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The UFM1 system, a ubiquitin-like conjugation system is crucial for endoplasmic reticulum (ER) homeostasis. Here, authors found that CYB5R3 is covalently conjugated with UFM1, which becomes a signal for ER-phagy, a selective autophagy of ER.

Abstract Recent studies have shown that adipose tissue is an immunological organ. While inflammation in energy-storing white adipose tissues has been the focus of intense research, the regulatory mechanisms of inflammation in heat-producing brown adipose tissues remain largely unknown. We previously identified apoptosis signal-regulating kinase 1 (ASK1) as a critical regulator of brown adipocyte maturation; the PKA-ASK1-p38 axis facilitates uncoupling protein 1 (UCP1) induction cell-autonomously. Here, we show that ASK1 suppresses an innate immune pathway and contributes to maintenance of brown adipocytes. We report a novel chemical pull-down method for endogenous kinases using analog sensitive kinase allele (ASKA) technology and identify an ASK1 interactor in brown adipocytes, receptor-interacting serine/threonine-protein kinase 2 (RIPK2). ASK1 disrupts the RIPK2 signaling complex and inhibits the NOD-RIPK2 pathway to downregulate the production of inflammatory cytokines. As a potential biological significance, an in vitro model for intercellular regulation suggests that ASK1 facilitates the expression of UCP1 through the suppression of inflammatory cytokine production. In parallel to our previous report on the PKA-ASK1-p38 axis, our work raises the possibility of an auxiliary role of ASK1 in brown adipocyte maintenance through neutralizing the thermogenesis-suppressive effect of the NOD-RIPK2 pathway.

ABSTRACT Influenza viral particles are assembled at the plasma membrane concomitantly with Rab11a-mediated endocytic transport of viral ribonucleoprotein complexes (vRNPs). The mechanism of spatiotemporal regulation of viral budozone formation and its regulatory molecules on the endocytic vesicles remain unclear. Here, we performed a proximity-based proteomics approach for Rab11a and found that ARHGAP1, a Rho GTPase-activating protein, is transported through the Rab11a-mediated apical transport of vRNP. ARHGAP1 stabilized actin filaments in infected cells for the lateral clustering of hemagglutinin (HA) molecules, a viral surface membrane protein, to the budozone. Disruption of the HA clustering results in the production of virions with low HA content, and such virions were less resistant to protease and had enhanced antigenicity, presumably because reduced clustering of viral membrane proteins exposes hidden surfaces. Collectively, these results demonstrate that Rab11a-mediated endocytic transport of ARHGAP1 with vRNPs stimulates budozone formation to ensure the integrity of virion surface required for viral survival. IMPORTANCE The endocytic transport of the influenza viral genome triggers the clustering of viral membrane proteins at the plasma membrane to form the viral budozone. However, host factors that promote viral budozone formation in concert with viral genome transport have not been identified. Here, we found that ARHGAP1, a negative regulator of the Rho family protein, is transported with the viral genome and stabilizes actin filaments to promote budozone formation. We have shown that ARHGAP1-mediated efficient formation of viral budozone was crucial for the clustering of viral HA protein to the progeny viral particles. The clustering of HA proteins on the virions is responsible for the structural integrity of the viral particles, which promotes viral stability and viral immune evasion. This study highlights the molecular mechanism that works in concert with viral genome packaging to ensure the structural integrity of viral particles.

Here the authors use chemical proteomics to identify the herpes simplex virus 1 encoded proteome in infected cells. Functional characterization of one of the nine identified proteins, designated piUL49, shows that it acts as neurovirulence factor in mice by regulating a virally encoded dUTPase.

Boosting energy expenditure by harnessing the activity of brown adipocytes is a promising strategy for combatting the global epidemic of obesity. Many studies have revealed that the β3-adrenergic receptor agonist is a potent activator of brown adipocytes, even in humans, and PKA and p38 MAPK have been demonstrated for regulating the transcription of a wide range of critical genes such as Ucp1. We previously revealed that the PKA-ASK1-p38 axis is activated in immature brown adipocytes and contributes to functional maturation. However, the downstream mechanisms of PKA that initiate the p38 MAPK cascade are still mostly unknown in mature brown adipocytes. Here, we identified the ASK family as a crucial signaling molecule bridging PKA and MAPK in mature brown adipocytes. Mechanistically, the phosphorylation of ASK1 at threonine 99 and serine 993 is critical in PKA-dependent ASK1 activation. Additionally, PKA also activates ASK2, which contributes to MAPK regulation. These lines of evidence provide new details for tailoring a βAR-dependent brown adipocyte activation strategy.

Yes-associated protein (YAP) is a growth-promoting transcription co-activator that regulates the malignancy of various cancers, however its regulation is not fully understood. Here the authors show that phosphatdylserine at recycling endosomes regulates YAP signalling pathway.

