Characterization of nucleolar SUMO isopeptidases unveils a general p53-independent checkpoint of impaired ribosome biogenesis.

Ribosome biogenesis is a multi-step process, in which a network of trans-acting factors ensures the coordinated assembly of pre-ribosomal particles in order to generate functional ribosomes. Ribosome biogenesis is tightly coordinated with cell proliferation and its perturbation activates a p53-dependent cell-cycle checkpoint. How p53-independent signalling networks connect impaired ribosome biogenesis to the cell-cycle machinery has remained largely enigmatic. We demonstrate that inactivation of the nucleolar SUMO isopeptidases SENP3 and SENP5 disturbs distinct steps of 40S and 60S ribosomal subunit assembly pathways, thereby triggering the canonical p53-dependent impaired ribosome biogenesis checkpoint. However, inactivation of SENP3 or SENP5 also induces a p53-independent checkpoint that converges on the specific downregulation of the key cell-cycle regulator CDK6. We further reveal that impaired ribosome biogenesis generally triggers the downregulation of CDK6, independent of the cellular p53 status. Altogether, these data define the role of SUMO signalling in ribosome biogenesis and unveil a p53-independent checkpoint of impaired ribosome biogenesis.

Small-molecule SUMO inhibition for biomarker-informed B-cell lymphoma therapy.

Aberrant activity of the SUMOylation pathway has been associated with MYC overexpression and poor prognosis in aggressive B-cell lymphoma (BCL) and other malignancies. Recently developed small-molecule inhibitors of SUMOylation (SUMOi) target the heterodimeric E1 SUMO activation complex (SAE1/UBA2). Here, we report that activated MYC signaling is an actionable molecular vulnerability in vitro and in a preclinical murine in vivo model of MYC-driven BCL. While SUMOi conferred direct effects on MYC-driven lymphoma cells, SUMO inhibition also resulted in substantial remodeling of various subsets of the innate and specific immunity in vivo. Specifically, SUMOi increased the number of memory B cells as well as cytotoxic and memory T cells, subsets that are attributed a key role within a coordinated anti-tumor immune response. In summary, our data constitute pharmacologic SUMOi as a powerful therapy in a subset of BCL causing massive remodeling of the normal B-cell and T-cell compartment.

Activated SUMOylation restricts MHC class I antigen presentation to confer immune evasion in cancer.

Activated SUMOylation is a hallmark of cancer. Starting from a targeted screening for SUMO-regulated immune evasion mechanisms, we identified an evolutionarily conserved function of activated SUMOylation, which attenuated the immunogenicity of tumor cells. Activated SUMOylation allowed cancer cells to evade CD8+ T cell-mediated immunosurveillance by suppressing the MHC class I (MHC-I) antigen-processing and presentation machinery (APM). Loss of the MHC-I APM is a frequent cause of resistance to cancer immunotherapies, and the pharmacological inhibition of SUMOylation (SUMOi) resulted in reduced activity of the transcriptional repressor scaffold attachment factor B (SAFB) and induction of the MHC-I APM. Consequently, SUMOi enhanced the presentation of antigens and the susceptibility of tumor cells to CD8+ T cell-mediated killing. Importantly, SUMOi also triggered the activation of CD8+ T cells and thereby drove a feed-forward loop amplifying the specific antitumor immune response. In summary, we showed that activated SUMOylation allowed tumor cells to evade antitumor immunosurveillance, and we have expanded the understanding of SUMOi as a rational therapeutic strategy for enhancing the efficacy of cancer immunotherapies.

Formate metabolism in the acetogenic bacterium Acetobacterium woodii.

