Large neutral amino acid levels tune perinatal neuronal excitability and survival.

Little is known about the critical metabolic changes that neural cells have to undergo during development and how temporary shifts in this program can influence brain circuitries and behavior. Inspired by the discovery that mutations in SLC7A5, a transporter of metabolically essential large neutral amino acids (LNAAs), lead to autism, we employed metabolomic profiling to study the metabolic states of the cerebral cortex across different developmental stages. We found that the forebrain undergoes significant metabolic remodeling throughout development, with certain groups of metabolites showing stage-specific changes, but what are the consequences of perturbing this metabolic program? By manipulating Slc7a5 expression in neural cells, we found that the metabolism of LNAAs and lipids are interconnected in the cortex. Deletion of Slc7a5 in neurons affects the postnatal metabolic state, leading to a shift in lipid metabolism. Additionally, it causes stage- and cell-type-specific alterations in neuronal activity patterns, resulting in a long-term circuit dysfunction.

Atypical chemokine receptor 1 on nucleated erythroid cells regulates hematopoiesis.

Healthy individuals of African ancestry have neutropenia that has been linked with the variant rs2814778(G) of the gene encoding atypical chemokine receptor 1 (ACKR1). This polymorphism selectively abolishes the expression of ACKR1 in erythroid cells, causing a Duffy-negative phenotype. Here we describe an unexpected fundamental role for ACKR1 in hematopoiesis and provide the mechanism that links its absence with neutropenia. Nucleated erythroid cells had high expression of ACKR1, which facilitated their direct contact with hematopoietic stem cells. The absence of erythroid ACKR1 altered mouse hematopoiesis including stem and progenitor cells, which ultimately gave rise to phenotypically distinct neutrophils that readily left the circulation, causing neutropenia. Individuals with a Duffy-negative phenotype developed a distinct profile of neutrophil effector molecules that closely reflected the one observed in the ACKR1-deficient mice. Thus, alternative physiological patterns of hematopoiesis and bone marrow cell outputs depend on the expression of ACKR1 in the erythroid lineage, findings with major implications for the selection advantages that have resulted in the paramount fixation of the ACKR1 rs2814778(G) polymorphism in Africa.

Shared behavioural impairments in visual perception and place avoidance across different autism models are driven by periaqueductal grey hypoexcitability in Setd5 haploinsufficient mice.

Despite the diverse genetic origins of autism spectrum disorders (ASDs), affected individuals share strikingly similar and correlated behavioural traits that include perceptual and sensory processing challenges. Notably, the severity of these sensory symptoms is often predictive of the expression of other autistic traits. However, the origin of these perceptual deficits remains largely elusive. Here, we show a recurrent impairment in visual threat perception that is similarly impaired in 3 independent mouse models of ASD with different molecular aetiologies. Interestingly, this deficit is associated with reduced avoidance of threatening environments-a nonperceptual trait. Focusing on a common cause of ASDs, the Setd5 gene mutation, we define the molecular mechanism. We show that the perceptual impairment is caused by a potassium channel (Kv1)-mediated hypoexcitability in a subcortical node essential for the initiation of escape responses, the dorsal periaqueductal grey (dPAG). Targeted pharmacological Kv1 blockade rescued both perceptual and place avoidance deficits, causally linking seemingly unrelated trait deficits to the dPAG. Furthermore, we show that different molecular mechanisms converge on similar behavioural phenotypes by demonstrating that the autism models Cul3 and Ptchd1, despite having similar behavioural phenotypes, differ in their functional and molecular alteration. Our findings reveal a link between rapid perception controlled by subcortical pathways and appropriate learned interactions with the environment and define a nondevelopmental source of such deficits in ASD.

The atypical chemokine receptor CCRL1 shapes functional CCL21 gradients in lymph nodes.

Afferent lymph-borne dendritic cells essentially rely on the chemokine receptor CCR7 for their transition from the subcapsular lymph node sinus into the parenchyma, a migratory step driven by putative gradients of CCR7 ligands. We found that lymph node fringes indeed contained physiological gradients of the chemokine CCL21, which depended on the expression of CCRL1, the atypical receptor for the CCR7 ligands CCL19 and CCL21. Lymphatic endothelial cells lining the ceiling of the subcapsular sinus, but not those lining the floor, expressed CCRL1, which scavenged chemokines from the sinus lumen. This created chemokine gradients across the sinus floor and enabled the emigration of dendritic cells. In vitro live imaging revealed that spatially confined expression of CCRL1 was necessary and sufficient for the creation of functional chemokine gradients.

