Turn off MathJax
Article Contents
Liting Zhang, Yan Mo, Shimin Mo, Ming Xia, Chaoliang Wei. Simultaneous detection of dynamic calcium signaling and ERK activity in living cells. Biophysics Reports. doi: 10.52601/bpr.2023.230038
Citation: Liting Zhang, Yan Mo, Shimin Mo, Ming Xia, Chaoliang Wei. Simultaneous detection of dynamic calcium signaling and ERK activity in living cells. Biophysics Reports. doi: 10.52601/bpr.2023.230038

Simultaneous detection of dynamic calcium signaling and ERK activity in living cells

doi: 10.52601/bpr.2023.230038
More Information
  • Corresponding author: chaoliangwei@126.com (C. Wei)
  • Received Date: 17 November 2023
  • Accepted Date: 26 December 2023
  • Available Online: 26 February 2024
  • Calcium (Ca2+) is a universal second messenger in eukaryotic cells, and extracellular regulated protein kinases (ERK) are the core component of the mitogen-activated protein kinase (MAPK) signaling cascade. Both are involved in numerous physiological and pathological processes, such as organogenesis, tumorigenesis, proliferation, migration and apoptosis. Over the past decade, it has been found that calcium signaling can regulate the ERK activity through multiple mechanisms, and conversely, ERK signaling transduction can also affect the triggering and intensity of calcium signaling. However, there are few reports on how to perform real-time synchronous detection of these two signals. Here we described a method for dynamically and synchronously recording calcium signals and ERK activity in living cells, utilizing stable expression of multiple genetically-encoded probes and multi-channel synchronous detection technology using confocal microscopy. The protocol can be useful to address the spatiotemporal encoding dynamic mechanism of calcium signaling and ERK activity in single or multiple cells, and to reveal the interaction and causal characteristics of these two signals.

  • Liting Zhang, Yan Mo, Shimin Mo, Ming Xia and Chaoliang Wei declare that they have no conflict of interest.
    This article does not contain any studies with human or animal subjects performed by any of the authors.
    Liting Zhang and Yan Mo contributed equally to this work.

