Clathrin-Mediated endocytosis, post-endocytic trafficking and their regulatory controls in plants

M. Narasimhan, Clathrin-Mediated Endocytosis, Post-Endocytic Trafficking and Their Regulatory Controls in Plants , IST Austria, 2019.

Download
OA Supplementary_movie_1.avi 5.40 MB
OA 3.7_supplementary_movie_10.avi 5.93 MB
OA 3.7_supplementary_movie_9.avi 9.57 MB
All

Thesis | Published | English
Department
Series Title
IST Austria Thesis
Abstract
Clathrin-Mediated Endocytosis (CME) is an aspect of cellular trafficking that is constantly regulated for mediating developmental and physiological responses. The main aim of my thesis is to decipher the basic mechanisms of CME and post-endocytic trafficking in the whole multicellular organ systems of Arabidopsis. The first chapter of my thesis describes the search for new components involved in CME. Tandem affinity purification was conducted using CLC and its interacting partners were identified. Amongst the identified proteins were the Auxilin-likes1 and 2 (Axl1/2), putative uncoating factors, for which we made a full functional analysis. Over-expression of Axl1/2 causes extreme modifications in the dynamics of the machinery proteins and inhibition of endocytosis altogether. However the loss of function of the axl1/2 did not present any cellular or physiological phenotype, meaning Auxilin-likes do not form the major uncoating machinery. The second chapter of my thesis describes the establishment/utilisation of techniques to capture the dynamicity and the complexity of CME and post-endocytic trafficking. We have studied the development of endocytic pits at the PM – specifically, the mode of membrane remodeling during pit development and the role of actin in it, given plant cells possess high turgor pressure. Utilizing the improved z-resolution of TIRF and VAEM techniques, we captured the time-lapse of the endocytic events at the plasma membrane; and using particle detection software, we quantitatively analysed all the endocytic trajectories in an unbiased way to obtain the endocytic rate of the system. This together with the direct analysis of cargo internalisation from the PM provided an estimate on the endocytic potential of the cell. We also developed a methodology for ultrastructural analysis of different populations of Clathrin-Coated Structures (CCSs) in both PM and endomembranes in unroofed protoplasts. Structural analysis, together with the intensity profile of CCSs at the PM show that the mode of CCP development at the PM follows ‘Constant curvature model’; meaning that clathrin polymerisation energy is a major contributing factor of membrane remodeling. In addition, other analyses clearly show that actin is not required for membrane remodeling during invagination or any other step of CCP development, despite the prevalent high turgor pressure. However, actin is essential in orchestrating the post-endocytic trafficking of CCVs facilitating the EE formation. We also observed that the uncoating process post-endocytosis is not immediate; an alternative mechanism of uncoating – Sequential multi-step process – functions in the cell. Finally we also looked at one of the important physiological stimuli modulating the process – hormone, auxin. auxin has been known to influence CME before. We have made a detailed study on the concentration-time based effect of auxin on the machinery proteins, CCP development, and the specificity of cargoes endocytosed. To this end, we saw no general effect of auxin on CME at earlier time points. However, very low concentration of IAA, such as 50nM, accelerates endocytosis of specifically PIN2 through CME. Such a tight regulatory control with high specificity to PIN2 could be essential in modulating its polarity.
Publishing Year
Date Published
2019-02-04
Page
138
IST-REx-ID

Cite this

Narasimhan M. Clathrin-Mediated Endocytosis, Post-Endocytic Trafficking and Their Regulatory Controls in Plants . IST Austria; 2019. doi:10.15479/at:ista:th1075
Narasimhan, M. (2019). Clathrin-Mediated endocytosis, post-endocytic trafficking and their regulatory controls in plants . IST Austria. https://doi.org/10.15479/at:ista:th1075
Narasimhan, Madhumitha. Clathrin-Mediated Endocytosis, Post-Endocytic Trafficking and Their Regulatory Controls in Plants . IST Austria, 2019. https://doi.org/10.15479/at:ista:th1075.
M. Narasimhan, Clathrin-Mediated endocytosis, post-endocytic trafficking and their regulatory controls in plants . IST Austria, 2019.
Narasimhan M. 2019. Clathrin-Mediated endocytosis, post-endocytic trafficking and their regulatory controls in plants , IST Austria, 138p.
Narasimhan, Madhumitha. Clathrin-Mediated Endocytosis, Post-Endocytic Trafficking and Their Regulatory Controls in Plants . IST Austria, 2019, doi:10.15479/at:ista:th1075.
All files available under the following license(s):
Creative Commons Attribution 4.0 International Public License (CC-BY 4.0):
Main File(s)
File Name
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
c958f27dd752712886e7e2638b847a3c
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
8786fdc29c62987c0aad3c866a4d3691
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
25f784c5159d6f4d966b2f9b371ebaf6
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
917069272a7a08d1f38224d5e12765d6
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
81e74f5ca0ad70050504f18192236dc0
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
47eb37b27a2930252713924307ea8c6f
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
f68f66721041ce84e331959c9a5779c3
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
67c01cefab51b363c5e214fe4cd671f3
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
e5a397edbee05b8821e2b19b3c1a9260
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
32d92b2a9277f956fdb0b42351d07c0b
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
efe7001f5d9a8c61e631e12d5f324ade
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
eeb0a5603c6449c5f34eacd5ff0b3a16
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
8e7c00ef6223bf0e177deb168338af13
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
3636006a7cb709a7543d6581e359b28d
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
39ca5519a6e9a38356e7b3704004fea7
File Name
Access Level
OA Open Access
Date Uploaded
2019-04-09
MD5 Checksum
4fcdaa3a6c645514a3b3205f0f69dc76

Source File
Access Level
Restricted Closed Access
Date Uploaded
2019-04-09
MD5 Checksum
268f0b6bad21d5f0d671e5d4b88104a7

Export

Marked Publications

Open Data IST Research Explorer

Search this title in

Google Scholar