An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells
Hereditary spastic paraplegias (HSPs) are a heterogeneous group of mono-genetic inherited neurological disorders, whose primary manifestation is the disruption of the pyramidal system, observed as a progressive impaired gait and leg spasticity in patients. Despite the large list of genes linked to t...
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| Format: | Article |
| Language: | English |
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MDPI AG
2024-03-01
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| Series: | Cells |
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| Online Access: | https://www.mdpi.com/2073-4409/13/7/577 |
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| author | M. Elena Garcia-Pardo Jeremy C. Simpson Niamh C. O’Sullivan |
| author_facet | M. Elena Garcia-Pardo Jeremy C. Simpson Niamh C. O’Sullivan |
| author_sort | M. Elena Garcia-Pardo |
| collection | DOAJ |
| description | Hereditary spastic paraplegias (HSPs) are a heterogeneous group of mono-genetic inherited neurological disorders, whose primary manifestation is the disruption of the pyramidal system, observed as a progressive impaired gait and leg spasticity in patients. Despite the large list of genes linked to this group, which exceeds 80 loci, the number of cellular functions which the gene products engage is relatively limited, among which endoplasmic reticulum (ER) morphogenesis appears central. Mutations in genes encoding ER-shaping proteins are the most common cause of HSP, highlighting the importance of correct ER organisation for long motor neuron survival. However, a major bottleneck in the study of ER morphology is the current lack of quantitative methods, with most studies to date reporting, instead, on qualitative changes. Here, we describe and apply a quantitative image-based screen to identify genetic modifiers of ER organisation using a mammalian cell culture system. An analysis reveals significant quantitative changes in tubular ER and dense sheet ER organisation caused by the siRNA-mediated knockdown of HSP-causing genes <i>ATL1</i> and <i>RTN2</i>. This screen constitutes the first attempt to examine ER distribution in cells in an automated and high-content manner and to detect genes which impact ER organisation. |
| first_indexed | 2024-04-24T10:47:38Z |
| format | Article |
| id | doaj.art-481c15ea008d4d289ca6e9b1b96eb7de |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-04-24T10:47:38Z |
| publishDate | 2024-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-481c15ea008d4d289ca6e9b1b96eb7de2024-04-12T13:16:26ZengMDPI AGCells2073-44092024-03-0113757710.3390/cells13070577An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human CellsM. Elena Garcia-Pardo0Jeremy C. Simpson1Niamh C. O’Sullivan2UCD School of Biomolecular and Biomedical Science, UCD Conway Institute, University College Dublin, 4 Dublin, IrelandCell Screening Laboratory, UCD School of Biology and Environmental Science, University College Dublin, 4 Dublin, IrelandUCD School of Biomolecular and Biomedical Science, UCD Conway Institute, University College Dublin, 4 Dublin, IrelandHereditary spastic paraplegias (HSPs) are a heterogeneous group of mono-genetic inherited neurological disorders, whose primary manifestation is the disruption of the pyramidal system, observed as a progressive impaired gait and leg spasticity in patients. Despite the large list of genes linked to this group, which exceeds 80 loci, the number of cellular functions which the gene products engage is relatively limited, among which endoplasmic reticulum (ER) morphogenesis appears central. Mutations in genes encoding ER-shaping proteins are the most common cause of HSP, highlighting the importance of correct ER organisation for long motor neuron survival. However, a major bottleneck in the study of ER morphology is the current lack of quantitative methods, with most studies to date reporting, instead, on qualitative changes. Here, we describe and apply a quantitative image-based screen to identify genetic modifiers of ER organisation using a mammalian cell culture system. An analysis reveals significant quantitative changes in tubular ER and dense sheet ER organisation caused by the siRNA-mediated knockdown of HSP-causing genes <i>ATL1</i> and <i>RTN2</i>. This screen constitutes the first attempt to examine ER distribution in cells in an automated and high-content manner and to detect genes which impact ER organisation.https://www.mdpi.com/2073-4409/13/7/577endoplasmic reticulumhigh-content imagingautomated image analysissiRNA screenhuman cells |
| spellingShingle | M. Elena Garcia-Pardo Jeremy C. Simpson Niamh C. O’Sullivan An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells Cells endoplasmic reticulum high-content imaging automated image analysis siRNA screen human cells |
| title | An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells |
| title_full | An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells |
| title_fullStr | An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells |
| title_full_unstemmed | An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells |
| title_short | An Automated Imaging-Based Screen for Genetic Modulators of ER Organisation in Cultured Human Cells |
| title_sort | automated imaging based screen for genetic modulators of er organisation in cultured human cells |
| topic | endoplasmic reticulum high-content imaging automated image analysis siRNA screen human cells |
| url | https://www.mdpi.com/2073-4409/13/7/577 |
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