A lineage-specific STAT5BN642H mouse model to study NK-cell leukemia.

ABSTRACT: Patients with T- and natural killer (NK)-cell neoplasms frequently have somatic STAT5B gain-of-function mutations. The most frequent STAT5B mutation is STAT5BN642H, which is known to drive murine T-cell leukemia, although its role in NK-cell malignancies is unclear. Introduction of the STAT5BN642H mutation into human NK-cell lines enhances their potential to induce leukemia in mice. We have generated a mouse model that enables tissue-specific expression of STAT5BN642H and have selectively expressed the mutated STAT5B in hematopoietic cells (N642Hvav/+) or exclusively in NK cells (N642HNK/NK). All N642Hvav/+ mice rapidly develop an aggressive T/NKT-cell leukemia, whereas N642HNK/NK mice display an indolent NK-large granular lymphocytic leukemia (NK-LGLL) that progresses to an aggressive leukemia with age. Samples from patients with NK-cell leukemia have a distinctive transcriptional signature driven by mutant STAT5B, which overlaps with that of murine leukemic N642HNK/NK NK cells. To our knowledge, we have generated the first reliable STAT5BN642H-driven preclinical mouse model that displays an indolent NK-LGLL progressing to aggressive NK-cell leukemia. This novel in vivo tool will enable us to explore the transition from an indolent to an aggressive disease and will thus permit the study of prevention and treatment options for NK-cell malignancies.

Lymphatic Endothelial Cells Control Initiation of Lymph Node Organogenesis.

Lymph nodes (LNs) are strategically situated throughout the body at junctures of the blood vascular and lymphatic systems to direct immune responses against antigens draining from peripheral tissues. The current paradigm describes LN development as a programmed process that is governed through the interaction between mesenchymal lymphoid tissue organizer (LTo) cells and hematopoietic lymphoid tissue inducer (LTi) cells. Using cell-type-specific ablation of key molecules involved in lymphoid organogenesis, we found that initiation of LN development is dependent on LTi-cell-mediated activation of lymphatic endothelial cells (LECs) and that engagement of mesenchymal stromal cells is a succeeding event. LEC activation was mediated mainly by signaling through receptor activator of NF-κB (RANK) and the non-canonical NF-κB pathway and was steered by sphingosine-1-phosphate-receptor-dependent retention of LTi cells in the LN anlage. Finally, the finding that pharmacologically enforced interaction between LTi cells and LECs promotes ectopic LN formation underscores the central LTo function of LECs.

Central Nervous System Stromal Cells Control Local CD8(+) T Cell Responses during Virus-Induced Neuroinflammation.

Stromal cells generate a complex cellular scaffold that provides specialized microenvironments for lymphocyte activation in secondary lymphoid organs. Here, we assessed whether local activation of stromal cells in the central nervous system (CNS) is mandatory to transfer immune recognition from secondary lymphoid organs into the infected tissue. We report that neurotropic virus infection in mice triggered the establishment of such stromal cell niches in the CNS. CNS stromal cell activation was dominated by a rapid and vigorous production of CC-motif chemokine receptor (CCR) 7 ligands CCL19 and CCL21 by vascular endothelial cells and adjacent fibroblastic reticular cell (FRC)-like cells in the perivascular space. Moreover, CCR7 ligands produced by CNS stromal cells were crucial to support recruitment and local re-activation of antiviral CD8(+) T cells and to protect the host from lethal neuroinflammatory disease, indicating that CNS stromal cells generate confined microenvironments that control protective T cell immunity.

Coincident pre- and postsynaptic activation induces dendritic filopodia via neurotrypsin-dependent agrin cleavage.

The synaptic serine protease neurotrypsin is essential for cognitive function, as its deficiency in humans results in severe mental retardation. Recently, we demonstrated the activity-dependent release of neurotrypsin from presynaptic terminals and proteolytical cleavage of agrin at the synapse. Here we show that the activity-dependent formation of dendritic filopodia is abolished in hippocampal neurons from neurotrypsin-deficient mice. Administration of the neurotrypsin-dependent 22 kDa fragment of agrin rescues the filopodial response. Detailed analyses indicated that presynaptic action potential firing is necessary for the release of neurotrypsin, whereas postsynaptic NMDA receptor activation is necessary for the neurotrypsin-dependent cleavage of agrin. This contingency characterizes the neurotrypsin-agrin system as a coincidence detector of pre- and postsynaptic activation. As the resulting dendritic filopodia are thought to represent precursors of synapses, the neurotrypsin-dependent cleavage of agrin at the synapse may be instrumental for a Hebbian organization and remodeling of synaptic circuits in the CNS.

