A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus.
We describe the use of a secreted form of Aequoria victoria green fluorescent protein (secGFP) in a non-invasive live cell assay of membrane traffic in Arabidopsis thaliana. We show that in comparison to GFP-HDEL, which accumulates in the endoplasmic reticulum (ER), secGFP generates a weak fluoresce...
| Автори: | , , , |
|---|---|
| Формат: | Journal article |
| Мова: | English |
| Опубліковано: |
2004
|
| _version_ | 1826294100606844928 |
|---|---|
| author | Zheng, H Kunst, L Hawes, C Moore, I |
| author_facet | Zheng, H Kunst, L Hawes, C Moore, I |
| author_sort | Zheng, H |
| collection | OXFORD |
| description | We describe the use of a secreted form of Aequoria victoria green fluorescent protein (secGFP) in a non-invasive live cell assay of membrane traffic in Arabidopsis thaliana. We show that in comparison to GFP-HDEL, which accumulates in the endoplasmic reticulum (ER), secGFP generates a weak fluorescence signal when transported to the apoplast. The fluorescence of secGFP in the apoplast can be increased by growth of seedlings on culture medium buffered at pH 8.1, suggesting that apoplastic pH is responsible, at least in part, for the low fluorescence intensity of seedlings expressing secGFP. Inhibition of secGFP transport between the ER and plasma membrane (PM), either by Brefeldin A (BFA) treatment or by genetic intervention results in increased intracellular secGFP accumulation accompanied by an increase in the secGFP fluorescence intensity. secGFP thus provides a valuable tool for forward and reverse genetic analysis of membrane traffic and endomembrane organisation in Arabidopsis. Using this assay for quantitative sublethal perturbation of secGFP transport, we identify a role for root hair defective 3 (RHD3) in transport of secreted and Golgi markers between the ER and the Golgi apparatus. |
| first_indexed | 2024-03-07T03:40:25Z |
| format | Journal article |
| id | oxford-uuid:bdb4d52b-db8a-4af0-a5bb-9d787c946bc9 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T03:40:25Z |
| publishDate | 2004 |
| record_format | dspace |
| spelling | oxford-uuid:bdb4d52b-db8a-4af0-a5bb-9d787c946bc92022-03-27T05:33:47ZA GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:bdb4d52b-db8a-4af0-a5bb-9d787c946bc9EnglishSymplectic Elements at Oxford2004Zheng, HKunst, LHawes, CMoore, IWe describe the use of a secreted form of Aequoria victoria green fluorescent protein (secGFP) in a non-invasive live cell assay of membrane traffic in Arabidopsis thaliana. We show that in comparison to GFP-HDEL, which accumulates in the endoplasmic reticulum (ER), secGFP generates a weak fluorescence signal when transported to the apoplast. The fluorescence of secGFP in the apoplast can be increased by growth of seedlings on culture medium buffered at pH 8.1, suggesting that apoplastic pH is responsible, at least in part, for the low fluorescence intensity of seedlings expressing secGFP. Inhibition of secGFP transport between the ER and plasma membrane (PM), either by Brefeldin A (BFA) treatment or by genetic intervention results in increased intracellular secGFP accumulation accompanied by an increase in the secGFP fluorescence intensity. secGFP thus provides a valuable tool for forward and reverse genetic analysis of membrane traffic and endomembrane organisation in Arabidopsis. Using this assay for quantitative sublethal perturbation of secGFP transport, we identify a role for root hair defective 3 (RHD3) in transport of secreted and Golgi markers between the ER and the Golgi apparatus. |
| spellingShingle | Zheng, H Kunst, L Hawes, C Moore, I A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title | A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title_full | A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title_fullStr | A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title_full_unstemmed | A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title_short | A GFP-based assay reveals a role for RHD3 in transport between the endoplasmic reticulum and Golgi apparatus. |
| title_sort | gfp based assay reveals a role for rhd3 in transport between the endoplasmic reticulum and golgi apparatus |
| work_keys_str_mv | AT zhengh agfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT kunstl agfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT hawesc agfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT moorei agfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT zhengh gfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT kunstl gfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT hawesc gfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus AT moorei gfpbasedassayrevealsaroleforrhd3intransportbetweentheendoplasmicreticulumandgolgiapparatus |