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Tumor necrosis factor receptor (TNFR)-associated factors (TRAFs) are a family of structurally related proteins that transduces signals from members of TNFR superfamily and various other immune receptors. Major downstream signaling events mediated by the TRAF molecules include activation of the transcription factor nuclear factor κB (NF-κB) and the mitogen-activated protein kinases (MAPKs). In addition, some TRAF family members, particularly TRAF2 and TRAF3, serve as negative regulators of specific signaling pathways, such as the noncanonical NF-κB and proinflammatory toll-like receptor pathways. Thus, TRAFs possess important and complex signaling functions in the immune system and play an important role in regulating immune and inflammatory responses. This review will focus on the role of TRAF proteins in the regulation of NF-κB and MAPK signaling pathways.

The galectin 3 binding protein (LGALS3BP, also known as 90K) is a ubiquitous multifunctional secreted glycoprotein originally identified in cancer progression. It remains unclear how 90K functions in innate immunity during viral infections. In this study, we found that viral infections resulted in elevated levels of 90K. Further studies demonstrated that 90K expression suppressed virus replication by inducing IFN and pro-inflammatory cytokine production. Upon investigating the mechanisms behind this event, we found that 90K functions as a scaffold/adaptor protein to interact with TRAF6, TRAF3, TAK1 and TBK1. Furthermore, 90K enhanced TRAF6 and TRAF3 ubiquitination and served as a specific ubiquitination substrate of TRAF6, leading to transcription factor NF-κB, IRF3 and IRF7 translocation from the cytoplasm to the nucleus. Conclusions: 90K is a virus-induced protein capable of binding with the TRAF6 and TRAF3 complex, leading to IFN and pro-inflammatory production.

The general transcription factor IID (TFIID) plays a central role in the initiation of RNA polymerase II (Pol II)-dependent transcription by nucleating pre-initiation complex (PIC) assembly at the core promoter. TFIID comprises the TATA-binding protein (TBP) and 13 TBP-associated factors (TAF1–13), which specifically interact with a variety of core promoter DNA sequences. Here we present the structure of human TFIID in complex with TFIIA and core promoter DNA, determined by single-particle cryo-electron microscopy at sub-nanometre resolution. All core promoter elements are contacted by subunits of TFIID, with TAF1 and TAF2 mediating major interactions with the downstream promoter. TFIIA bridges the TBP–TATA complex with lobe B of TFIID. We also present the cryo-electron microscopy reconstruction of a fully assembled human TAF-less PIC. Superposition of common elements between the two structures provides novel insights into the general role of TFIID in promoter recognition, PIC assembly, and transcription initiation. A sub-nanometre resolution cryo-EM structure of human TFIID bound to TFIIA and core promoter DNA and a model of the TFIID-based pre-initiation complex. Promoter-bound TFIID structure This paper describes the interaction of the human general transcription factor IID (TFIID) with core promoter DNA. TFIID has a central role in the initiation of RNA polymerase II dependent transcription by nucleating assembly of the pre-initiation complex (PIC) at the promoter. Eva Nogales and colleagues determine a sub-nanometre resolution cryo-electron microscopy structure of TFIID bound to TFIIA and core promoter DNA. They also present a cryo-electron microscopy reconstruction of a fully assembled human PIC lacking TBP-associated factors (TAFs). By superimposing common elements between the TFIID–TFIIA–promoter structure and the TAF-less PIC structure, they propose a structural model for the complete TFIID-based PIC, and provide insights into the role of TFIID in promoter recognition, PIC assembly, and transcription initiation.

Membraneless organelles or condensates form through liquid–liquid phase separation1–4, which is thought to underlie gene transcription through condensation of the large-scale nucleolus5–7 or in smaller assemblies known as transcriptional condensates8–11. Transcriptional condensates have been hypothesized to phase separate at particular genomic loci and locally promote the biomolecular interactions underlying gene expression. However, there have been few quantitative biophysical tests of this model in living cells, and phase separation has not yet been directly linked with dynamic transcriptional outputs12,13. Here, we apply an optogenetic approach to show that FET-family transcriptional regulators exhibit a strong tendency to phase separate within living cells, a process that can drive localized RNA transcription. We find that TAF15 has a unique charge distribution among the FET family members that enhances its interactions with the C-terminal domain of RNA polymerase II. Nascent C-terminal domain clusters at primed genomic loci lower the energetic barrier for nucleation of TAF15 condensates, which in turn further recruit RNA polymerase II to drive transcriptional output. These results suggest that positive feedback between interacting transcriptional components drives localized phase separation to amplify gene expression.Wei et al. show that clusters of unphosphorylated RNA polymerase II seed the nucleation of phase-separated condensates of TAF15, which further recruit RNA polymerase II to amplify transcriptional activation.

