Stimulation of the Gαq/phospholipase Cβ1 signaling pathway returns differentiated cells to a stem‐like state

Abstract

Phospholipase Cβ1 is activated by Gαq to generate calcium signals in response to hormones and neurotransmitters. Besides carrying out this plasma membrane function, PLCβ1 has a cytosolic population that helps to drive the differentiation of PC12 cells by inhibiting a nuclease that promotes RNA‐induced silencing (C3PO). Here, we show that down‐regulating PLCβ1 or reducing its cytosolic population by activating Gαq to localize it to the plasma membrane returns differentiated PC12 and SK‐N‐SH cells to an undifferentiated state. In this state, PC12 cells have a spherical morphology, resume proliferation, and express the stem cell transcription factors nanog and Oct4. Similar changes are seen when C3PO is down‐regulated. This return to a stem‐like state is accompanied by shifts in multiple miR populations. Surprisingly, de‐differentiation can be induced by extended stimulation of Gαq where cells return to a spherical morphology and levels of specific miRs return to their undifferentiated values. In complementary studies, we followed the real‐time hydrolysis of a fluorescent‐tagged miR in cells where PLCβ1 or C3PO were down‐regulated in PC12 cells and find substantial differences in miR processing in the undifferentiated and differentiated states. Taken together, our studies suggest that PLCβ1, through its ability to regulate C3PO and endogenous miR populations, mediates the differentiation of two types of cultured neuronal cells.

Activation of Gαq sequesters specific transcripts into Ago2 particles

“…PLCβ1 is associated with loss of proliferation and differentiation 35,36 and these effects are likely to be due to its interaction with C3PO and Ago2, as well as mediating calcium signals. We have previously identified miRs associated PLCβ1 down-regulation in PC12 cells 37 and found . that down-regulating PLCβ1 levels results in substantial reduction in miR101a and miR101b, which may regulate ATP synthase 5B 38 and miR-22 levels by 20%, which regulates Chgb 39 .…”

Neurotransmitter Activation Sequesters Specific mRNAs Into Stress Granules

“…PLCβ1 is associated with loss of proliferation and differentiation [38,39] and these effects are likely to be due to its interaction with C3PO and Ago2, as well as mediating calcium signals. We have previously identi ed miRs associated PLCβ1 down-regulation in PC12 cells [40] and found that down-regulating PLCβ1 levels results in substantial reduction in miR101a and miR101b, which may regulate ATP synthase 5B [41]and miR-22 levels by 20%, which regulates Chgb [42]. The loss of these miRs, along with the observation that loss of cytosolic PLCβ1 or sustained neurotransmitter activation of Gαq returns PC12 cells to the undifferentiated state, may indicate that the increased number of stress granules may sequester mRNAs important for de ning cell phenotype.…”

“…Western blotting: We used an establish procedure [40] with modi cations. Samples were grown in 100 mm dishes plates and were treated with MG132 at a nal concentration of 250 nM 24 hours before collection.…”

“…Phase modulation FLIM measurements were performed on the dual-channel confocal fast FLIM (Alba version 5, ISS Inc.) using a two-photon titanium-sapphire laser and a Nikon Eclipse Ti-U inverted microscope as described [40]. The lifetime of the laser was calibrated each time before experiments by measuring the lifetime of Atto 425 in water with a lifetime of 3.61 ns at 80, 160, and 240 MHz.…”

Effect of PLC-β1/CaM signaling pathway mediated by AT1R on the occurrence and development of hepatocellular carcinoma

“…Extracellular factors such as Ang II, epinephrine, histamine and bradykinin can bind to corresponding receptors to activate PLC-β1 protein, further promote CaM expression and conformational changes in cells, thus regulating intracellular Ca 2+ concentration and calcium kinase activity, producing a series of biological effects (Fig. 2 ) [ 17 ]. Recent studies have found that the contents of PLC-β1 and CaM in HCC, colon cancer and breast cancer tissues are significantly increased, which can promote the growth, invasion and movement of tumor cells [ 18 – 20 ].…”