Biology subjects

Takagi, E.

Publications and source records attributed to Takagi, E..

3 recordsLinked to original sources

bioRxiv2025

Impaired IFNγ responsiveness of lung monocyte-derived cells limits immunity to Mycobacterium tuberculosis

Lung mononuclear phagocyte (MNP) subsets differ in their ability to restrict Mycobacterium tuberculosis (Mtb) during chronic infection, yet the mechanisms underlying this difference are not well defined. Here, we show that CD11clo monocyte-derived cells (MNC1), the subset of lung cells that is most permissive for Mtb viability during chronic infection, express lower levels of interferon-gamma (IFN{gamma}) signaling proteins, resulting in reduced responses to IFN{gamma} compared to alveolar macrophages (AM) and CD11chi MNC2. Moreover, type I IFN signaling suppresses IFN{gamma}-mediated MHC-II expression, impairing antigen-specific CD4 T cell activation by MNC1 cells. Importantly, prior immunity conferred by contained Mtb infection enhances IFN{gamma} responsiveness of monocyte-derived cells, reducing bacterial burdens in lungs and within MNC subsets. Our findings indicate that heterogeneous IFN{gamma} responsiveness is exploited by Mtb for persistence in vivo. Overcoming or bypassing impaired IFN{gamma} responsiveness may guide the development of more effective TB vaccines and host-directed therapies.

immunology↗

bioRxiv2024

Coordinated Regulation of Renal Glucose Reabsorption and Gluconeogenesis by mTORC2 and Potassium

BackgroundThe kidney proximal tubule is uniquely responsible for reabsorption of filtered glucose and gluconeogenesis (GNG). Insulin stimulates glucose transport and suppresses GNG in the proximal tubule, however, the signaling mechanisms and coordinated regulation of these processes remain poorly understood. The kinase complex mTORC2 is critical for regulation of growth, metabolism, solute transport, and electrolyte homeostasis in response to a wide array of inputs. Here we examined its role in the regulation of renal glucose reabsorption and GNG. MethodsRictor, an essential component of mTORC2, was knocked out using the Pax8-LC1 system to generate inducible tubule specific Rictor knockout (TRKO) mice. These animals were subjected to fasting, refeeding, and variation in dietary K+. Metabolic parameters including glucose homeostasis and renal function were assessed in balance cages. Kidneys and livers were also harvested for molecular analysis of gluconeogenic enzymes, mTORC2-regulated targets, and plasma membrane glucose transporters. ResultsOn a normal chow diet, TRKO mice had marked glycosuria despite indistinguishable blood glucose relative to WT controls. Kidney plasma membrane showed lower SGLT2 and SGLT1 in the fed state, supporting reduced renal glucose reabsorption. Additional metabolic testing provided evidence for renal insulin resistance with elevated fasting insulin, impaired pyruvate tolerance, elevated hemoglobin A1c, and increased renal gluconeogenic enzymes in the fasted and fed states. These effects were correlated with reduced downstream phosphorylation of Akt and the transcription factor FOXO4, identifying a novel role of FOXO4 in the kidney. Interestingly, high dietary K+ prevented glycosuria and excessive GNG in TRKO mice, despite persistent reduction in mTORC2 substrate phosphorylation. ConclusionRenal tubule mTORC2 is critical for coordinated regulation of sodium-glucose cotransport by SGLT2 and SGLT1 as well as renal GNG. Dietary K+ promotes glucose reabsorption and suppresses GNG independently of insulin signaling and mTORC2, potentially providing an alternative signaling mechanism in states of insulin resistance. SIGNIFICANCE STATEMENTThe kidney contributes to regulation of blood glucose through reabsorption of filtered glucose and gluconeogenesis. This study shows that mTORC2 and dietary potassium coordinate the regulation of sodium-glucose cotransport and glucose production in the kidney via independent mechanisms. New insights into the regulation of these processes in the kidney offer promising implications for diabetes mellitus management and treatment.

physiology↗

bioRxiv2022

Structural features of mTORC2 that control substrate-specific activities.

mTORC2 is a multi-subunit kinase complex that is central to multiple essential signaling pathways. Two core subunits, Rictor and mSin1 distinguish it from its relative, mTORC1 and support context-dependent phosphorylation of its substrates. mTORC2 structures have been determined previously, however, important questions remain, particularly regarding structural determinants of substrate specificity and context dependent activities. We used cryo-EM to obtain high resolution structures of the human mTORC2 apo-complex, as well as structures in the presence of substrates, Akt and SGK1. Specific predictions suggested by substrate-induced structural changes were tested in functional assays. First, side chain interactions between Rictor and mTOR that prevent recruitment of mTORC1 substrates and confer resistance to the mTORC1 inhibitor rapamycin were visualized for the first time in the apo-state, demonstrating the steric occlusion that prevents mTORC2 interaction with mTORC1 substrates and rapamycin. Also in the apo-state, mSin1 was seen to form extensive contacts with Rictor, including a pair of short -helices nestled between two Rictor helical repeat clusters, followed by an extended strand, which makes multiple weak contacts with Rictor helical cluster 1. In co-complex structures, SGK1, but not Akt, markedly altered the conformation of the mSin1 N-terminal extended strand, disrupting multiple weak interactions while inducing a large rotation of mSin1/Arg-83, which comes to interact with a negative patch within Rictor. Mutation of Arg-83 to Ala selectively disrupted mTORC2 dependent phosphorylation of SGK1 but not of Akt, supporting context-dependent substrate selection. These findings provide new structural and functional insights into mTORC2 specificity and context-dependent activities.

biochemistry↗