Acetogenic bacteria are already established as biocatalysts for production of high-value compounds from C1 substrates such as H2  + CO2 or CO. However, little is known about the physiology, biochemistry and bioenergetics of acetogenesis from formate, an interesting feedstock for biorefineries. Here, we analysed formate metabolism in the model acetogen Acetobacterium woodii. Cells grew optimally on 200 mM formate to an optical density of 0.6. Formate was exclusively converted to acetate (and CO2 ) with a ratio of 4.4:1. Transcriptome analyses revealed genes/enzymes involved in formate metabolism. Strikingly, A. woodii has two genes potentially encoding a formyl-THF synthetase, fhs1 and fhs2. fhs2 forms an operon with a gene encoding a potential formate transporter, fdhC. Deletion of fhs2/fdhC led to a reduced growth rate, formate consumption and optical densities. Acetogenesis from H2  + CO2 was accompanied by transient formate production; strikingly, formate reutilization was completely abolished in the Δfhs2/fdhC mutant. Take together, our studies gave the first detailed insights into the formatotrophic lifestyle of A. woodii.

Energy conservation involving 2 respiratory circuits.

Chemiosmosis and substrate-level phosphorylation are the 2 mechanisms employed to form the biological energy currency adenosine triphosphate (ATP). During chemiosmosis, a transmembrane electrochemical ion gradient is harnessed by a rotary ATP synthase to phosphorylate adenosine diphosphate to ATP. In microorganisms, this ion gradient is usually composed of [Formula: see text], but it can also be composed of Na+ Here, we show that the strictly anaerobic rumen bacterium Pseudobutyrivibrio ruminis possesses 2 ATP synthases and 2 distinct respiratory enzymes, the ferredoxin:[Formula: see text] oxidoreductase (Rnf complex) and the energy-converting hydrogenase (Ech complex). In silico analyses revealed that 1 ATP synthase is [Formula: see text]-dependent and the other Na+-dependent, which was validated by biochemical analyses. Rnf and Ech activity was also biochemically identified and investigated in membranes of P. ruminis Furthermore, the physiology of the rumen bacterium and the role of the energy-conserving systems was investigated in dependence of 2 different catabolic pathways (the Embden-Meyerhof-Parnas or the pentose-phosphate pathway) and in dependence of Na+ availability. Growth of P. ruminis was greatly stimulated by Na+, and a combination of physiological, biochemical, and transcriptional analyses revealed the role of the energy conserving systems in P. ruminis under different metabolic scenarios. These data demonstrate the use of a 2-component ion circuit for [Formula: see text] bioenergetics and a 2nd 2-component ion circuit for Na+ bioenergetics in a strictly anaerobic rumen bacterium. In silico analyses infer that these 2 circuits are prevalent in a number of other strictly anaerobic microorganisms.

Alanine, a Novel Growth Substrate for the Acetogenic Bacterium Acetobacterium woodii.

Acetogenic bacteria are an ecophysiologically important group of strictly anaerobic bacteria that grow lithotrophically on H2 plus CO2 or on CO or heterotrophically on different substrates such as sugars, alcohols, aldehydes, or acids. Amino acids are rarely used. Here, we describe that the model acetogen Acetobacterium woodii can use alanine as the sole carbon and energy source, which is in contrast to the description of the type strain. The alanine degradation genes have been identified and characterized. A key to alanine degradation is an alanine dehydrogenase which has been characterized biochemically. The resulting pyruvate is further degraded to acetate by the known pathways involving the Wood-Ljungdahl pathway. Our studies culminate in a metabolic and bioenergetic scheme for alanine-dependent acetogenesis in A. woodiiIMPORTANCE Peptides and amino acids are widespread in nature, but there are only a few reports that demonstrated use of amino acids as carbon and energy sources by acetogenic bacteria, a central and important group in the anaerobic food web. Our finding that A. woodii can perform alanine oxidation coupled to reduction of carbon dioxide not only increases the number of substrates that can be used by this model acetogen but also raises the possibility that other acetogens may also be able to use alanine. Indeed, the alanine genes are also present in at least two more acetogens, for which growth on alanine has not been reported so far. Alanine may be a promising substrate for industrial fermentations, since acid formation goes along with the production of a base (NH3) and pH regulation is a minor issue.