Figure 1.
Analysis of the fluorescence intensity resulting from in situ hybridisation of poly(A) RNA in the meristematic cells of lupines roots.
There were significant differences in intensity (p<0.05) between the cytoplasm (MC), nucleus (MN) and nuclear bodies (MNB).
Figure 2.
Double labelling of poly(A) RNA and the PANA antigen in the chromocentric (Lupinus) (A–C) and reticular (Allium) (D–F) nuclei of root cells.
In the nucleoplasm of both species, poly(A) RNA is present in nuclear structures (arrows) and does not colocalise with speckles. Representative examples of Pearson correlation coefficients for weak and non-colocalisation of poly(A) mRNA with the PANA antigen in Lupinus luteus and Allium cepa cells (G). A scale bar representing 2 µm is shown. The percentages of weak and non-colocalisation of poly(A) RNA-rich bodies with the PANA antigen are indicated by the Pearson correlation coefficient (H). Error bars represent standard error. Double labelling of poly(A) RNA and U2 snRNA in Lupinus (I–K) and Allium (L–N) cells. Accumulation of poly(A) RNA in nuclear bodies rich in U2 snRNA (arrows). Bar, 5 µm. N-nucleus, Nu- nucleolus
Figure 3.
Double labelling of poly(A) RNA and the U2B” protein in Lupinus (A–C) and Allium (D–F) root cells.
Poly(A) RNA present within Cajal bodies stained an anti-U2B” antibody (arrows). Immunogold labelling of poly(A) RNA in the Cajal bodies (CBs) of Lupinus root cells. Gold particles were mainly observed in the Cajal body (CB) (G). Simultaneous localisation of U2B”:GFP and poly(A) RNA in transgenic Arabidopsis thaliana root cells. Strong colocalisation in Cajal bodies (H–J). Localisation of poly(A) RNA with Atcoilin:RFP in Arabidopsis thaliana root cells. Accumulation of poly(A) in Cajal bodies (K–M). Bar, 5 µm. N-nucleus, Nu- nucleolus, C- cytoplasm.
Figure 4.
Double labelling of a mixture of four mRNAs (A, C) and distinct genes: cyclin B1 mRNA (D, F), peroxidase mRNA (G, I), cytokinin-specific binding protein mRNA (J, L) and pectin methylesterase mRNA (M, O), with Sm proteins (B, E, H, K, N) in Lupinus cells.
The arrows indicate colocalisation of mRNA transcripts with CBs. A stronger signal was observed in the cytoplasm than in the nuclei after in situ hybridisation with the mixture of probes (A, C) and with a probe against cytokinin-specific binding protein mRNA (J, L). % indicates the percentage of nuclei showing the representative immunolocalisation pattern. Bar, 5 µm. N-nucleus, Nu- nucleolus, C- cytoplasm.
Figure 5.
Double labelling of poly(A) RNA (A) and the elongation form of RNA polymerase II (B) in Lupinu root cells.
Cajal body (arrow) rich in poly(A) RNA near the nucleolus do not colocalise with RNA polymerase II. Double labelling of a newly formed transcript (E, H) and poly(A) RNA (D, G) in lupine cells. If the transcript does not leave the nucleus, no signal occurs in the CB (arrow) (D–F). In cells in which the newly formed RNA is transported to the cytoplasm, a weak signal is observed in the Cajal body (arrow) (G–I). Bar, 5 µm. Fragment of the nucleus from Fig 2I (J). BrU-containing RNA localises at the periphery and in small spots in the middle of CB (J). Bar, 1 µm. The percentage of Cajal bodies rich in poly(A) RNA that colocalise with BrU-containing RNA (K, L). The data for experimental treatments and control were conducted, analyzed and plotted as in Figure 2G–H. A scale bar representing 2 µm is shown. Error bars represent standard error. N- nucleus, Nu- nucleolus, C- cytoplasm.
Figure 6.
Localisation of poly(A) RNA in control lupine root cells (A) and after submersion to tap water for 3 h (B).
Arrows indicate CBs, and arrow heads indicate cytoplasmic granules. Bar, 5 µm. Nu- nucleolus. Quantitative analysis of the fluorescence intensity associated with the localisation of poly(A) RNA. Significant differences in the signal intensity in the nuclei and CBs between control and hypoxia-treated cells (C) MNu- control nucleus, MCB control Cajal bodies, HypNu and HypCB- nucleus and Cajal bodies after hypoxia treatment, respectively (p = 0.05).