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Roxane Spiegelhalder, PhD

@roxyspi.bsky.social

474 followers 198 following 8 posts

plant scientist turned human geneticist🧬🖥 Human Rights | Feminism | Climate Justice she/her

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

Finally, we saw that the dose-dependent rescue of GC and SC formation in sid/bdmute-1 by 3xGFP-BdMUTE was mirrored in a gradual complementation of stomatal physiology - linking once more SC presence and GC morphology to the improved kinetics of graminoid stomata. (7/8)

gSW of WT, the four sid/bdmute-1;BdMUTEp:3xGFP-BdMUTE (M3GM) lines (++, +, - and - -) and sid/bdmute-1 under changing light conditions sowing a gradual rescue of the sid phenotype.

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

In intermediate lines, we saw stomata with 0, 1 or 2 SCs. In complexes with only 1 SC,GCs that lacked a SC did not form proper dumbbell shape. The GC flanked by the SCs, however, formed a better dumbbell. Thus, SC presence influenced GC morphogenesis independent of BdMUTE. (6/8)

3D confocal stacks of mature stomatal complexes of the M3GM- line without SCs (E), with one SC (F) and with two SCs (G). XZ Cross sections through the apices and middle and YZ sections are displayed. visible is a larger circumference and more cell wall staining in GC cross sections of cells not flanked by SCs

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

In weak lines, SCs were not recruited but GC abortion fully rescued. This seemed to correlate w/ rescue of skewed divisions in dev. GCs w/o requiring SC recruitment, suggesting a cell-autonomous role in division plane orientation in GMCs that does not require SC recruitment (5/8)

plot of total “skewness” ratios per GMC as a sum of apical and basal “skewness” ratios. A total ratio < 1.6 (dashed horizontal line) represents a skewed division (indicated as red dots). n = 7-8 individuals per genotype and 66-82 stomata per genotype (dots are stomata). Schematic representation of the division phenotypes “Straight” division represented by ratios > 1.6 (i.e. all WT stomata) were used as a WT cutoff. GCs with a division plane orientation < 1.6 and > 0 were classified as “skewed” and a transverse division is shown as ratio = 0.

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

Even though strong lines almost fully rescued sid/bdmute-1, we were able to show that BdMUTE with reduced mobility resulted in severely delayed SC recruitment in the strong lines indicating that BdMUTE mobility is indeed required to recruit SCs. (4/8)

confocal images of 2nd leaf developmental zones in MYM and M3GM++ showing undivided (“U”) and divided (“D”) GMCs and present SCs (arrowhead) or missing SCs (!) are indicated. MYM line shows present SCs flanking all undivided GMCs whereas M3GM++ shows divided GCs that still sometimes lack one SC. Cell walls are stained with PI (magenta), fluorophores shown in yellow (YFP) and green (3xGFP

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

Strikingly, we found a broad range of expression strengths in the mobility impaired BdMUTE complementation lines. The stronger the line was expressed, the better it rescued the mutant sid/bdmute-1 phenotype (3/8)

Bar plot showing quantification of stomatal phenotypes in the mature 3rd leaf of WT, sid/bdmute-1 and sid/bdmute-1 complemented with the different reporters. A gradual complemetation of both phenotypes (GC and SC) can be seen

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

We used a set of sid/bdmute-1 complementation lines with functional, mobile BdMUTE (single YFP-tagged) and BdMUTE with impaired mobility (triple GFP-tagged) to untangle roles in the two independent cell lineages (2/8)

Representative confocal images of stage 3 leaf zones of the lines MYM, M3GM++, M3GM+, M3GM-, M3GM- - and MMY with signal intensity of reporter proteins shown as “fire” heatmap and propidium iodide (PI)-stained cell walls shown in gray. MUTE in GMCs shows different intensities in the four GFP lines, mobile signal in the lateral cell files is only visible in the single-tag YFP lines.

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Roxane Spiegelhalder, PhD @roxyspi.bsky.social · 06/05/2024

Excited to share our new #preprint on BdMUTE and its dual role in recruiting subsidiary cells (SCs) and orienting guard cell (GC) division in #Brachypodium: biorxiv.org/content/10.1... We set out to answer questions about how BdMUTE influences grass stomatal development: (1/8)

Schematic displaying stomatal development in wild type (WT). Stomatal development starts with guard mother cell (GMC; blue) formation. BdMUTE is expressed in the GMC (light blue nucleus) and moves into the lateral cells (light yellow nucleus) to establish subsidiary mother cells (SMCs). SMCs then polarize (yellow gradient) and divide asymmetrically to form lateral SCs (yellow). Finally, the GMC divides symmetrically to generate two GCs (light green), which form a pore and mature into the characteristic dumbbell shape (dark green). Open questions regarding BdMUTE’s roles and mechanisms are indicated.

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