ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons
Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in live human neurons and to shed new light on the role of dysfunctional transport in neurodegenerative disorders. Here, we describe an approach for live imaging of axonal and dendritic transport...
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| Veröffentlicht in: | Traffic (Copenhagen, Denmark) Denmark), 2020-01, Vol.21 (1), p.138-155 |
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| creator | Boecker, Clemens Alexander Olenick, Mara A. Gallagher, Elizabeth R. Ward, Michael E. Holzbaur, Erika L. F. |
| description | Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in live human neurons and to shed new light on the role of dysfunctional transport in neurodegenerative disorders. Here, we describe an approach for live imaging of axonal and dendritic transport in iPSC‐derived cortical neurons. We use transfection and transient expression of genetically‐encoded fluorescent markers to characterize the motility of Rab‐positive vesicles, including early, late and recycling endosomes, as well as autophagosomes and mitochondria in iPSC‐derived neurons. Comparing transport parameters of these organelles with data from primary rat hippocampal neurons, we uncover remarkable similarities. In addition, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study the transport of endogenously labeled vesicles, as a parallel approach to the transient overexpression of fluorescently labeled organelle markers.
Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in human neurons. Here, we describe an approach for live‐imaging axonal and dendritic transport in iPSC‐derived cortical neurons. We characterize the motility of endosomes, autophagosomes and mitochondria. Comparing transport parameters of these organelles with data from rat hippocampal neurons, we uncover remarkable similarities. Furthermore, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study transport of endogenously labeled vesicles. |
| doi_str_mv | 10.1111/tra.12701 |
| format | Article |
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Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in human neurons. Here, we describe an approach for live‐imaging axonal and dendritic transport in iPSC‐derived cortical neurons. We characterize the motility of endosomes, autophagosomes and mitochondria. Comparing transport parameters of these organelles with data from rat hippocampal neurons, we uncover remarkable similarities. Furthermore, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study transport of endogenously labeled vesicles.</description><identifier>ISSN: 1398-9219</identifier><identifier>EISSN: 1600-0854</identifier><identifier>DOI: 10.1111/tra.12701</identifier><identifier>PMID: 31603614</identifier><language>eng</language><publisher>Former Munksgaard: John Wiley & Sons A/S</publisher><subject>Animals ; Axonal transport ; Axons - metabolism ; Biological Transport ; Cells, Cultured ; Dendritic transport ; Endosomes ; Fluorescent indicators ; Green fluorescent protein ; Hippocampus ; Induced Pluripotent Stem Cells ; Inhibitory postsynaptic potentials ; Intracellular ; iPSCs ; iPSC‐derived neurons ; live imaging ; Membrane proteins ; Mitochondria ; Neurodegenerative diseases ; Neurons ; NGN2 ; Organelles ; Phagosomes ; Pluripotency ; Protein turnover ; Rats ; Stem cell transplantation ; Stem cells ; Transfection ; Vesicles</subject><ispartof>Traffic (Copenhagen, Denmark), 2020-01, Vol.21 (1), p.138-155</ispartof><rights>2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd</rights><rights>2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.</rights><rights>2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3881-d21fe5e29b05c25bc197ff8bff5fa6fa5cb299cb598594ab4035d87d522e37873</citedby><cites>FETCH-LOGICAL-c3881-d21fe5e29b05c25bc197ff8bff5fa6fa5cb299cb598594ab4035d87d522e37873</cites><orcidid>0000-0001-5389-4114</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1111%2Ftra.12701$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1111%2Ftra.12701$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,776,780,1411,1427,27901,27902,45550,45551,46384,46808</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/31603614$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Boecker, Clemens Alexander</creatorcontrib><creatorcontrib>Olenick, Mara A.</creatorcontrib><creatorcontrib>Gallagher, Elizabeth R.</creatorcontrib><creatorcontrib>Ward, Michael E.</creatorcontrib><creatorcontrib>Holzbaur, Erika L. F.</creatorcontrib><title>ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons</title><title>Traffic (Copenhagen, Denmark)</title><addtitle>Traffic</addtitle><description>Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in live human neurons and to shed new light on the role of dysfunctional transport in neurodegenerative disorders. Here, we describe an approach for live imaging of axonal and dendritic transport in iPSC‐derived cortical neurons. We use transfection and transient expression of genetically‐encoded fluorescent markers to characterize the motility of Rab‐positive vesicles, including early, late and recycling endosomes, as well as autophagosomes and mitochondria in iPSC‐derived neurons. Comparing transport parameters of these organelles with data from primary rat hippocampal neurons, we uncover remarkable similarities. In addition, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study the transport of endogenously labeled vesicles, as a parallel approach to the transient overexpression of fluorescently labeled organelle markers.
Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in human neurons. Here, we describe an approach for live‐imaging axonal and dendritic transport in iPSC‐derived cortical neurons. We characterize the motility of endosomes, autophagosomes and mitochondria. Comparing transport parameters of these organelles with data from rat hippocampal neurons, we uncover remarkable similarities. Furthermore, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study transport of endogenously labeled vesicles.</description><subject>Animals</subject><subject>Axonal transport</subject><subject>Axons - metabolism</subject><subject>Biological Transport</subject><subject>Cells, Cultured</subject><subject>Dendritic transport</subject><subject>Endosomes</subject><subject>Fluorescent indicators</subject><subject>Green fluorescent protein</subject><subject>Hippocampus</subject><subject>Induced Pluripotent Stem Cells</subject><subject>Inhibitory postsynaptic potentials</subject><subject>Intracellular</subject><subject>iPSCs</subject><subject>iPSC‐derived neurons</subject><subject>live imaging</subject><subject>Membrane proteins</subject><subject>Mitochondria</subject><subject>Neurodegenerative diseases</subject><subject>Neurons</subject><subject>NGN2</subject><subject>Organelles</subject><subject>Phagosomes</subject><subject>Pluripotency</subject><subject>Protein turnover</subject><subject>Rats</subject><subject>Stem cell transplantation</subject><subject>Stem cells</subject><subject>Transfection</subject><subject>Vesicles</subject><issn>1398-9219</issn><issn>1600-0854</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2020</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kc1KxDAQgIMo7rp68AUk4EUPdfPTbBNvq_gHC4Ks55K2yVJpm5q06t58BJ_RJ3HWXT0IhkAm4ZuPyQxCh5ScUVjjzuszyhJCt9CQTgiJiBTxNsRcyUgxqgZoL4QnQggTcbyLBhwoPqHxENVz56oL93aOZ-WLwXe1XpTNAjuLywa0uamqvtIeO7_QDVwMhtcmtM53QMAu-twUuK16X7auM02HQ2dqvEr8fP8ojAdtgRvTe9eEfbRjdRXMweYcocfrq_nlbTS7v7m7nM6inEtJo4JRa4RhKiMiZyLLqUqslZm1wuqJ1SLPmFJ5JpQUKtZZTLgoZFIIxgxPZMJH6GTtbb177k3o0roMq5LgD64PKeNEEE4SSByh4z_ok-t9A9UBxSSLFVUCqNM1lXsXgjc2bX1Za79MKUlXM0ihLen3DIA92hj7rDbFL_nTdADGa-C1rMzyf1M6f5iulV_IvpJ5</recordid><startdate>202001</startdate><enddate>202001</enddate><creator>Boecker, Clemens Alexander</creator><creator>Olenick, Mara A.</creator><creator>Gallagher, Elizabeth R.</creator><creator>Ward, Michael E.</creator><creator>Holzbaur, Erika L. F.</creator><general>John Wiley & Sons A/S</general><general>Wiley Subscription Services, Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QP</scope><scope>7TK</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0001-5389-4114</orcidid></search><sort><creationdate>202001</creationdate><title>ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons</title><author>Boecker, Clemens Alexander ; Olenick, Mara A. ; Gallagher, Elizabeth R. ; Ward, Michael E. ; Holzbaur, Erika L. F.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3881-d21fe5e29b05c25bc197ff8bff5fa6fa5cb299cb598594ab4035d87d522e37873</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2020</creationdate><topic>Animals</topic><topic>Axonal transport</topic><topic>Axons - metabolism</topic><topic>Biological Transport</topic><topic>Cells, Cultured</topic><topic>Dendritic transport</topic><topic>Endosomes</topic><topic>Fluorescent indicators</topic><topic>Green fluorescent protein</topic><topic>Hippocampus</topic><topic>Induced Pluripotent Stem Cells</topic><topic>Inhibitory postsynaptic