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Lysosomes serve as cellular degradation and signaling centers that coordinate the turnover of macromolecules with cell metabolism. The adaptation of cellular lysosome content and activity via the induction of lysosome biogenesis is therefore key to cell physiology and to counteract disease. Previous work has established a pathway for the induction of lysosome biogenesis in signaling-inactive starved cells that is based on the repression of mTORC1-mediated nutrient signaling. How lysosomal biogenesis is facilitated in signaling-active fed cells is poorly understood. A recent study by Malek et al. (2022) partially fills this gap by unraveling a nutrient signaling-independent pathway for lysosome biogenesis that operates in signaling-active cells. This pathway involves the receptor-mediated activation of phospholipase C, inositol (1,4,5)-triphosphate (IP3)-triggered release of calcium ions from endoplasmic reticulum stores, and the calcineurin-induced activation of transcription factor EB (TFEB) and its relative TFE3 to induce lysosomal gene expression independent of calcium in the lysosome lumen. These findings contribute to our understanding of how lysosome biogenesis and function are controlled in response to environmental changes and cell signaling and may conceivably be of relevance for our understanding and the treatment of lysosome-related diseases as well as for aging and neurodegeneration. [ABSTRACT FROM AUTHOR]

Abstract: Calcium (Ca2+) is involved in a myriad of cellular functions in the brain including synaptic plasticity. However, the role of intracellular Ca2+ stores in memory processing remains poorly defined. The current study explored a role for glutamate-dependent intracellular Ca2+ release in memory processing via blockade of metabotropic glutamate receptor subtype 1 (mGluR1) and inositol (1,4,5)-trisphosphate receptors (IP3Rs). Using a single-trial discrimination avoidance task developed for the young chick, administration of the specific and potent mGluR1 antagonist JNJ16259685 (500nM, immediately post-training, ic), or the IP3R antagonist Xestospongin C (5μM, immediately post-training, ic), impaired retention from 90min post-training. These findings are consistent with mGluR1 activating IP3Rs to release intracellular Ca2+ required for long-term memory formation and have been interpreted within an LTP2 model. The consequences of different patterns of retention loss following ryanodine receptor (RyR) and IP3R inhibition are discussed. [Copyright &y& Elsevier]

Abstract: In the human lens, stimulation of tyrosine-kinase coupled growth factor receptors such as epidermal growth factor receptor (EGFR) can induce calcium release from endoplasmic reticulum (ER) stores. The present study investigated the impact of calcium store inactivation on EGFR signalling, cell growth and death in a well-characterised human lens cell line (FHL124). FHL124 cells were routinely cultured in Eagle''s minimum essential medium (EMEM) supplemented with 10% foetal calf serum (FCS) and seeded on 24-well plates (DNA and protein synthesis), tissue culture dishes (growth assay, western immunoblot), and glass coverslips (immunocytochemistry). DNA and protein synthesis rates were quantified by measuring the incorporation of 3H-thymidine and 35S-methionine into FHL124 cells in serum-free EMEM or EMEM supplemented with thapsigargin (Tg) (100nm and 1μm). Longer-term growth was assessed by quantifying the increase in area over time of a circular patch of seeded cells. EGFR was identified using anti-EGFR mouse monoclonal antibody and visualised by fluorescence microscopy with ALEXA 488 conjugated secondary antibody. Programmed cell death was determined by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay method. Activation of the mitogen-activated protein kinase (MAPK) signalling protein extracellular signal-regulated kinase (ERK) and the cell cycle proteins CDK2 and P27kip1 were detected by western immunoblot techniques. Inactivation by ≥100nm Tg inhibited both protein and DNA synthesis although the effect on the latter was greatest. The cell cycle activator CDK2 was reduced by Tg, while the inhibitor P27kip1 was increased along with the percentage of apoptotic cells. A single, maximal epidermal growth factor (EGF) (10ngml−1) exposure induced receptor internalization and increased ERK phosphorylation. Both internalisation and ERK activation were unaffected by the presence of Tg. However, reduced internalisation and ERK activation followed repeated EGF applications in the presence of Tg. Additionally, ERK activation by submaximal EGF concentrations was reduced by store depletion. An intact endoplasmic reticulum calcium store therefore plays a significant role in human lens cell survival and growth. [Copyright &y& Elsevier]

Cytosolic Ca2+ controls a wide range of cellular events. The versatility of this second messenger depends on its ability to form diverse spatial and temporal patterns, including waves and oscillations. Ca2+-signaling patterns are thought to be determined in part by the subcellular distribution of inositol (1,4,5)-trisphosphate receptors [Ins(1,4,5)P3Rs] but little is currently known about how the localization of the Ins(1,4,5)P3R itself is regulated. Here, we report that the recruitment of GFP-tagged Ins(1,4,5)P3Rs in the vicinity of tight junctions in Madin-Darby canine kidney (MDCK) cells requires the N-terminal domain. Stable expression of this domain in polarized MDCK cells induced a flattened morphology, affected cytokinesis, accelerated cell migration in response to monolayer wounding and interfered with the cortical targeting of myosin IIA. In addition, downregulation of myosin IIA in polarized MDCK cells was found to mimic the effects of stable expression of the N-terminal part of Ins(1,4,5)P3R on cell shape and to alter localization of endogenous Ins(1,4,5)P3Rs. Taken together, these results support a model in which the recruitment of Ins(1,4,5)P3Rs at the apex of the lateral membrane in polarized MDCK cells, involves myosin IIA and might be important for the regulation of cortical actin dynamics. [ABSTRACT FROM AUTHOR]

Inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] originating in the vascular smooth-muscle cells (VSMCs) has been shown to modulate the Ca2+ stores in endothelial cells (ECs). However, the reverse is not found, suggesting that Ins(1,4,5)P3 movement might be unidirectional across gap junctions at the myoendothelial junction (MEJ), or that distribution of the Ins(1,4,5)P3 receptor [Ins(1,4,5)P3-R] is different between the two cell types. To study trans-junctional communication at the MEJ, we used a vascular-cell co-culture model system and selectively modified the connexin composition in gap junctions in the two cell types. We found no correlation between modification of connexin expression and Ins(1,4,5)P3 signaling between ECs and VSMCs. We next explored the distribution of Ins(1,4,5)P3-R isoforms in the two cell types and found that Ins(1,4,5)P3-R1 was selectively localized to the EC side of the MEJ. Using siRNA, selective knockdown of Ins(1,4,5)P3-R1 in ECs eliminated the secondary Ins(1,4,5)P3-induced response in these cells. By contrast, siRNA knockdown of Ins(1,4,5)P3-R2 or Ins(1,4,5)P3-R3 in ECs did not alter the EC response to VSMC stimulation. The addition of 5-phosphatase inhibitor (5-PI) to ECs that were transfected with Ins(1,4,5)P3-R1 siRNA rescued the Ins(1,4,5)P3 response, indicating that metabolic degradation of Ins(1,4,5)P3 is an important part of EC-VSMC coupling. To test this concept, VSMCs were loaded with 5-PI and BAPTA-loaded ECs were stimulated, inducing an Ins(1,4,5)P3-mediated response in VSMCs; this indicated that Ins(1,4,5)P3 is bidirectional across the gap junction at the MEJ. Therefore, localization of Ins(1,4,5)P3-R1 on the EC side of the MEJ allows the ECs to respond to Ins(1,4,5)P3 from VSMCs, whereas Ins(1,4,5)P3 moving from ECs to VSMCs is probably metabolized before binding to a receptor. This data implicates the MEJ as being a unique cell-signaling domain in the vasculature. [ABSTRACT FROM AUTHOR]