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doi: 10.1242/10.1242/dev.00359

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Quiescent center formation in maize roots is associated with an auxin-regulated oxidizing environment

Department of Plant and Microbial Biology, 111 Koshland Hall, University of California, Berkeley, CA 94720, USA

View larger version (115K): [in a new window]   Fig. 1. Views of maize root quiescent center (QC). (A) Autoradiograph of a median longitudinal section (MLS) of a maize root supplied with [3H]thymidine for 8 hours. Note prominent QC and root cap (RC). (B) MLS showing QC and RC detached together from root proper. (C) Whole maize root with QC and RC dissected free of the root proper. (D) MLS showing detached QC; the RC has already been excised. The extent of the proximal meristem (PM) is also shown. Scale bar, 100 µm.

View larger version (109K): [in a new window]   Fig. 2. Localization of auxin and cell division activity in the maize root apex. Scale bar, 100 µm. (A-D) Control roots, intact, no treatments, stained for (A) histology (Orange G and Safranin), (B) BrdU and (C) auxin. Note that the auxin maximum (dark purple color) localizes in the columella initials/QC. (D) Control, as in C, but without incubation with the primary antibody. QC, quiescent center; PM, proximal meristem; RC, root cap. (E-P) Roots treated with NPA for (E-H) 24 hours; (I-L) 48 hours; (M-P) 24 hours NPA followed by no NPA for 24 hours and stained for auxin (E,I,M) and BrdU (F,G,J,K,N,O). G,K,O are enlargements of F,J,N respectively. H,L and P show the histology. rcj, root cap junction. (Q-V) Roots from which the root cap has been excised (Q,R) 24 hours, (S,T) 48 hours and (U,V) 72 hours earlier. Arrow in S points to heavily stained, auxin-rich region of the developing columella. (Q,S,U) auxin; (R,T,V) BrdU.

View larger version (77K): [in a new window]   Fig. 3. Whole mounts of excised quiescent centers (QC) with or without the adjacent proximal meristem (PM); from roots pre-treated for 2-3 hours with the redox sensitive dye carboxy H2DCFDA. Fluorescence indicates a relatively oxidizing environment. (A,B) An isolated QC with its distal face showing. Photographed in white light (A) or in UV light (B). (C,D) QC and adjacent PM. Photographed simultaneously with white and UV light (C), or in UV light only (D). Note the metaxylem initial (xi arrow) on the distal face of the PM. Scale bar: 100 µm.

View larger version (64K): [in a new window]   Fig. 4. Whole mounts of excised quiescent centers (QC), with and without the adjacent proximal meristem (PM), from roots pre-treated for 2-3 hours with the redox sensitive dye carboxy H2DCFDA. C-J are from roots exposed to NPA (for 24 or 48 hours), and then either treated with the dye, or allowed to grow for an additional amount of time, with no NPA, and then treated with the dye. (A,B) Control, no NPA. Photographed in white light only (A) or in white +UV (B). (C,D) QC at the end of the 24-hour NPA treatment. Photographed in white light (C) or in UV (D). The extent of fluorescence has decreased relative to the control (B). (E-H) QCs and a PM from roots allowed to grow for 24 hours after the end of the NPA treatment. Photographed in white light (E,G), or in UV (F,H). At this time QCs show variable fluorescence. The PMs essentially show no fluorescence (F). G and H are from a QC showing essentially no fluorescence. (I,J) QC and adjacent PM from roots at the end of a 48-hour NPA treatment. Photographed in white (I) or in UV light (J). Note that the PM region is now highly fluorescent, with no fluorescence in the QC. Scale bar, 100 µm.

View larger version (91K): [in a new window]   Fig. 5. Whole mounts of excised quiescent centers (QC) from roots that have had their root cap (RC) excised, and subsequent regeneration allowed to proceed for various periods of time. During the last 2-3 hours of regeneration roots are treated with the redox sensitive dye carboxy H2DCFDA. (A,B) Control, time zero; RC excised just prior to placing roots in dye. Photographed in white light (A) or white plus UV (B). (C,D) 24 hours after removal of the RC. Photographed in white light (C) or UV (D). Note the disappearance of fluorescence from the QC, indicating a less oxidizing environment. (E,F) 48 hours after removal of the RC. Photographed in white light (E) or UV (F). A low level of fluorescence is detected in the center of the tissue. (G,H) 72 hours after removal of the RC. Photographed in white light (G) or UV light (H). The fluorescence has expanded to include an area similar to that of the control (Fig. 7B). Scale bar, 100 µm.

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