Fig. 5: CURT1 depletion causes cell division defects in Chlamydomonas. | Nature Communications

Fig. 5: CURT1 depletion causes cell division defects in Chlamydomonas.

From: CurT/CURT1 proteins are involved in cell and chloroplast division coordination of cyanobacteria and green algae

Fig. 5

a Schematic maps of mutant CrCURT1 alleles with exons being indicated as black boxes. The maps are drawn to scale in units of base pairs (bp, see scale bar). Primer-binding sites for reverse-transcriptase PCR (RT-PCR) are indicated as half-arrows. Aph7, hygromycin resistance gene. b RT-PCR indicates absence of CrCURT1A, CrCURT1B, and CrCURT1C transcripts in the triple mutant. The cDNA for G-protein β-subunit-like protein (GBLP) was employed as internal control85. Sizes of respective amplification products in base pairs (bp) are indicated. Sampling of mRNA through one cycle in light/dark culture and subsequent RT-PCR was done twice independently (n = 2), yielding similar results. c Chlamydomonas curt1abc mutants display increased average duplication time (p = 3.9 × 10−3; two-sided Student’s t-test). Error bars indicate standard deviations of n = 4 independent biological replicates. d Transmission (top) and confocal laser scanning micrographs (bottom) of Chlamydomonas WT and curt1abc mutant cells cultivated in liquid TAP media. Magenta represents chlorophyll fluorescence (Chl FL). The curt1abc mutant displays variable size in vegetative cells (left) and atypical cell division products (right). The experiment was performed twice with similar results. e Cell-size distributions of non-dividing Chlamydomonas WT (n = 80) and curt1abc mutants (n = 86) shown in (d). Cell size was recorded as cell-body length excluding flagella using Fiji MicrobeJ. f Cell-size distributions of Chlamydomonas WT and curt1abc mutants cultivated in liquid TAP media as recorded by flow cytometry. Apparent particle size (x axis) was estimated from forward scatter (FSC) using a size calibration kit (F13838, Invitrogen). Red fluorescence (FL) intensity (primary y axis, arbitrary (arb.) units; excitation 532 nm, emission 680 ± 15 nm) is shown as detector output values without absolute calibration. Dot plots are colored by point density (secondary y axis, arb. units relative to maximum density). Dashed lines mark average particle sizes (dot) ± standard deviation (whiskers); *** indicates statistically significant difference (p = 8.4 × 10-84; two-sided Student’s t-test; n = 4977 cells each). The experiment was performed twice with similar results.

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