bims-lymeca Biomed News
on Lysosome metabolism in cancer
Issue of 2022‒07‒17
four papers selected by
Harilaos Filippakis
University of New England

  1. Nature. 2022 Jul 13.
      Mechanistic target of rapamycin complex 1 (mTORC1) controls growth by regulating anabolic and catabolic processes in response to environmental cues, including nutrients1,2. Amino acids signal to mTORC1 through the Rag GTPases, which are regulated by several protein complexes, including GATOR1 and GATOR2. GATOR2, which has five components (WDR24, MIOS, WDR59, SEH1L and SEC13), is required for amino acids to activate mTORC1 and interacts with the leucine and arginine sensors SESN2 and CASTOR1, respectively3-5. Despite this central role in nutrient sensing, GATOR2 remains mysterious as its subunit stoichiometry, biochemical function and structure are unknown. Here we used cryo-electron microscopy to determine the three-dimensional structure of the human GATOR2 complex. We found that GATOR2 adopts a large (1.1 MDa), two-fold symmetric, cage-like architecture, supported by an octagonal scaffold and decorated with eight pairs of WD40 β-propellers. The scaffold contains two WDR24, four MIOS and two WDR59 subunits circularized via two distinct types of junction involving non-catalytic RING domains and α-solenoids. Integration of SEH1L and SEC13 into the scaffold through β-propeller blade donation stabilizes the GATOR2 complex and reveals an evolutionary relationship to the nuclear pore and membrane-coating complexes6. The scaffold orients the WD40 β-propeller dimers, which mediate interactions with SESN2, CASTOR1 and GATOR1. Our work reveals the structure of an essential component of the nutrient-sensing machinery and provides a foundation for understanding the function of GATOR2 within the mTORC1 pathway.
  2. Methods Mol Biol. 2022 ;2473 285-306
      Lysosomes are membrane-bound organelles that degrade diverse biomolecules and regulate a multitude of other essential processes including cell growth and metabolism, signaling, plasma membrane repair and infection. Such diverse functions of lysosomes are highly coordinated in space and time and are therefore tightly coupled to the directional transport of the organelles within the cytoplasm. Thus, robust quantitative assessments of lysosome positioning within the cell provide a valuable tool for researchers interested in understanding these multifunctional organelles. Here, we present point-by-point methodology to measure lysosome positioning by two straight forward and widely used techniques: shell analysis and line scan.
    Keywords:  ImageJ analysis; Line scan; Lysosome distribution; Lysosome positioning measurements; Lysosome transport; Organelle positioning; Shell analysis
  3. Autophagy. 2022 Jul 09.
      The conjugation of Atg8-family proteins with phospholipids on the double-membrane phagophore is one of the hallmarks of macroautopahgy/autophagy. However, in the past decades, Atg8-family proteins are also found on single-membrane structures, including the phagosome, endosome and lysosome. While the physiological importance of the non-canonical Atg8-family protein conjugation has been demonstrated, the mechanism of this process and the underlying regulation are still not very clear. In a recent paper, Hooper et al. found that during LC3-associated phagocytosis, reactive oxygen species are required for V-ATPase assembly, which is essential for the subsequent LC3 conjugation to the phagosome. Enhanced V-ATPase assembly and the direct engagement of ATG16L1 are also observed in a wide range of non-canonical Atg8-family protein conjugation processes, defining the V-ATPase and ATG16L1 as taking part in a common mechanism.
    Keywords:  ATG16L1; Atg8 conjugation; LAP; ROS; V-ATPase; non-canonical autophagy
  4. Mol Cancer Res. 2022 Jul 14. pii: mcr.22.0026. [Epub ahead of print]
      Exchange Proteins directly Activated by cAMP (EPACs) belong to a family of RAP guanine nucleotide exchange factors (RAPGEF). EPAC1/2 (RAPGEF3/4) activate RAP1 and the alternative cAMP signaling pathway. We previously showed that the differential growth response of primary and metastatic melanoma cells to cAMP is mediated by EPAC. However, the mechanisms responsible for this differential response to EPAC signaling are not understood. In this study, we show that pharmacological inhibition or siRNA-mediated knockdown of EPAC selectively inhibits the growth and survival of primary melanoma cells by downregulation of cell cycle proteins and inhibiting the cell cycle progression independent of ERK1/2 phosphorylation. EPAC inhibition results in upregulation of AKT phosphorylation but a downregulation of mTORC1 activity and its downstream effectors. We also show that EPAC regulates both glycolysis and oxidative phosphorylation, and production of mitochondrial reactive oxygen species, preferentially in primary melanoma cells. Employing a series of genetically matched primary and lymph node metastatic (LNM) melanoma cells, and distant organ metastatic melanoma (MM) cells, we show that the LNM and MM cells become progressively less responsive and refractory to EPAC inhibition suggesting loss of dependency on EPAC signaling correlates with melanoma progression. Analysis of TCGA dataset showed that lower RAPGEF3, RAPGEF4 mRNA expression in primary tumor is a predictor of better disease-free survival of patients diagnosed with primary melanoma suggesting that EPAC signaling facilitates tumor progression and EPAC is a useful prognostic marker. These data highlight EPAC signaling as a potential target for prevention of melanoma progression. Implications: This study establishes loss of dependency on EPAC-mTORC1 signaling as hallmark of primary melanoma evolution and targeting this escape mechanism is a promising strategy for metastatic melanoma.