Journal: bioRxiv

Article Title: Tubulin tyrosination/detyrosination regulates the sorting of intraflagellar transport trains on axonemal microtubule doublets

doi: 10.1101/2024.05.03.592312

Figure Lengend Snippet: A) Illustration of the reconstitution setup. Chlamydomonas cells are overlaid on synthetically polymerized microtubules adhered to a coverslip in a droplet of motility buffer on a TIRF microscope. A detergent back-filled capillary micropipette is immersed in the buffer from the top and centred using a three-axis micromanipulator. B) Legend showing components of IFT trains ( Top ), and illustrative representations of an anterograde ( Middle ) and retrograde train ( Bottom ). C) Illustration of relative positions of microtubules, cells, and capillary pipette. D) Illustration of in vivo IFT train motility ( Left ) and ex vivo IFT train motility on polarity-marked microtubules ( Right ). ⍰ Detergent shot from capillary pipette solubilizes ciliary membrane. ⍰ IFT trains are released into the surrounding environment. Some motile trains continue to move (black arrow) on the demembranated cilia (parent axoneme), while ⍰ others land and move (red arrows) on the polarity-marked microtubules (see also Video S3 ).

Article Snippet: Solution ejection parameters (ejection pressure, time pulse) were controlled by a micropipette pump (PV-850, World Precision Instruments, UK).

Techniques: Microscopy, Transferring, In Vivo, Ex Vivo, Membrane

Journal: bioRxiv

Article Title: Tubulin tyrosination/detyrosination regulates the sorting of intraflagellar transport trains on axonemal microtubule doublets

doi: 10.1101/2024.05.03.592312

Figure Lengend Snippet: ( A ) Representative transilluminated still of a field of view showing relative positions of the Chlamydomonas cells (Yellow), microtubules (Cyan), and overhanging capillary micropipette (blue). Red X marks the spot on the coverslip directly under the needle. ( B ) Time series showing diffusion of liquid (dilute Rhodamine) ejected from capillary pipette over time. The force of ejection does not displace microtubules in the neighbourhood. Relative position and centre of capillary pipette as shown. ‘Area of effect’ is a circle of radius 15 μm with X as the centre. See also Video S1 . ( C ) Kymograph (white diagonal line in (B)) and two-dimension line intensity plots illustrating kinetics of diffusion of ejected liquid. The concentration of the ejected liquid falls exponentially in space and time from the centre of the needle within the area of effect (red square bracket). ( D ) Montage and kymograph of rapid ciliary demembranation (white arrowheads). Cilia proximal to the centre of the capillary (red X) demembranates first followed by the distal cilia. Cell body traced in yellow for reference. See also Video S2 . ( E ) Single box plot of sample of the speed of demembranation of cilia as shown in (D).

Article Snippet: Solution ejection parameters (ejection pressure, time pulse) were controlled by a micropipette pump (PV-850, World Precision Instruments, UK).

Techniques: Diffusion-based Assay, Transferring, Concentration Assay

Journal: STAR Protocols

Article Title: Targeting endogenous proteins for spatial and temporal knockdown using auxin-inducible degron in Caenorhabditis elegans

doi: 10.1016/j.xpro.2022.102028

Figure Lengend Snippet:

Article Snippet: Microinjector , Carl Zeiss , Axiovery 200M.

Techniques: Virus, Recombinant, Gel Extraction, Plasmid Preparation, Purification, CRISPR, Software, Selection, Dissection, Microscopy, Fluorescence