TLR4 signals via MyD88 and TRIF to activate proinflammatory cytokine and type I interferon responses. Karin and colleagues show that different ubiquitination of TRAF3 via K48 or K63 dictates the ensuing immune response. Balanced production of type I interferons and proinflammatory cytokines after engagement of Toll-like receptors (TLRs), which signal through adaptors containing a Toll–interleukin 1 receptor (TIR) domain, such as MyD88 and TRIF, has been proposed to control the pathogenesis of autoimmune disease and tumor responses to inflammation. Here we show that TRAF3, a ubiquitin ligase that interacts with both MyD88 and TRIF, regulated the production of interferon and proinflammatory cytokines in different ways. Degradative ubiquitination of TRAF3 during MyD88-dependent TLR signaling was essential for the activation of mitogen-activated protein kinases (MAPKs) and production of inflammatory cytokines. In contrast, TRIF-dependent signaling triggered noncanonical TRAF3 self-ubiquitination that activated the interferon response. Inhibition of degradative ubiquitination of TRAF3 prevented the expression of all proinflammatory cytokines without affecting the interferon response.

Tumor necrosis factor receptor-associated factor 2 (TRAF2) is a critical mediator of tumor necrosis factor-α (TNF-α) signaling. However, the regulatory mechanisms of TRAF2 are not fully understood. Here we show evidence that TRAF2 requires brefeldin A-inhibited guanine nucleotide-exchange factor 1 (BIG1) to be recruited into TNF receptor 1 (TNFR1) signaling complexes. In BIG1 knockdown cells, TNF-α-induced c-Jun N-terminal kinase (JNK) activation was attenuated and the sensitivity to TNF-α-induced apoptosis was increased. Since these trends correlated well with those of TRAF2 deficient cells as previously demonstrated, we tested whether BIG1 functions as an upstream regulator of TRAF2 in TNFR1 signaling. As expected, we found that knockdown of BIG1 suppressed TNF-α-dependent ubiquitination of TRAF2 that is required for JNK activation, and impaired the recruitment of TRAF2 to the TNFR1 signaling complex (complex I). Moreover, we found that the recruitment of TRAF2 to the death-inducing signaling complex termed complex II was also impaired in BIG1 knockdown cells. These results suggest that BIG1 is a key component of the machinery that drives TRAF2 to the signaling complexes formed after TNFR1 activation. Thus, our data demonstrate a novel and unexpected function of BIG1 that regulates TNFR1 signaling by targeting TRAF2.

Tumor necrosis factor receptor-associated factors (TRAFS) were initially discovered as adaptor proteins that couple the tumor necrosis factor receptor family to signaling pathways. More recently they have also been shown to be signal transducers of Toll/interleukin-1 family members. Six members of the TRAF family have been identified. All TRAF proteins share a C-terminal homology region termed the TRAF domain that is capable of binding to the cytoplasmic domain of receptors, and to other TRAF proteins. In addition, TRAFs 2-6 have RING and zinc finger motifs that are important for signaling downstream events. TRAF proteins are thought to be important regulators of cell death and cellular responses to stress, and TRAF2, TRAF5 and TRAF6 have been demonstrated to mediate activation of NF-kappaB and JNK. TRAF proteins are expressed in normal and diseased tissue in a regulated fashion, suggesting that they play an important role in physiological and pathological processes.