Apoptosis signal-regulating kinase 1 (ASK1) is an oxidative stress-responsive kinase that is regulated by various interacting molecules and post-translational modifications. However, how these molecules and modifications cooperatively regulate ASK1 activity remains largely unknown. Here, we showed that tripartite motif 48 (TRIM48) orchestrates the regulation of oxidative stress-induced ASK1 activation. A pull-down screen identified a TRIM48-interacting partner, protein arginine methyltransferase 1 (PRMT1), which negatively regulates ASK1 activation by enhancing its interaction with thioredoxin (Trx), another ASK1-negative regulator. TRIM48 facilitates ASK1 activation by promoting K48-linked polyubiquitination and degradation of PRMT1. TRIM48 knockdown suppressed oxidative stress-induced ASK1 activation and cell death, whereas forced expression promoted cancer cell death in mouse xenograft model. These results indicate that TRIM48 facilitates oxidative stress-induced ASK1 activation and cell death through ubiquitination-dependent degradation of PRMT1. This study provides a cell death mechanism fine-tuned by the crosstalk between enzymes that engage various types of post-translational modifications.

Mutations inCHCHD2 are associated with Parkinson’s disease. Here the authors investigate the physiological and pathological roles of CHCHD2 in Drosophilaand mammalian cells, and find that it regulates mitochondrial respiration through stabilizing cytochrome c.

ZFP36L2 promotes the destruction of AU-rich element-containing transcripts, while its regulation and functional significance in cell cycle control are scarcely identified. We show that ZFP36L2 is a cell cycle-regulated CCCH protein, the abundance of which is regulated post-translationally at the respective stages of the cell cycle. Indeed, ZFP36L2 protein was eliminated after release from M phase, and ZYG11B-based E3 ligase plays a role in its polyubiquitination in interphase. Although ZFP36L2 is dispensable for normal cell cycle progression, we found that endogenous ZFP36L2 played a key role in cisplatin-induced S-phase arrest, a process in which the suppression of G1/S cyclins is necessary. The accumulation of ZFP36L2 was stimulated under DNA replication stresses and altered interactions with a subset of RNA-binding proteins. Notably, silencing endogenous ZFP36L2 led to impaired cell viability in the presence of cisplatin-induced DNA lesions. Thus, we propose that ZFP36L2 is a key protein that controls S-phase progression in the case of genome instability.

Cells possess ER quality control systems to adapt to ER stress and maintain their function. ER-stress-induced pre-emptive quality control (ER pQC) selectively degrades ER proteins via translocational attenuation during ER stress. However, the molecular mechanism underlying this process remains unclear. Here, we find that most newly synthesized endogenous transthyretin proteins are rerouted to the cytosol without cleavage of the signal peptide, resulting in proteasomal degradation in hepatocytes during ER stress. Derlin family proteins (Derlins), which are ER-associated degradation components, reroute specific ER proteins, but not ER chaperones, from the translocon to the proteasome through interactions with the signal recognition particle (SRP). Moreover, the cytosolic chaperone Bag6 and the AAA-ATPase p97 contribute to the degradation of ER pQC substrates. These findings demonstrate that Derlins-mediated substrate-specific rerouting and Bag6- and p97-mediated effective degradation contribute to the maintenance of ER homeostasis without the need for translocation.

Mbtd1 (mbt domain containing 1) encodes a nuclear protein containing a zinc finger domain and four malignant brain tumor (MBT) repeats. We previously generated Mbtd1-deficient mice and found that MBTD1 is highly expressed in fetal hematopoietic stem cells (HSCs) and sustains the number and function of fetal HSCs. However, since Mbtd1-deficient mice die soon after birth possibly due to skeletal abnormalities, its role in adult hematopoiesis remains unclear. To address this issue, we generated Mbtd1 conditional knockout mice and analyzed adult hematopoietic tissues deficient in Mbtd1. We observed that the numbers of HSCs and progenitors increased and Mbtd1-deficient HSCs exhibited hyperactive cell cycle, resulting in a defective response to exogenous stresses. Mechanistically, we found that MBTD1 directly binds to the promoter region of FoxO3a, encoding a forkhead protein essential for HSC quiescence, and interacts with components of TIP60 chromatin remodeling complex and other proteins involved in HSC and other stem cell functions. Restoration of FOXO3a activity in Mbtd1-deficient HSCs in vivo rescued cell cycle and pool size abnormalities. These findings indicate that MBTD1 is a critical regulator for HSC pool size and function, mainly through the maintenance of cell cycle quiescence by FOXO3a. [ABSTRACT FROM AUTHOR]

In macroautophagy, membrane structures called autophagosomes engulf substrates and deliver them for lysosomal degradation. Autophagosomes enwrap a variety of targets with diverse sizes, from portions of cytosol to larger organelles. However, the mechanism by which autophagosome size is controlled remains elusive. We characterized a novel ER membrane protein, ERdj8, in mammalian cells. ERdj8 localizes to a meshwork-like ER subdomain along with phosphatidylinositol synthase (PIS) and autophagy-related (Atg) proteins. ERdj8 overexpression extended the size of the autophagosome through its DnaJ and TRX domains. ERdj8 ablation resulted in a defect in engulfing larger targets. C. elegans, in which the ERdj8 orthologue dnj-8 was knocked down, could perform autophagy on smaller mitochondria derived from the paternal lineage but not the somatic mitochondria. Thus, ERdj8 may play a critical role in autophagosome formation by providing the capacity to target substrates of diverse sizes for degradation. [ABSTRACT FROM AUTHOR]