  • loading
  • Chen TW, Wardill TJ, Sun Y, Pulver SR, Renninger SL, Baohan A, Schreiter ER, Kerr RA, Orger MB, Jayaraman V, Looger LL, Svoboda K, Kim DS (2013) Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499: 295−300 doi: 10.1038/nature12354
    Grewal SS, Horgan AM, York RD, Withers GS, Banker GA, Stork PJ (2000) Neuronal calcium activates a Rap1 and B-Raf signaling pathway via the cyclic adenosine monophosphate-dependent protein kinase. J Biol Chem 275: 3722−3728 doi: 10.1074/jbc.275.5.3722
    Grienberger C, Konnerth A (2012) Imaging calcium in neurons. Neuron 73: 862−885 doi: 10.1016/j.neuron.2012.02.011
    Gudipaty SA, Lindblom J, Loftus PD, Redd MJ, Edes K, Davey CF, Krishnegowda V, Rosenblatt J (2017) Mechanical stretch triggers rapid epithelial cell division through Piezo1. Nature 543: 118−121 doi: 10.1038/nature21407
    Guo YJ, Pan WW, Liu SB, Shen ZF, Xu Y, Hu LL (2020) ERK/MAPK signalling pathway and tumorigenesis. Exp Ther Med 19: 1997−2007
    Hammad AS, Machaca K (2021) Store operated calcium entry in cell migration and cancer metastasis. Cells 10: 1246. https://doi.org/10.3390/cells10051246
    Heredia DJ, De Angeli C, Fedi C, Gould TW (2020) Calcium signaling in Schwann cells. Neurosci Lett 729: 134959. https://doi.org/10.1016/j.neulet.2020.134959
    Hirata E, Kiyokawa E (2016) Future perspective of single-molecule FRET biosensors and intravital FRET microscopy. Biophys J 111: 1103−1111 doi: 10.1016/j.bpj.2016.01.037
    Klepeis VE, Cornell-Bell A, Trinkaus-Randall V (2001) Growth factors but not gap junctions play a role in injury-induced Ca2 + waves in epithelial cells. J Cell Sci 114: 4185−4195 doi: 10.1242/jcs.114.23.4185
    Kosugi S, Hasebe M, Tomita M, Yanagawa H (2008) Nuclear export signal consensus sequences defined using a localization-based yeast selection system. Traffic 9: 2053−2062 doi: 10.1111/j.1600-0854.2008.00825.x
    Kudo T, Jeknić S, Macklin DN, Akhter S, Hughey JJ, Regot S, Covert MW (2018) Live-cell measurements of kinase activity in single cells using translocation reporters. Nat Protoc 13: 155−169 doi: 10.1038/nprot.2017.128
    Maryu G, Matsuda M, Aoki K (2016) Multiplexed fluorescence imaging of ERK and Akt activities and cell-cycle progression. Cell Struct Funct 41: 81−92 doi: 10.1247/csf.16007
    Miura H, Matsuda M, Aoki K (2014) Development of a FRET biosensor with high specificity for Akt. Cell Struct Funct 39: 9−20 doi: 10.1247/csf.13018
    Mottet D, Michel G, Renard P, Ninane N, Raes M, Michiels C (2002) ERK and calcium in activation of HIF-1. Ann N Y Acad Sci 973: 448−453 doi: 10.1111/j.1749-6632.2002.tb04681.x
    Nakamura A, Goto Y, Kondo Y, Aoki K (2021) Shedding light on developmental ERK signaling with genetically encoded biosensors. Development 148: dev199767. https://doi.org/10.1242/dev.199767
    Palmer AE, Tsien RY (2006) Measuring calcium signaling using genetically targetable fluorescent indicators. Nat Protoc 1: 1057−1065 doi: 10.1038/nprot.2006.172
    Paredes RM, Etzler JC, Watts LT, Zheng W, Lechleiter JD (2008) Chemical calcium indicators. Methods 46: 143−151 doi: 10.1016/j.ymeth.2008.09.025
    Regot S, Hughey JJ, Bajar BT, Carrasco S, Covert MW (2014) High-sensitivity measurements of multiple kinase activities in live single cells. Cell 157: 1724−1734 doi: 10.1016/j.cell.2014.04.039
    Roe MW, Lemasters JJ, Herman B (1990) Assessment of Fura-2 for measurements of cytosolic free calcium. Cell Calcium 11: 63−73 doi: 10.1016/0143-4160(90)90060-8
    Rosado JA, Sage SO (2002) The ERK cascade, a new pathway involved in the activation of store-mediated calcium entry in human platelets. Trends Cardiovasc Med 12: 229−234 doi: 10.1016/S1050-1738(02)00161-5
    Samson SC, Khan AM, Mendoza MC (2022) ERK signaling for cell migration and invasion. Front Mol Biosci 9: 998475. https://doi.org/10.3389/fmolb.2022.998475
    Smaili SS, Pereira GJ, Costa MM, Rocha KK, Rodrigues L, do Carmo LG, Hirata H, Hsu YT (2013) The role of calcium stores in apoptosis and autophagy. Curr Mol Med 13: 252−265 doi: 10.2174/156652413804810772
    Sukumaran P, Nascimento Da Conceicao V, Sun Y, Ahamad N, Saraiva LR, Selvaraj S, Singh BB (2021) Calcium signaling regulates autophagy and apoptosis. Cells 10: 2125. https://doi.org/10.3390/cells10082125
    Takada H, Furuya K, Sokabe M (2014) Mechanosensitive ATP release from hemichannels and Ca²⁺ influx through TRPC6 accelerate wound closure in keratinocytes. J Cell Sci 127: 4159−4171
    Yang S, Liu G (2017) Targeting the Ras/Raf/MEK/ERK pathway in hepatocellular carcinoma. Oncol Lett 13: 1041−1047 doi: 10.3892/ol.2017.5557
  • 加载中

Catalog

    通讯作者: 陈斌, bchen63@163.com
    • 1. 

      沈阳化工大学材料科学与工程学院 沈阳 110142

    1. 本站搜索
    2. 百度学术搜索
    3. 万方数据库搜索
    4. CNKI搜索

    Figures(5)  / Tables(1)

    Article Metrics

    Article views (195) PDF downloads(8) Cited by()
    Proportional views
    Related

    /

    DownLoad:  Full-Size Img  PowerPoint
    Return
    Return