Frontotemporal lobar degeneration (FTLD) causes frontotemporal dementia (FTD), the most common form of dementia after Alzheimer’s disease, and is often also associated with motor disorders 1 . The pathological hallmarks of FTLD are neuronal inclusions of specific, abnormally assembled proteins 2 . In the majority of cases the inclusions contain amyloid filament assemblies of TAR DNA-binding protein 43 (TDP-43) or tau, with distinct filament structures characterizing different FTLD subtypes 3 , 4 . The presence of amyloid filaments and their identities and structures in the remaining approximately 10% of FTLD cases are unknown but are widely believed to be composed of the protein fused in sarcoma (FUS, also known as translocated in liposarcoma). As such, these cases are commonly referred to as FTLD–FUS. Here we used cryogenic electron microscopy (cryo-EM) to determine the structures of amyloid filaments extracted from the prefrontal and temporal cortices of four individuals with FTLD–FUS. Surprisingly, we found abundant amyloid filaments of the FUS homologue TATA-binding protein-associated factor 15 (TAF15, also known as TATA-binding protein-associated factor 2N) rather than of FUS itself. The filament fold is formed from residues 7–99 in the low-complexity domain (LCD) of TAF15 and was identical between individuals. Furthermore, we found TAF15 filaments with the same fold in the motor cortex and brainstem of two of the individuals, both showing upper and lower motor neuron pathology. The formation of TAF15 amyloid filaments with a characteristic fold in FTLD establishes TAF15 proteinopathy in neurodegenerative disease. The structure of TAF15 amyloid filaments provides a basis for the development of model systems of neurodegenerative disease, as well as for the design of diagnostic and therapeutic tools targeting TAF15 proteinopathy. Cryogenic electron microscopy structures of amyloid filaments extracted from patient brains reveal that the protein TAF15 forms filaments that characterize certain cases of frontotemporal lobar degeneration.

SAGA (Spt–Ada–Gcn5–acetyltransferase) is a 19-subunit complex that stimulates transcription via two chromatin-modifying enzymatic modules and by delivering the TATA box binding protein (TBP) to nucleate the pre-initiation complex on DNA, a pivotal event in the expression of protein-encoding genes 1 . Here we present the structure of yeast SAGA with bound TBP. The core of the complex is resolved at 3.5 Å resolution (0.143 Fourier shell correlation). The structure reveals the intricate network of interactions that coordinate the different functional domains of SAGA and resolves an octamer of histone-fold domains at the core of SAGA. This deformed octamer deviates considerably from the symmetrical analogue in the nucleosome and is precisely tuned to establish a peripheral site for TBP, where steric hindrance represses binding of spurious DNA. Complementary biochemical analysis points to a mechanism for TBP delivery and release from SAGA that requires transcription factor IIA and whose efficiency correlates with the affinity of DNA to TBP. We provide the foundations for understanding the specific delivery of TBP to gene promoters and the multiple roles of SAGA in regulating gene expression. Structural studies on the yeast transcription coactivator complex SAGA (Spt–Ada–Gcn5–acetyltransferase) provide insights into the mechanism of initiation of regulated transcription by this multiprotein complex, which is conserved among eukaryotes.

Eukaryotic cells use autophagy to recycle cellular components. During autophagy, autophagosomes deliver cytoplasmic contents to the vacuole or lysosome for breakdown. Mammalian cells regulate the dynamics of autophagy via ubiquitin-mediated proteolysis of autophagy proteins. Here, we show that the Tumor necrosis factor Receptor-Associated Factor (TRAF) family proteins TRAF1a and TRAF1b (previously named MUSE14 and MUSE13, respectively) help regulate autophagy via ubiquitination. Upon starvation, cytoplasmic TRAF1a and TRAF1b translocated to autophagosomes. Knockout lines showed reduced tolerance to nutrient deficiency, increased salicylic acid and reactive oxygen species levels, and constitutive cell death in rosettes, resembling the phenotypes of autophagy-defective mutants. Starvation-activated autophagosome accumulation decreased in root cells, indicating that TRAF1a and TRAF1b function redundantly in regulating autophagosome formation. TRAF1a and TRAF1b interacted in planta with ATG6 and the RING finger E3 ligases SINAT1, SINAT2, and SINAT6 (with a truncated RING-finger domain). SINAT1 and SINAT2 require the presence of TRAF1a and TRAF1b to ubiquitinate and destabilize AUTOPHAGY PROTEIN6 (ATG6) in vivo. Conversely, starvation-induced SINAT6 reduced SINAT1- and SINAT2-mediated ubiquitination and degradation of ATG6. Consistently, / and knockout mutants exhibited increased tolerance and sensitivity, respectively, to nutrient starvation. Therefore, TRAF1a and TRAF1b function as molecular adaptors that help regulate autophagy by modulating ATG6 stability in Arabidopsis.