IFIT1 is an antiviral protein that recognizes 5'-triphosphate RNA.

Antiviral innate immunity relies on the recognition of microbial structures. One such structure is viral RNA that carries a triphosphate group on its 5' terminus (PPP-RNA). By an affinity proteomics approach with PPP-RNA as the 'bait', we found that the antiviral protein IFIT1 (interferon-induced protein with tetratricopeptide repeats 1) mediated binding of a larger protein complex containing other IFIT family members. IFIT1 bound PPP-RNA with nanomolar affinity and required the arginine at position 187 in a highly charged carboxy-terminal groove of the protein. In the absence of IFIT1, the growth and pathogenicity of viruses containing PPP-RNA was much greater. In contrast, IFIT proteins were dispensable for the clearance of pathogens that did not generate PPP-RNA. On the basis of this specificity and the great abundance of IFIT proteins after infection, we propose that the IFIT complex antagonizes viruses by sequestering specific viral nucleic acids.

Phenotypic characterization of disease-initiating stem cells in JAK2- or CALR-mutated myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPN) are characterized by uncontrolled expansion of myeloid cells, disease-related mutations in certain driver-genes including JAK2, CALR, and MPL, and a substantial risk to progress to secondary acute myeloid leukemia (sAML). Although behaving as stem cell neoplasms, little is known about disease-initiating stem cells in MPN. We established the phenotype of putative CD34+ /CD38- stem cells and CD34+ /CD38+ progenitor cells in MPN. A total of 111 patients with MPN suffering from polycythemia vera, essential thrombocythemia, or primary myelofibrosis (PMF) were examined. In almost all patients tested, CD34+ /CD38- stem cells expressed CD33, CD44, CD47, CD52, CD97, CD99, CD105, CD117, CD123, CD133, CD184, CD243, and CD274 (PD-L1). In patients with PMF, MPN stem cells often expressed CD25 and sometimes also CD26 in an aberrant manner. MPN stem cells did not exhibit substantial amounts of CD90, CD273 (PD-L2), CD279 (PD-1), CD366 (TIM-3), CD371 (CLL-1), or IL-1RAP. The phenotype of CD34+ /CD38- stem cells did not change profoundly during progression to sAML. The disease-initiating capacity of putative MPN stem cells was confirmed in NSGS mice. Whereas CD34+ /CD38- MPN cells engrafted in NSGS mice, no substantial engraftment was produced by CD34+ /CD38+ or CD34- cells. The JAK2-targeting drug fedratinib and the BRD4 degrader dBET6 induced apoptosis and suppressed proliferation in MPN stem cells. Together, MPN stem cells display a unique phenotype, including cytokine receptors, immune checkpoint molecules, and other clinically relevant target antigens. Phenotypic characterization of neoplastic stem cells in MPN and sAML should facilitate their enrichment and the development of stem cell-eradicating (curative) therapies.

Hepatocytic p62 suppresses ductular reaction and tumorigenesis in mouse livers with mTORC1 activation and defective autophagy.

BACKGROUND & AIMS: Either activation of mTORC1 due to loss of Tsc1 (tuberous sclerosis complex 1) or defective hepatic autophagy due to loss of Atg5 leads to spontaneous liver tumorigenesis in mice. The purpose of this study was to investigate the mechanisms by which autophagy contributes to the hepatic metabolic changes and tumorigenesis mediated by mTORC1 activation. METHODS: Atg5 Flox/Flox (Atg5F/F) and Tsc1F/F mice were crossed with albumin-Cre mice to generate liver-specific Atg5 knockout (L-Atg5 KO), L-Tsc1 KO and L-Atg5/Tsc1 double KO (DKO) mice. These mice were crossed with p62/Sqstm1F/F (p62) and whole body Nrf2 KO mice to generate L-Atg5/Tsc1/p62 and L-Atg5/Tsc1-Nrf2 triple KO mice. These mice were housed for various periods up to 12 months, and blood and liver tissues were harvested for biochemical and histological analysis RESULTS: Deletion of Atg5 in L-Tsc1 KO mice inhibited liver tumorigenesis but increased mortality and was accompanied by drastically enhanced hepatic ductular reaction (DR), hepatocyte degeneration and metabolic reprogramming. Deletion of p62 reversed DR, hepatocyte degeneration and metabolic reprogramming as well as the mortality of L-Atg5/Tsc1 DKO mice, but unexpectedly promoted liver tumorigenesis via activation of a group of oncogenic signaling pathways. Nrf2 ablation markedly improved DR with increased hepatocyte population and improved metabolic reprogramming and survival of the L-Atg5/Tsc1 DKO mice without tumor formation. Decreased p62 and increased mTOR activity were also observed in a subset of human hepatocellular carcinomas. CONCLUSIONS: These results reveal previously undescribed functions of hepatic p62 in suppressing tumorigenesis and regulating liver cell repopulation and metabolic reprogramming resulting from persistent mTORC1 activation and defective autophagy. LAY SUMMARY: Metabolic liver disease and viral hepatitis are common chronic liver diseases and risk factors of hepatocellular carcinoma, which are often associated with impaired hepatic autophagy and increased mTOR activation. Using multiple genetically engineered mouse models of defective hepatic autophagy and persistent mTOR activation, we dissected the complex mechanisms behind this observation. Our results uncovered an unexpected novel tumor suppressor function of p62/Sqstm1, which regulated liver cell repopulation, ductular reaction and metabolic reprogramming in liver tumorigenesis.

