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The liver exhibits a remarkable capacity to regenerate following injury. Despite this unique attribute, toxic injury is a leading cause of liver failure. The temporal processes by which the liver senses injury and initiates regeneration remain unclear. Here, we developed a transgenic zebrafish model wherein hepatocyte-specific expression of uracil phosphoribosyltransferase (UPRT) enabled the implementation of SLAM-ITseq to investigate the nascent transcriptome during initiation of liver injury and regeneration. Using this approach, we identified a rapid metabolic transition from the fed to the fasted state that was followed by induction of the nuclear erythroid 2-related factor (Nrf2) antioxidant program. We find that activation of Nrf2 in hepatocytes is required to induce the pentose phosphate pathway (PPP) and improve survival following liver injury. Mechanistically, we demonstrate that inhibition of the PPP disrupts nucleotide biosynthesis to prevent liver regeneration. Together, these studies provide fundamental insights into the mechanism by which early metabolic adaptation to injury facilitates tissue regeneration.

Chromatin regulation involves the selective recruitment of chromatin factors to facilitate DNA repair, replication and transcription. Here we demonstrate the utility of coupling unbiased functional genomics with chromatin immunoprecipitation (CRISPR-ChIP) to identify the factors associated with active chromatin modifications in mammalian cells. Specifically, an integrated reporter containing a cis-regulatory element of interest and a single guide RNA provide a chromatinized template for a direct readout for regulators of histone modifications associated with actively transcribed genes such as H3K4me3 and H3K79me2. With CRISPR-ChIP, we identify all the nonredundant COMPASS complex members required for H3K4me3 and demonstrate that RNA polymerase II is dispensable for the maintenance of H3K4me3. As H3K79me2 has a putative oncogenic function in leukemia cells driven by MLL translocations, using CRISPR-ChIP we reveal a functional partitioning of H3K79 methylation into two distinct regulatory units: an oncogenic DOT1L complex directed by the MLL fusion protein in a Menin-dependent manner and a separate endogenous DOT1L complex, where catalytic activity is directed by MLLT10. Overall, CRISPR-ChIP provides a powerful tool for the unbiased interrogation of the mechanisms underpinning chromatin regulation.

UNLABELLED: Therapies that enhance antitumor immunity have altered the natural history of many cancers. Consequently, leveraging nonoverlapping mechanisms to increase immunogenicity of cancer cells remains a priority. Using a novel enzymatic inhibitor of the RNA methyl-transferase METTL3, we demonstrate a global decrease in N6-methyladenosine (m6A) results in double-stranded RNA (dsRNA) formation and a profound cell-intrinsic interferon response. Through unbiased CRISPR screens, we establish dsRNA-sensing and interferon signaling are primary mediators that potentiate T-cell killing of cancer cells following METTL3 inhibition. We show in a range of immunocompetent mouse models that although METTL3 inhibition is equally efficacious to anti-PD-1 therapy, the combination has far greater preclinical activity. Using SPLINTR barcoding, we demonstrate that anti-PD-1 therapy and METTL3 inhibition target distinct malignant clones, and the combination of these therapies overcomes clones insensitive to the single agents. These data provide the mole-cular and preclinical rationale for employing METTL3 inhibitors to promote antitumor immunity in the clinic. SIGNIFICANCE: This work demonstrates that METTL3 inhibition stimulates a cell-intrinsic interferon response through dsRNA formation. This immunomodulatory mechanism is distinct from current immunotherapeutic agents and provides the molecular rationale for combination with anti-PD-1 immune-checkpoint blockade to augment antitumor immunity. This article is featured in Selected Articles from This Issue, p. 2109.

Precise control of activating H3K4me3 and repressive H3K27me3 histone modifications at bivalent promoters is essential for normal development and frequently corrupted in cancer. By coupling a cell surface readout of bivalent MHC class I gene expression with whole-genome CRISPR-Cas9 screens, we identify specific roles for MTF2-PRC2.1, PCGF1-PRC1.1 and Menin-KMT2A/B complexes in maintaining bivalency. Genetic loss or pharmacological inhibition of Menin unexpectedly phenocopies the effects of polycomb disruption, resulting in derepression of bivalent genes in both cancer cells and pluripotent stem cells. While Menin and KMT2A/B contribute to H3K4me3 at active genes, a separate Menin-independent function of KMT2A/B maintains H3K4me3 and opposes polycomb-mediated repression at bivalent genes. Release of KMT2A from active genes following Menin targeting alters the balance of polycomb and KMT2A at bivalent genes, facilitating gene activation. This functional partitioning of Menin-KMT2A/B complex components reveals therapeutic opportunities that can be leveraged through inhibition of Menin.

