Fig 2: Lignification of S. dulcamara, Tomato cv. Marmande and S. tuberosum cv. Desirée tissues upon R. solanacearum infection. (A) Representative composed images of S. dulcamara, tomato, and potato taproot transversal sections obtained 9 days after inoculation with R. solanacearum-GFP or mock treatment. First and second row: microscope bright-field images after lignin staining with phloroglucinol HCl (magenta colouration). Third row: fluorescence microscopy images after inoculation to assess the extent of bacterial colonisation. Images were obtained using a Leica DM6 microscope. Scale bars indicate 0.5 mm. (B) Quantification of the phloroglucinol HCL stain -indicative of lignin content- in the vascular area from the images shown in A performed with the ImageJ software. ∗indicates statistical differences (p-value < 0.05; T-student significant test α = 0.05).

Fig 3: nse-4 mutants showed defective inter-sister repair. (A) Micrographs of the diakinesis chromosome with fragments (white arrows) in the double mutants. (B) Representative micrographs showing chromosome structure at diakinesis in nse-4(tm7158);syp-2(ok307). Chromosome bodies are grossly damaged, but surprisingly, some nuclei showed wild-type phenotype. (C) Quantification of chromosome fragments at diakinesis (** p < 0.01, **** p < 0.0001, chi-square Fisher’s Exact Test). The sample size (n) = 200 for each genotype. Chromosome fragments were scored using Leica DM6 B at 100× objective with oil immersion. (D) Quantification of DAPI-stained bodies of nse-4(tm7141);syp-2(ok307) diakinesis nuclei. Nse-4(tm7141) mutation rescued about 15% of the syp-2 diakinesis phenotype. The sample size (n) = 200 for each genotype. DAPI bodies were scored using Leica DM6 B at 100× objective with oil immersion. Images were captured using Zeiss confocal microscope LSM 800 with Airyscan at 63× objective with oil immersion (scale bar = 10 μm).

Fig 4: (A) Expression of SLC22A8 (OAT3) and it´s allelic variants in HEK293T cells line. Wild type protein was tagged with C-terminal GFP tag and cells was transiently transfected by polyethyleneimine lipofection. Samples was fixed by 4% paraformaldehyde and picture was take by Leica DM6 microscope in 400× magnification. Plasmatic membrane localization was observed in all variants. Cell nucleus was stain by DAPI (blue). Higher magnification of allelic variant R149C of transporter OAT3 C-terminus tagged with GFP. Protein was expressed in HEK293 cell line. Cell nucleus was stain with DAPI. I. Fluorescence signal of GFP tagged protein. II. Cytoplasmatic membrane staining by PKH26 stain. (B) Immunoblotting of HEK293T transfected by OAT3 WT and its allelic variant. Fusion protein OAT3-mGFP had majority band with molecular weight 90 kDa. CapZ (β-actinin) protein is used as loading control and it had one major band with 42 kDa molecular weight. (C) Transport of 14C radiolabeled uric acid by HEK293 cells transfected by OAT3 WT and its allelic variants. The data are shown as pmol uric acid transported into the cytoplasm per one minute and per total protein in cell lysate. Data are expressed with ±SD, n = 2. Statistical analyses for significant differnces was calculate by Students t-test (*, p < 0.05). (D) Topological model of human OAT3 protein (encoded by gene SLC22A8). Allelic variants p.R149C, p.V448I and p.R513Q (single amino acid exchange) are colored by pink color. Putative glycosylation motifs are colored by green (for OAT3 is no experimental evidence for glycosylation). Edited on the base [19].

Fig 5: Glial humanization of the mouse brain. (A) Human lymphoid and brain progenitor cells were transplanted to mice at birth. Blue arrow indicates injection of neural progenitor cells (NPC) into the lateral ventricle (LV). Red arrow indicates intrahepatic (ih) injection of hematopoietic stem/progenitor cells (HSPC). (B) Representative view under a 1× objective lens from a 5-μm paraffin-embedded sagittal section stained for human-specific GFAP (brown). Asterisks indicate fields where cell images were captured for display (C) and quantified (E). (C) Staining with human astrocyte-specific antibodies (hu-GFAP, red) and total glial fibrillary acidic protein (GFAP; green) was performed. Representative images from different brain regions are shown. Original images were collected by a Leica DM6 system at 1000× magnification. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI; blue). Scale bar: 20 μm. (D) Oligodendrocyte-specific nuclear staining (OLIG2; green) and total human-specific nuclear (hN, red) staining was performed, and representative images are shown. The original magnification was 400×. Scale bar: 50 μm. The dashed line in D demarcates the CC from the FC. (E,F) Quantitative data showing the number of human glial cells from one of four experiments with 10 animals. The average percentage of human cells was assessed on two to four representative sagittal sections from each mouse for a selected field of view of the same brain region under 400× magnification. Distribution and proportion of hu-GFAP+ astrocytes among the total number of GFAP+ cells (E); distribution and proportion of hN+ cells positive for OLIG2 (F). Each symbol represents an individual mouse. Data are mean± s.e.m. *P<0.05, **P<0.01, ***P<0.001 by Kruskal–Wallis test. Animals were analyzed at 6 months of age and reconstitution characteristics are shown in Table 1.