Maturation of lymph node fibroblastic reticular cells from myofibroblastic precursors is critical for antiviral immunity.

The stromal scaffold of the lymph node (LN) paracortex is built by fibroblastic reticular cells (FRCs). Conditional ablation of lymphotoxin-ß receptor (LTßR) expression in LN FRCs and their mesenchymal progenitors in developing LNs revealed that LTßR-signaling in these cells was not essential for the formation of LNs. Although T cell zone reticular cells had lost podoplanin expression, they still formed a functional conduit system and showed enhanced expression of myofibroblastic markers. However, essential immune functions of FRCs, including homeostatic chemokine and interleukin-7 expression, were impaired. These changes in T cell zone reticular cell function were associated with increased susceptibility to viral infection. Thus, myofibroblasic FRC precursors are able to generate the basic T cell zone infrastructure, whereas LTßR-dependent maturation of FRCs guarantees full immunocompetence and hence optimal LN function during infection.

BRD4 degradation blocks expression of MYC and multiple forms of stem cell resistance in Ph+ chronic myeloid leukemia.

In most patients with chronic myeloid leukemia (CML) clonal cells can be kept under control by BCR::ABL1 tyrosine kinase inhibitors (TKI). However, overt resistance or intolerance against these TKI may occur. We identified the epigenetic reader BRD4 and its downstream-effector MYC as growth regulators and therapeutic targets in CML cells. BRD4 and MYC were found to be expressed in primary CML cells, CD34+ /CD38- leukemic stem cells (LSC), and in the CML cell lines KU812, K562, KCL22, and KCL22T315I . The BRD4-targeting drug JQ1 was found to suppress proliferation in KU812 cells and primary leukemic cells in the majority of patients with chronic phase CML. In the blast phase of CML, JQ1 was less effective. However, the BRD4 degrader dBET6 was found to block proliferation and/or survival of primary CML cells in all patients tested, including blast phase CML and CML cells exhibiting the T315I variant of BCR::ABL1. Moreover, dBET6 was found to block MYC expression and to synergize with BCR::ABL1 TKI in inhibiting the proliferation in the JQ1-resistant cell line K562. Furthermore, BRD4 degradation was found to overcome osteoblast-induced TKI resistance of CML LSC in a co-culture system and to block interferon-gamma-induced upregulation of the checkpoint antigen PD-L1 in LSC. Finally, dBET6 was found to suppress the in vitro survival of CML LSC and their engraftment in NSG mice. Together, targeting of BRD4 and MYC through BET degradation sensitizes CML cells against BCR::ABL1 TKI and is a potent approach to overcome multiple forms of drug resistance in CML LSC.

Influence of sperm cryopreservation on sperm motility and proAKAP4 concentration in mice.

