Nuclear protein in testis (Nut) is a universal oncogenic driver in the highly aggressive NUT midline carcinoma, whose physiological function in male germ cells has been unclear. Here we show that expression of Nut is normally restricted to post-meiotic spermatogenic cells, where its presence triggers p300-dependent genome-wide histone H4 hyperacetylation, which is essential for the completion of histone-to-protamine exchange. Accordingly, the inactivation of Nut induces male sterility with spermatogenesis arrest at the histone-removal stage. Nut uses p300 and/or CBP to enhance acetylation of H4 at both K5 and K8, providing binding sites for the first bromodomain of Brdt, the testis-specific member of the BET family, which subsequently mediates genome-wide histone removal. Altogether, our data reveal the detailed molecular basis of the global histone hyperacetylation wave, which occurs before the final compaction of the male genome.

BackgroundTumor orchestrated metabolic changes in the microenvironment limit generation of anti-tumor immune responses. Availability of arginine, a semi-essential amino acid, is critical for lymphocyte proliferation and function. Levels of arginine are regulated by the enzymes arginase 1,2 and nitric oxide synthase (NOS). However, the role of arginase activity in lung tumor maintenance has not been investigated in clinically relevant orthotopic tumor models.MethodsRNA sequencing (RNA-seq) of sorted cell populations from mouse lung adenocarcinomas derived from immunocompetent genetically engineered mouse models (GEMM)s was performed. To complement mouse studies, a patient tissue microarray consisting of 150 lung adenocarcinomas, 103 squamous tumors, and 54 matched normal tissue were stained for arginase, CD3, and CD66b by multiplex immunohistochemistry. Efficacy of a novel arginase inhibitor compound 9 in reversing arginase mediated T cell suppression was determined in splenocyte ex vivo assays. Additionally, the anti-tumor activity of this compound was determined in vitro and in an autochthonous immunocompetent KrasG12D GEMM of lung adenocarcinoma model.ResultsAnalysis of RNA-seq of sorted myeloid cells suggested that arginase expression is elevated in myeloid cells in the tumor as compared to the normal lung tissue. Accordingly, in the patient samples arginase 1 expression was mainly localized in the granulocytic myeloid cells and significantly elevated in both lung adenocarcinoma and squamous tumors as compared to the controls. Our ex vivo analysis demonstrated that myeloid derived suppressor cell (MDSC)s cause T cell suppression by arginine depletion, and suppression of arginase activity by a novel ARG1/2 inhibitor, compound 9, led to restoration of T cell function by increasing arginine. Treatment of KrasG12D GEMM of lung cancer model with compound 9 led to a significant tumor regression associated with increased T cell numbers and function, while it had no activity across several murine and human non-small cell (NSCLC) lung cancer lines in vitro.ConclusionsWe show that arginase expression is elevated in mouse and patient lung tumors. In a KRASG12D GEMM arginase inhibition diminished growth of established tumors. Our data suggest arginase as an immunomodulatory target that should further be investigated in lung tumors with high arginase activity.Electronic supplementary materialThe online version of this article (10.1186/s40425-019-0504-5) contains supplementary material, which is available to authorized users.

Ubiquitin-specific protease 14 (USP14) is one of the major proteasome-associated deubiquitinating enzymes critical for proteome homeostasis. However, substrates of USP14 remain largely unknown, hindering the understanding of its functional roles. Here we conduct a comprehensive proteome, ubiquitinome and interactome analysis for USP14 substrate screening. Bioinformatics analysis reveals broad new potential roles of USP14, especially in lipid and carbohydrate metabolism. Among the potential substrates identified, we show that fatty acid synthase (FASN), a key enzyme involved in hepatic lipogenesis, is a bona fide substrate of USP14. USP14 directly interacts with and increases FASN stability. As a result, overexpression of USP14 promotes liver triglyceride accumulation in C57BL/6 mice, whereas genetic ablation or pharmacological inhibition of USP14 ameliorates hepatosteatosis, hyperglycemia and insulin resistance in obese mice. In conclusion, our findings reveal for the first time an indispensable role of USP14 in hepatosteatosis through FASN stabilization.

Type 2 diabetes (T2D) is a major chronic healthcare concern worldwide. Emerging evidence suggests that a histone-modification-mediated epigenetic mechanism underlies T2D. Nevertheless, the dynamics of histone marks in T2D have not yet been carefully analyzed. Using a mass spectrometry-based label-free and chemical stable isotope labeling quantitative proteomic approach, we systematically profiled liver histone post-translational modifications (PTMs) in a prediabetic high-fat diet-induced obese (DIO) mouse model. We identified 170 histone marks, 30 of which were previously unknown. Interestingly, about 30% of the histone marks identified in DIO mouse liver belonged to a set of recently reported lysine acylation modifications, including propionylation, butyrylation, malonylation, and succinylation, suggesting possible roles of these newly identified histone acylations in diabetes and obesity. These histone marks were detected without prior affinity enrichment with an antibody, demonstrating that the histone acylation marks are present at reasonably high stoichiometry. Fifteen histone marks differed in abundance in DIO mouse liver compared with liver from chow-fed mice in label-free quantification, and six histone marks in stable isotope labeling quantification. Analysis of hepatic histone modifications from metformin-treated DIO mice revealed that metformin, a drug widely used for T2D, could reverse DIO-stimulated histone H3K36me2 in prediabetes, suggesting that this mark is likely associated with T2D development. Our study thus offers a comprehensive landscape of histone marks in a prediabetic mouse model, provides a resource for studying epigenetic functions of histone modifications in obesity and T2D, and suggest a new epigenetic mechanism for the physiological function of metformin.