An influenza virus-triggered SUMO switch orchestrates co-opted endogenous retroviruses to stimulate host antiviral immunity

Dynamic small ubiquitin-like modifier (SUMO) linkages to diverse cellular protein groups are critical to orchestrate resolution of stresses such as genome damage, hypoxia, or proteotoxicity. Defense against pathogen insult (often reliant upon host recognition of "non-self" nucleic acids) is also modulated by SUMO, but the underlying mechanisms are incompletely understood. Here, we used quantitative SILAC-based proteomics to survey pan-viral host SUMOylation responses, creating a resource of almost 600 common and unique SUMO remodeling events that are mounted during influenza A and B virus infections, as well as during viral innate immune stimulation. Subsequent mechanistic profiling focused on a common infection-induced loss of the SUMO-modified form of TRIM28/KAP1, a host transcriptional repressor. By integrating knockout and reconstitution models with system-wide transcriptomics, we provide evidence that influenza virus-triggered loss of SUMO-modified TRIM28 leads to derepression of endogenous retroviral (ERV) elements, unmasking this cellular source of "self" double-stranded (ds)RNA. Consequently, loss of SUMO-modified TRIM28 potentiates canonical cytosolic dsRNA-activated IFN-mediated defenses that rely on RIG-I, MAVS, TBK1, and JAK1. Intriguingly, although wild-type influenza A virus robustly triggers this SUMO switch in TRIM28, the induction of IFN-stimulated genes is limited unless expression of the viral dsRNA-binding protein NS1 is abrogated. This may imply a viral strategy to antagonize such a host response by sequestration of induced immunostimulatory ERV dsRNAs. Overall, our data reveal that a key nuclear mechanism that normally prevents aberrant expression of ERV elements (ERVs) has been functionally co-opted via a stress-induced SUMO switch to augment antiviral immunity.</p

SUMO-targeted ubiquitin E3 ligase RNF4 is required for the response of human cells to DNA damage

Here we demonstrate that RNF4, a highly conserved small ubiquitin-like modifier (SUMO)-targeted ubiquitin E3 ligase, plays a critical role in the response of mammalian cells to DNA damage. Human cells in which RNF4 expression was ablated by siRNA or chicken DT40 cells with a homozygous deletion of the RNF4 gene displayed increased sensitivity to DNA-damaging agents. Recruitment of RNF4 to double-strand breaks required its RING and SUMO interaction motif (SIM) domains and DNA damage factors such as NBS1, mediator of DNA damage checkpoint 1 (MDC1), RNF8, 53BP1, and BRCA1. In the absence of RNF4, these factors were still recruited to sites of DNA damage, but 53BP1, RNF8, and RNF168 displayed delayed clearance from such foci. SILAC-based proteomics of SUMO substrates revealed that MDC1 was SUMO-modified in response to ionizing radiation. As a consequence of SUMO modification, MDC1 recruited RNF4, which mediated ubiquitylation at the DNA damage site. Failure to recruit RNF4 resulted in defective loading of replication protein A (RPA) and Rad51 onto ssDNA. This appeared to be a consequence of reduced recruitment of the CtIP nuclease, resulting in inefficient end resection. Thus, RNF4 is a novel DNA damage-responsive protein that plays a role in homologous recombination and integrates SUMO modification and ubiquitin signaling in the cellular response to genotoxic stress

Sumoylation controls host anti-bacterial response to the gut invasive pathogen <em>Shigella flexneri</em>

International audienceShigella flexneri, the etiological agent of bacillary dysentery, invades the human colonic epithelium and causes its massive inflammatory destruction. Little is known about the post-translational modifications implicated in regulating the host defense pathway against Shigella. Here, we show that SUMO-2 impairs Shigella invasion of epithelial cells in vitro. Using mice haploinsufficient for the SUMO E2 enzyme, we found that sumoylation regulates intestinal permeability and is required to restrict epithelial invasion and control mucosal inflammation. Quantitative proteomics reveals that Shigella infection alters the sumoylation status of a restricted set of transcriptional regulators involved in intestinal functions and inflammation. Consistent with this, sumoylation restricts the pro-inflammatory transcriptional response of Shigella-infected guts. Altogether, our results show that the SUMO pathway is an essential component of host innate protection, as it reduces the efficiency of two key steps of shigellosis: invasion and inflammatory destruction of the intestinal epithelium

System-wide changes to SUMO modifications in response to heat shock

Covalent conjugation of the small ubiquitin-like modifier (SUMO) proteins to target proteins regulates many important eukaryotic cellular mechanisms. Although the molecular consequences of the conjugation of SUMO proteins are relatively well understood, little is known about the cellular signals that regulate the modification of their substrates. Here, we show that SUMO-2 and SUMO-3 are required for cells to survive heat shock. Through quantitative labeling techniques, stringent purification of SUMOylated proteins, advanced mass spectrometric technology, and novel techniques of data analysis, we quantified heat shock-induced changes in the SUMOylation state of 766 putative substrates. In response to heat shock, SUMO was polymerized into polySUMO chains and redistributed among a wide range of proteins involved in cell cycle regulation; apoptosis; the trafficking, folding, and degradation of proteins; transcription; translation; and DNA replication, recombination, and repair. This comprehensive proteomic analysis of the substrates of a ubiquitin-like modifier (Ubl) identifies a pervasive role for SUMO proteins in the biologic response to hyperthermic stress

An engineered IL-2 partial agonist promotes CD8+ T cell stemness

Adoptive transfer of antigen-specific T cells represents a major advance in cancer immunotherapy, with robust clinical outcomes in some patients(1). Both the number of transferred T cells and their differentiation state are critical determinants of effective responses(2,3). T cells can be expanded with T cell receptor (TCR)-mediated stimulation and interleukin-2, but this can lead to differentiation into effector T cells(4,5) and lower therapeutic efficacy(6), whereas maintenance of a more stem-cell-like state before adoptive transfer is beneficial(7). Here we show that H9T, an engineered interleukin-2 partial agonist, promotes the expansion of CD8(+) T cells without driving terminal differentiation. H9T led to altered STAT5 signalling and mediated distinctive downstream transcriptional, epigenetic and metabolic programs. In addition, H9T treatment sustained the expression of T cell transcription factor 1 (TCF-1) and promoted mitochondrial fitness, thereby facilitating the maintenance of a stem-cell-like state. Moreover, TCR-transgenic and chimeric antigen receptor-modified CD8(+) T cells that were expanded with H9T showed robust anti-tumour activity in vivo in mouse models of melanoma and acute lymphoblastic leukaemia. Thus, engineering cytokine variants with distinctive properties is a promising strategy for creating new molecules with translational potential