Background: The protein proAKAP4 is crucial for sperm motility and has been suggested as an indicator of male fertility. We determined the relationship between proAKAP4 concentration and sperm motility parameters in mice, and investigated the effects of cryopreservation on these variables. Methods: Computer-assisted sperm analysis and ELISA were applied to determine sperm motility and proAKAP4 concentration in fresh and frozen-thawed epididymal sperm of SWISS, B6D2F1, C57BL/6N, and BALB/c mice. Results: ProAKAP4 levels ranged between 12 and 89 ng/ml and did not differ between fresh and frozen-thawed samples, or between strains. We found a negative relationship between proAKAP4 levels and some sperm motility parameters. Sperm traits differed between strains, and cryopreservation negatively affected sperm velocity but not sperm direction parameters. Conclusion: ProAKAP4 levels in epididymal mouse spermatozoa were unaffected by cryopreservation, highlighting the robustness of this parameter as a potentially time-independent marker for sperm motility and fertility. The high individual variation in proAKAP4 levels supports the potential role of proAKAP4 as a marker for sperm quality, though we found no positive, and even negative relationships between proAKAP4 levels and some sperm motility parameters. Future studies have to investigate the significance of proAKAP4 as an indicator for fertility in mice.

Different Roles of p62 (SQSTM1) Isoforms in Keratin-Related Protein Aggregation.

p62/Sequestosome-1 (p62) is a multifunctional adaptor protein and is also a constant component of disease-associated protein aggregates, including Mallory-Denk bodies (MDBs), in steatohepatitis and hepatocellular carcinoma. We investigated the interaction of the two human p62 isoforms, p62-H1 (full-length isoform) and p62-H2 (partly devoid of PB1 domain), with keratins 8 and 18, the major components of MDBs. In human liver, p62-H2 is expressed two-fold higher compared to p62-H1 at the mRNA level and is present in slightly but not significantly higher concentrations at the protein level. Co-transfection studies in CHO-K1 cells, PLC/PRF/5 cells as well as p62- total-knockout and wild-type mouse fibroblasts revealed marked differences in the cytoplasmic distribution and aggregation behavior of the two p62 isoforms. Transfection-induced overexpression of p62-H2 generated large cytoplasmic aggregates in PLC/PRF/5 and CHO-K1 cells that mostly co-localized with transfected keratins resembling MDBs or (transfection without keratins) intracytoplasmic hyaline bodies. In fibroblasts, however, transfected p62-H2 was predominantly diffusely distributed in the cytoplasm. Aggregation of p62-H2 and p62ΔSH2 as well as the interaction with K8 (but not with K18) involves acquisition of cross-ß-sheet conformation as revealed by staining with luminescent conjugated oligothiophenes. These results indicate the importance of considering p62 isoforms in protein aggregation disease.

Lymphangiogenesis in a mouse model of renal transplant rejection extends life span of the recipients.

Renal allograft rejection can be prevented by immunological tolerance, which may be associated with de novo formed lymphatic vessels in the donor kidney after transplantation in man. A suitable mouse model of renal allograft rejection in which lymphangiogenesis can be deliberately induced in the graft is critical for elucidating the mechanisms responsible for the association between attenuated transplant rejection and abundance of lymphatic vessels. Here we describe the development of a novel mouse model of rapid renal transplant rejection in which transgenic induction of lymphangiogenesis in the immune-incompatible graft greatly extends its survival time. Thus, our novel approach may facilitate exploitation of lymphangiogenesis in the grafted organ.

High activation of STAT5A drives peripheral T-cell lymphoma and leukemia.

Recurrent gain-of-function mutations in the transcription factors STAT5A and much more in STAT5B were found in hematopoietic malignancies with the highest proportion in mature T- and natural killer-cell neoplasms (peripheral T-cell lymphoma, PTCL). No targeted therapy exists for these heterogeneous and often aggressive diseases. Given the shortage of models for PTCL, we mimicked graded STAT5A or STAT5B activity by expressing hyperactive Stat5a or STAT5B variants at low or high levels in the hematopoietic system of transgenic mice. Only mice with high activity levels developed a lethal disease resembling human PTCL. Neoplasia displayed massive expansion of CD8+ T cells and destructive organ infiltration. T cells were cytokine-hypersensitive with activated memory CD8+ T-lymphocyte characteristics. Histopathology and mRNA expression profiles revealed close correlation with distinct subtypes of PTCL. Pronounced STAT5 expression and activity in samples from patients with different subsets underline the relevance of JAK/STAT as a therapeutic target. JAK inhibitors or a selective STAT5 SH2 domain inhibitor induced cell death and ruxolitinib blocked T-cell neoplasia in vivo We conclude that enhanced STAT5A or STAT5B action both drive PTCL development, defining both STAT5 molecules as targets for therapeutic intervention.

Metabolic disease and ABHD6 alter the circulating bis(monoacylglycerol)phosphate profile in mice and humans.

