Structure-guided Mutagenesis of OSCAs Reveals Mechanosensitive Activation

Temel Kavramlar

The author explores the structural elements contributing to mechanosensitivity in OSCA channels, highlighting the role of the amphipathic helix and lipid-interacting residues in response to mechanical stimuli.

Özet

The study investigates the mechanosensitivity of OSCA/TMEM63 ion channels through cryo-EM analysis and mutagenesis experiments. The research focuses on the involvement of specific structural elements, such as the amphipathic helix and lipid-interacting residues, in channel activation by mechanical stimuli. Results indicate a selective disruption of poke-activated currents without affecting stretch-activated currents, suggesting functional diversity within the OSCA family.
The dimeric two-pore OSCA/TMEM63 family has been identified as mechanically activated ion channels. The study aims to understand how different domains contribute to mechanical signal transduction in these channels. By using cryo-electron microscopy, the authors solved the structure of Arabidopsis thaliana (At) OSCA3.1 and performed mutagenesis studies on conserved mechanosensitive features of OSCA1.2.
Results show that mutations in key residues within the amphipathic helix selectively disrupt poke-induced currents while leaving stretch-activated currents unaffected. Additionally, replacing potential lipid-interacting lysine residues with isoleucine abrogates poke responses but not stretch responses in OSCA1.2.
Overall, this research sheds light on the molecular mechanisms underlying mechanosensitivity in OSCAs and highlights specific structural elements crucial for channel activation by mechanical stimuli.

İstatistikler

The EGFP was removed by PreScission Protease cleavage, leaving a stretch of ten residues on the C-terminus.
Purified protein was reconstituted into nanodiscs and subjected to cryo-EM analysis at 2.6 Å resolution.
A total dose of ∼50 electrons per Å2 was used during data collection.
Cryo-EM maps have been deposited to EMDB under accession number EMD-41911.
The final model comprises residues 2-102, 154-231, 307-388, 410-476, and 489-697 out of 724 residues in OSCA3.1 sequence.

Alıntılar

"The loss of sensitivity of cells expressing mutant constructs to poking suggests that the amphipathic helix plays an important role in mechanically activated responses."
"Substituting potential lipid-interacting lysine residues for isoleucine selectively impairs poke-induced responses without affecting stretch responsiveness."
"The results suggest that changes in specific features could fine-tune the response to certain mechanical stimuli within the large family."

Önemli Bilgiler Şuradan Elde Edildi

Structure-guided mutagenesis of OSCAs reveals differential activation to mechanical stimuli

by Jojoa Cruz,S... : www.biorxiv.org 10-03-2023

https://www.biorxiv.org/content/10.1101/2023.10.03.560740v2

Daha Derin Sorular

How do differences in membrane composition between plant cells and HEK293 cells impact channel activation by mechanical stimuli?

The differences in membrane composition between plant cells and HEK293 cells can have a significant impact on the activation of channels by mechanical stimuli. Plant cell membranes typically contain a different lipid composition compared to mammalian cell membranes like those of HEK293 cells. These variations in lipid content can influence how ion channels, such as OSCA/TMEM63, respond to mechanical stimuli.
For instance, lipids play a crucial role in stabilizing and modulating the function of ion channels. In TRAAK channels, occlusion of the pore by a lipid acyl chain has been proposed as a gating mechanism. Similarly, in SWELL1 channels, lipids block the pore in the closed state. The interaction between specific residues on TM4 and TM6b with lipid phosphate head groups may contribute to channel activation during poking stimulus.
Given these insights into how lipids interact with ion channels to regulate their function, it is plausible that variations in membrane composition could alter the sensitivity or response characteristics of mechanosensitive ion channels like OSCA/TMEM63 when comparing plant cell membranes to those of HEK293 cells.

How might understanding mechanosensitivity at a molecular level contribute to advancements in biophysics or medical research?

Understanding mechanosensitivity at a molecular level holds great promise for contributing to advancements in both biophysics and medical research. Here are some ways this knowledge could be beneficial:

Biophysics Advancements:

Structural Insights: Detailed structural studies provide valuable information about how mechanically activated ion channels like OSCAs are organized at atomic levels.
Mechanistic Understanding: By elucidating the mechanisms through which these ion channels sense and transduce mechanical forces, researchers can gain deeper insights into fundamental biological processes related to mechanotransduction.
Functional Diversity: Identifying key structural elements involved in mechanosensation helps uncover sources of functional diversity within protein families like OSCA/TMEM63.

Medical Research Implications:

Drug Development: Targeting mechanosensitive ion channels implicated in various diseases could lead to novel therapeutic strategies.
Disease Mechanisms: Understanding how mutations or dysregulation affect channel function under mechanical stress may shed light on disease pathogenesis linked to impaired mechanotransduction pathways.
Biomedical Engineering: Insights into how proteins respond mechanically can inform the design of biomimetic materials or devices for biomedical applications.

Overall, unraveling the molecular basis of mechanosensitivity not only enhances our understanding of basic biological processes but also opens up avenues for innovative approaches towards addressing health-related challenges through biophysical and medical research endeavors.