potentials</topic><topic>Intracellular</topic><topic>iPSCs</topic><topic>iPSC‐derived neurons</topic><topic>live imaging</topic><topic>Membrane proteins</topic><topic>Mitochondria</topic><topic>Neurodegenerative diseases</topic><topic>Neurons</topic><topic>NGN2</topic><topic>Organelles</topic><topic>Phagosomes</topic><topic>Pluripotency</topic><topic>Protein turnover</topic><topic>Rats</topic><topic>Stem cell transplantation</topic><topic>Stem cells</topic><topic>Transfection</topic><topic>Vesicles</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Boecker, Clemens Alexander</creatorcontrib><creatorcontrib>Olenick, Mara A.</creatorcontrib><creatorcontrib>Gallagher, Elizabeth R.</creatorcontrib><creatorcontrib>Ward, Michael E.</creatorcontrib><creatorcontrib>Holzbaur, Erika L. 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F.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons</atitle><jtitle>Traffic (Copenhagen, Denmark)</jtitle><addtitle>Traffic</addtitle><date>2020-01</date><risdate>2020</risdate><volume>21</volume><issue>1</issue><spage>138</spage><epage>155</epage><pages>138-155</pages><issn>1398-9219</issn><eissn>1600-0854</eissn><abstract>Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in live human neurons and to shed new light on the role of dysfunctional transport in neurodegenerative disorders. Here, we describe an approach for live imaging of axonal and dendritic transport in iPSC‐derived cortical neurons. We use transfection and transient expression of genetically‐encoded fluorescent markers to characterize the motility of Rab‐positive vesicles, including early, late and recycling endosomes, as well as autophagosomes and mitochondria in iPSC‐derived neurons. Comparing transport parameters of these organelles with data from primary rat hippocampal neurons, we uncover remarkable similarities. In addition, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study the transport of endogenously labeled vesicles, as a parallel approach to the transient overexpression of fluorescently labeled organelle markers.
Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in human neurons. Here, we describe an approach for live‐imaging axonal and dendritic transport in iPSC‐derived cortical neurons. We characterize the motility of endosomes, autophagosomes and mitochondria. Comparing transport parameters of these organelles with data from rat hippocampal neurons, we uncover remarkable similarities. Furthermore, we generated lysosomal‐associated membrane protein 1 (LAMP1)‐enhanced green fluorescent protein (EGFP) knock‐in iPSCs and show that knock‐in neurons can be used to study transport of endogenously labeled vesicles.</abstract><cop>Former Munksgaard</cop><pub>John Wiley & Sons A/S</pub><pmid>31603614</pmid><doi>10.1111/tra.12701</doi><tpages>18</tpages><orcidid>https://orcid.org/0000-0001-5389-4114</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | Animals Axonal transport Axons - metabolism Biological Transport Cells, Cultured Dendritic transport Endosomes Fluorescent indicators Green fluorescent protein Hippocampus Induced Pluripotent Stem Cells Inhibitory postsynaptic potentials Intracellular iPSCs iPSC‐derived neurons live imaging Membrane proteins Mitochondria Neurodegenerative diseases Neurons NGN2 Organelles Phagosomes Pluripotency Protein turnover Rats Stem cell transplantation Stem cells Transfection Vesicles |
| title | ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons |
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