Targeting the cell membrane in established and emerging model organisms.

Area of Science:

• Cell Biology
• Developmental Biology
• Molecular Biology

Background:

• Transgenic tools are crucial for studying cells and organisms, but require species-specific components like cis-regulatory elements.
• Existing membrane-localizing signals show variable efficacy across different species, limiting their universal application.
• A need exists for broadly applicable tools to target proteins to the plasma membrane in diverse biological systems.

Purpose of the Study:

• To create and validate a toolkit of membrane-localizing tags for rapid screening in diverse organisms.
• To identify robust membrane-targeting signals applicable across a wide range of species and phyla.
• To establish universally effective membrane-localizing tags for enhanced transgenic research.

Main Methods:

• Generation of a toolkit comprising 11 distinct membrane-localizing tags, each fused to the mScarlet3 fluorescent protein.
• Utilizing a T7 promoter for mRNA production, enabling delivery into various embryos and cells.
• Collaborative testing of the toolkit across ten animal species representing diverse phyla and in choanoflagellates.

Main Results:

• Identification of robust membrane-localizing tags in all tested animal species and choanoflagellates.
• Demonstration that three specific tags (KRas, GAP43, and Src64B) exhibit functionality across all tested species.
• Validation of diverse targeting mechanisms, including signal peptides, lipid attachments, and lipid-binding domains.

Conclusions:

• The developed toolkit provides a valuable resource for rapidly screening membrane-localizing tags in diverse organisms.
• The identification of universally effective tags (KRas, GAP43, Src64B) significantly advances the development of cross-species transgenic tools.
• This work facilitates more efficient and standardized cellular and developmental studies across a broad spectrum of life.