expansion-dependent cell biological changes and toxicity. However, the current differentiation protocols are cumbersome and time consuming, making preparation of large quantities of cells for biochemical or screening assays difficult. Here, we report the generation of immortalized striatal precursor neurons (ISPNs) with normal (33) and expanded (180) CAG repeats from HD iPSCs, differentiated to a phenotype resembling medium spiny neurons (MSN), as a proof of principle for a more tractable patient-derived cell model.

C9orf72-associated SMCR8 protein binds in the ubiquitin pathway and with proteins linked with neurological disease.

A pathogenic GGGCCC hexanucleotide expansion in the first intron/promoter region of the C9orf72 gene is the most common mutation associated with amyotrophic lateral sclerosis (ALS). The C9orf72 gene product forms a complex with SMCR8 (Smith-Magenis Syndrome Chromosome Region, Candidate 8) and WDR41 (WD Repeat domain 41) proteins. Recent studies have indicated roles for the complex in autophagy regulation, vesicle trafficking, and immune response in transgenic mice, however a direct connection with ALS etiology remains unclear.

Stimulus-dependent modifications in astrocyte-derived extracellular vesicle cargo regulate neuronal excitability.

Extracellular vesicles have now emerged as key players in cell-to-cell communication. This is particularly important in the central nervous system, where glia-neuron cross-talk helps maintain normal neuronal function. Astrocyte-derived extracellular vesicles (ADEVs) secreted constitutively promote neurite outgrowth and neuronal survival.

Synthetic Circular RNA Functions as a miR-21 Sponge to Suppress Gastric Carcinoma Cell Proliferation.

MicroRNA (miR) sponges containing miR binding sequences constitute a potentially powerful molecular therapeutic strategy. Recently, naturally occurring circular RNAs (circRNAs) were shown to function as efficient miR sponges in cancer cells. We hypothesized that synthetic circRNA sponges could achieve therapeutic loss-of-function targeted against specific miRs.

The plasma proteome identifies expected and novel proteins correlated with micronutrient status in undernourished Nepalese children.

Micronutrient deficiencies are common in undernourished societies yet remain inadequately assessed due to the complexity and costs of existing assays. A plasma proteomics-based approach holds promise in quantifying multiple nutrient:protein associations that reflect biological function and nutritional status. To validate this concept, in plasma samples of a cohort of 500 6- to 8-y-old Nepalese children, we estimated cross-sectional correlations between vitamins A (retinol), D (25-hydroxyvitamin D), and E (α-tocopherol), copper, and selenium, measured by conventional assays, and relative abundance of their major plasma-bound proteins, measured by quantitative proteomics using 8-plex iTRAQ mass tags.

The cardiac acetyl-lysine proteome.

In the heart, lysine acetylation has been implicated in processes ranging from transcriptional control of pathological remodeling, to cardioprotection arising from caloric restriction. Given the emerging importance of this post-translational modification, we used a proteomic approach to investigate the broader role of lysine acetylation in the heart using a guinea pig model. Briefly, hearts were fractionated into myofilament-, mitochondrial- and cytosol-enriched fractions prior to proteolysis and affinity-enrichment of acetylated peptides.