Satellite and GPS tracking technology continues to reveal new migration patterns of birds which enables comparative studies of migration strategies and distributional information useful in conservation. Bar-tailed godwitsin the East Asian–Australasian Flyway Limosa lapponica baueri and L. l. menzbieri are known for their long non-stop flights, howeverthese populations are in steep decline. A third subspecies in this flyway, L. l. anadyrensis, breeds in the Anadyr River basin, Chukotka,Russia, and is morphologically distinct from menzbieri and baueri based on comparison of museum specimens collected from breeding areas. However, the non-breeding distribution, migration route and population size of anadyrensis are entirely unknown. Among 24 female bar-tailed godwits tracked in 2015–2018 from northwest Australia, the main non-breeding area for menzbieri, two birds migrated further eastthan the rest to breed in the Anadyr River basin, i.e. they belonged to the anadyrensis subspecies. During pre-breeding migration, all birds staged in the Yellow Sea and then flew to the breeding grounds in the eastern Russian Arctic. After breeding, these two birds migrated southwestward to stage in Russia on the Kamchatka Peninsula and on Sakhalin Island en route to the Yellow Sea. This contrasts with the other 22tracked godwits that followed the previously described route of menzbieri, i.e. they all migrated northwards to stage in the New Siberian Islands before turning south towards the Yellow Sea, and onwards to northwest Australia. Since the Kamchatka Peninsula was not used by any of the tracked menzbieri birds, the 4500 godwits counted in theKhairusova–Belogolovaya estuary in western Kamchatka may well be anadyrensis. Comparing migration patterns across the three bar-tailed godwits subspecies, the migration strategy of anadyrensis lies between that of menzbieri and baueri. Futur

UNLABELLED: Cancer cell metabolism is increasingly recognized as providing an exciting therapeutic opportunity. However, a drug that directly couples targeting of a metabolic dependency with the induction of cell death in cancer cells has largely remained elusive. Here we report that the drug-like small-molecule ironomycin reduces the mitochondrial iron load, resulting in the potent disruption of mitochondrial metabolism. Ironomycin promotes the recruitment and activation of BAX/BAK, but the resulting mitochondrial outer membrane permeabilization (MOMP) does not lead to potent activation of the apoptotic caspases, nor is the ensuing cell death prevented by inhibiting the previously established pathways of programmed cell death. Consistent with the fact that ironomycin and BH3 mimetics induce MOMP through independent nonredundant pathways, we find that ironomycin exhibits marked in vitro and in vivo synergy with venetoclax and overcomes venetoclax resistance in primary patient samples. SIGNIFICANCE: Ironomycin couples targeting of cellular metabolism with cell death by reducing mitochondrial iron, resulting in the alteration of mitochondrial metabolism and the activation of BAX/BAK. Ironomycin induces MOMP through a different mechanism to BH3 mimetics, and consequently combination therapy has marked synergy in cancers such as acute myeloid leukemia. This article is highlighted in the In This Issue feature, p. 587.

There is increasing recognition of the prognostic significance of tumor cell major histocompatibility complex (MHC) class II expression in anti-cancer immunity. Relapse of acute myeloid leukemia (AML) following allogeneic stem cell transplantation (alloSCT) has recently been linked to MHC class II silencing in leukemic blasts; however, the regulation of MHC class II expression remains incompletely understood. Utilizing unbiased CRISPR-Cas9 screens, we identify that the C-terminal binding protein (CtBP) complex transcriptionally represses MHC class II pathway genes, while the E3 ubiquitin ligase complex component FBXO11 mediates degradation of CIITA, the principal transcription factor regulating MHC class II expression. Targeting these repressive mechanisms selectively induces MHC class II upregulation across a range of AML cell lines. Functionally, MHC class II+ leukemic blasts stimulate antigen-dependent CD4+ T cell activation and potent anti-tumor immune responses, providing fundamental insights into the graft-versus-leukemia effect. These findings establish the rationale for therapeutic strategies aimed at restoring tumor-specific MHC class II expression to salvage AML relapse post-alloSCT and also potentially to enhance immunotherapy outcomes in non-myeloid malignancies.