Bis(monoacylglycerol)phosphate (BMP) is a phospholipid that is crucial for lipid degradation and sorting in acidic organelles. Genetic and drug-induced lysosomal storage disorders (LSDs) are associated with increased BMP concentrations in tissues and in the circulation. Data on BMP in disorders other than LSDs, however, are scarce, and key enzymes regulating BMP metabolism remain elusive. Here, we demonstrate that common metabolic disorders and the intracellular BMP hydrolase α/ß-hydrolase domain-containing 6 (ABHD6) affect BMP metabolism in mice and humans. In mice, dietary lipid overload strongly affects BMP concentration and FA composition in the liver and plasma, similar to what has been observed in LSDs. Notably, distinct changes in the BMP FA profile enable a clear distinction between lipid overload and drug-induced LSDs. Global deletion of ABHD6 increases circulating BMP concentrations but does not cause LSDs. In humans, nonalcoholic fatty liver disease and liver cirrhosis affect the serum BMP FA composition and concentration. Furthermore, we identified a patient with a loss-of-function mutation in the ABHD6 gene, leading to an altered circulating BMP profile. In conclusion, our results suggest that common metabolic diseases and ABHD6 affect BMP metabolism in mice and humans.

APMAP interacts with lysyl oxidase-like proteins, and disruption of Apmap leads to beneficial visceral adipose tissue expansion.

Adipocyte plasma membrane-associated protein (APMAP) has been described as an adipogenic factor in 3T3-L1 cells with unknown biochemical function; we therefore aimed to investigate the physiologic function of APMAP in vivo We generated Apmap-knockout mice and challenged them with an obesogenic diet to investigate their metabolic phenotype. We identified a novel truncated adipocyte-specific isoform of APMAP in mice that is produced by alternative transcription. Mice lacking the full-length APMAP protein, the only isoform that is expressed in humans, have an improved metabolic phenotype upon diet-induced obesity, indicated by enhanced insulin sensitivity, preserved glucose tolerance, increased respiratory exchange ratio, decreased inflammatory marker gene expression, and reduced adipocyte size. At the molecular level, APMAP interacts with the extracellular collagen cross-linking matrix proteins lysyl oxidase-like 1 and 3. On a high-fat diet, the expression of lysyl oxidase-like 1 and 3 is strongly decreased in Apmap-knockout mice, paralleled by reduced expression of profibrotic collagens and total collagen content in epididymal white adipose tissue, indicating decreased fibrotic potential. Together, our data suggest that APMAP is a novel regulator of extracellular matrix components, and establish that APMAP is a potential target to mitigate obesity-associated insulin resistance.-Pessentheiner, A. R., Huber, K., Pelzmann, H. J., Prokesch, A., Radner, F. P. W., Wolinski, H., Lindroos-Christensen, J., Hoefler, G., Rülicke, T., Birner-Gruenberger, R., Bilban, M., Bogner-Strauss, J. G. APMAP interacts with lysyl oxidase-like proteins, and disruption of Apmap leads to beneficial visceral adipose tissue expansion.

Sequestosome 1/p62 Protein Is Associated with Autophagic Removal of Excess Hepatic Endoplasmic Reticulum in Mice.

Xenobiotics exposure increases endoplasmic reticulum (ER) proliferation and cytochrome P-450 (CYP) induction to sustain metabolic requirements. Whether autophagy is essential for the removal of excess ER and CYP and whether an autophagy receptor is involved in this process in mammals remains elusive. In this study, we show that autophagy is induced in mouse livers after withdrawal of the hepatic mitogen 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP). Although isolated autophagosomes, autolysosomes, and lysosomes from mouse livers after withdrawal of TCPOBOP contained ER proteins, those in control mouse livers did not. Liver-specific Atg5 knockout mice had higher basal hepatic ER content that was further increased and sustained after withdrawal of TCPOBOP compared with wild-type mice. In addition to regulating ER degradation, our results also suggest that autophagy plays a role in regulating the homeostasis of hepatic CYP because blocking autophagy led to increased CYP2B10 accumulation either at the basal level or following TCPOBOP withdrawal. Furthermore, we found that the autophagy receptor protein sequestosome 1 (SQSTM1)/p62 is associated with the ER. After withdrawal of TCPOBOP, p62 knockout mice had increased ER content in the liver compared with wild-type mice. These results suggest that p62 may act as an autophagy receptor for the autophagic removal of excess ER in the mouse liver. Taken together, our results indicate that autophagy is important for the removal of excess ER and hepatic CYP enzymes in mouse livers, a process associated with the autophagy receptor protein p62.