Aim: Molluscs are important grazers, filter and deposit feeders, scavengers and predators, which in turn are food for shorebirds, fish and people. Some species, targeted as human food, have been cultured along the Chinese coast for hundreds of years. To examine whether aquacultural practices have meanwhile affected biodiversity gradients, we measured mollusc community structure along the coast of China in habitats which are intensively used by humans. Location: Chinese coast. Methods: We sampled 21 intertidal sites spanning 20 latitudinal degrees and 18,400 km of coastline. We assessed alpha diversity to verify whether mollusc communities exhibit the expected biodiversity gradient with latitude and beta diversity gradients with distance. To examine whether human activities such as transportation and culturing could have affected these patterns, we distinguished commercial from non-commercial mollusc species and compared the differences in distribution, density, alpha diversity and beta diversity. Results: We found non-commercial species showed the expected biodiversity gradients. Commercial species (a) dominated the intertidal mollusc communities at 19 of the 21 sites and compared with non-commercial species, (b) exhibited wider geographical distributions, (c) showed no significant change in Bray-Curtis index (abundance-based beta diversity) with either geographical or climatic distance, (d) exhibited lower average dissimilarities and (e) did not show a decrease in species richness and Shannon diversity with latitude. Combining all species, trends were the same as for the commercial species. Main conclusions: A few cultured species dominated the intertidal mollusc communities in high densities along the Chinese coastline, taking over the ecological roles of the native species but not driving them extinct. In this way, aquacultural practices have exerted a homogenizing influence strong enough to erase basic biodiversity gradients. Since molluscs are food for the grow

Shorebirds in the East Asian–Australasian Flyway are struggling because of the various threats they faced, notably habitat loss and deterioration in intertidal flats in East Asia. This thesis studies three species migrating between Northwest Australia and the breeding areas in the eastern Russian Arctic: The Great Knot, Red Knot and Bar-tailed Godwit, which depend on East Asian intertidal flats to fuel up during their migrations. To track the migration journeys, small satellite tags are attached onto the birds. We first describe migration routes and timing of the anadyrensis subspecies of the Bar-tailed Godwit which were unknown before this study. Then we describe how the Red Knot, previously thought to be a ‘long-jump’ migrant, in fact made several short stops during northward migration. To galvanize conservation actions in this flyway, we explore ways to apply satellite tracking of shorebirds in coastal conservation. Satellite tracking uncovers many potentially important but unrecognized shorebird sites, and also helps plan on-ground surveys at lesser-known sites. In collaboration with hydraulic engineers, we incorporate shorebird tracking data in ecological impact assessment of port construction and in ‘Building with Nature’ port design. Finally, we investigate what determines how shorebirds cope with habitat deteriorations. Site fidelity, which might deter birds to move away from a site when it deteriorates, is lower in Great Knots than in Bar-tailed Godwits. Moreover, more explorative great knots responded quicker to the collapse in prey stock at their main staging site.

Climate warming in the Arctic has led to warmer and earlier springs, and as a result, many food resources for migratory animals become available earlier in the season, as well as become distributed further northwards. To optimally profit from these resources, migratory animals are expected to arrive earlier in the Arctic, as well as shift their own spatial distributions northwards. Here, we review literature to assess whether Arctic migra-tory birds and mammals already show shifts in migration timing or distribution in response to the warming climate. Distribution shifts were most prominent in marine mammals, as expected from observed northward shifts of their resources. At least for many bird species, the ability to shift distributions is likely constrained by available habitat further north. Shifts in timing have been shown in many species of terrestrial birds and ungulates, as well as for polar bears. Within species, we found strong variation in shifts in timing and distributions between populations. Ou r review thus shows that many migratory animals display shifts in migration timing and spatial distribution in reaction to a warming Arctic. Importantly, we identify large knowledge gaps especially concerning distribution shifts and timing of autumn migration, especially for marine mammals. Our understanding of how migratory animals respond to climate change appears to be mostly limited by the lack of long-term monitoring studies.